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Sample records for functional total rna

  1. A Rapid and Efficient Method for Purifying High Quality Total RNA from Peaches (Prunus persica for Functional Genomics Analyses

    Directory of Open Access Journals (Sweden)

    LEE MEISEL

    2005-01-01

    Full Text Available Prunus persica has been proposed as a genomic model for deciduous trees and the Rosaceae family. Optimized protocols for RNA isolation are necessary to further advance studies in this model species such that functional genomics analyses may be performed. Here we present an optimized protocol to rapidly and efficiently purify high quality total RNA from peach fruits (Prunus persica. Isolating high-quality RNA from fruit tissue is often difficult due to large quantities of polysaccharides and polyphenolic compounds that accumulate in this tissue and co-purify with the RNA. Here we demonstrate that a modified version of the method used to isolate RNA from pine trees and the woody plant Cinnamomun tenuipilum is ideal for isolating high quality RNA from the fruits of Prunus persica. This RNA may be used for many functional genomic based experiments such as RT-PCR and the construction of large-insert cDNA libraries.

  2. Monocytic microRNA profile associated with coronary collateral artery function in chronic total occlusion patients.

    Science.gov (United States)

    Hakimzadeh, Nazanin; Elias, Joëlle; Wijntjens, Gilbert W M; Theunissen, Ruud; van Weert, Angela; Smulders, Martijn W; van den Akker, Nynke; Moerland, Perry D; Verberne, Hein J; Hoebers, Loes P; Henriques, Jose P S; van der Laan, Anja M; Ilhan, Mustafa; Post, Mark; Bekkers, Sebastiaan C A M; Piek, Jan J

    2017-05-08

    An expansive collateral artery network is correlated with improved survival in case of adverse cardiac episodes. We aimed to identify cellular microRNAs (miRNA; miR) important for collateral artery growth. Chronic total occlusion (CTO) patients (n = 26) were dichotomized using pressure-derived collateral flow index (CFI p ) measurements; high collateral capacity (CFI p  > 0.39; n = 14) and low collateral (CFI p  collateral capacity patients. Validation by real-time polymerase chain reaction demonstrated significantly decreased expression of miR339-5p in all stimulated monocyte phenotypes of low collateral capacity patients. MiR339-5p showed significant correlation with CFI p values in stimulated monocytes. Ingenuity pathway analysis of predicted gene targets of miR339-5p and differential gene expression data from high versus low CFI p patients (n = 20), revealed significant association with STAT3 pathway, and also suggested a possible regulatory role for this signaling pathway. These results identify a novel association between miR339-5p and coronary collateral function. Future work examining modulation of miR339-5p and downstream effects on the STAT3 pathway and subsequent collateral vessel growth are warranted.

  3. Phylogenetic and Functional Diversity of Total (DNA) and Expressed (RNA) Bacterial Communities in Urban Green Infrastructure Bioswale Soils

    Science.gov (United States)

    Lee, Angela; McGuire, Krista L.

    2017-01-01

    ABSTRACT New York City (NYC) is pioneering green infrastructure with the use of bioswales and other engineered soil-based habitats to provide stormwater infiltration and other ecosystem functions. In addition to avoiding the environmental and financial costs of expanding traditional built infrastructure, green infrastructure is thought to generate cobenefits in the form of diverse ecological processes performed by its plant and microbial communities. Yet, although plant communities in these habitats are closely managed, we lack basic knowledge about how engineered ecosystems impact the distribution and functioning of soil bacteria. We sequenced amplicons of the 16S ribosomal subunit, as well as seven genes associated with functional pathways, generated from both total (DNA-based) and expressed (RNA) soil communities in the Bronx, NYC, NY, in order to test whether bioswale soils host characteristic bacterial communities with evidence for enriched microbial functioning, compared to nonengineered soils in park lawns and tree pits. Bioswales had distinct, phylogenetically diverse bacterial communities, including taxa associated with nutrient cycling and metabolism of hydrocarbons and other pollutants. Bioswale soils also had a significantly greater diversity of genes involved in several functional pathways, including carbon fixation (cbbL-R [cbbL gene, red-like subunit] and apsA), nitrogen cycling (noxZ and amoA), and contaminant degradation (bphA); conversely, no functional genes were significantly more abundant in nonengineered soils. These results provide preliminary evidence that urban land management can shape the diversity and activity of soil communities, with positive consequences for genetic resources underlying valuable ecological functions, including biogeochemical cycling and degradation of common urban pollutants. IMPORTANCE Management of urban soil biodiversity by favoring taxa associated with decontamination or other microbial metabolic processes is a

  4. Phylogenetic and Functional Diversity of Total (DNA) and Expressed (RNA) Bacterial Communities in Urban Green Infrastructure Bioswale Soils.

    Science.gov (United States)

    Gill, Aman S; Lee, Angela; McGuire, Krista L

    2017-08-15

    New York City (NYC) is pioneering green infrastructure with the use of bioswales and other engineered soil-based habitats to provide stormwater infiltration and other ecosystem functions. In addition to avoiding the environmental and financial costs of expanding traditional built infrastructure, green infrastructure is thought to generate cobenefits in the form of diverse ecological processes performed by its plant and microbial communities. Yet, although plant communities in these habitats are closely managed, we lack basic knowledge about how engineered ecosystems impact the distribution and functioning of soil bacteria. We sequenced amplicons of the 16S ribosomal subunit, as well as seven genes associated with functional pathways, generated from both total (DNA-based) and expressed (RNA) soil communities in the Bronx, NYC, NY, in order to test whether bioswale soils host characteristic bacterial communities with evidence for enriched microbial functioning, compared to nonengineered soils in park lawns and tree pits. Bioswales had distinct, phylogenetically diverse bacterial communities, including taxa associated with nutrient cycling and metabolism of hydrocarbons and other pollutants. Bioswale soils also had a significantly greater diversity of genes involved in several functional pathways, including carbon fixation ( cbbL-R [ cbbL gene, red-like subunit] and apsA ), nitrogen cycling ( noxZ and amoA ), and contaminant degradation ( bphA ); conversely, no functional genes were significantly more abundant in nonengineered soils. These results provide preliminary evidence that urban land management can shape the diversity and activity of soil communities, with positive consequences for genetic resources underlying valuable ecological functions, including biogeochemical cycling and degradation of common urban pollutants. IMPORTANCE Management of urban soil biodiversity by favoring taxa associated with decontamination or other microbial metabolic processes is a

  5. Human Milk MicroRNA and Total RNA Differ Depending on Milk Fractionation.

    Science.gov (United States)

    Alsaweed, Mohammed; Hepworth, Anna R; Lefèvre, Christophe; Hartmann, Peter E; Geddes, Donna T; Hassiotou, Foteini

    2015-10-01

    MicroRNA have been recently discovered in human milk signifying potentially important functions for both the lactating breast and the infant. Whilst human milk microRNA have started to be explored, little data exist on the evaluation of sample processing, and analysis to ensure that a full spectrum of microRNA can be obtained. Human milk comprises three main fractions: cells, skim milk, and lipids. Typically, the skim milk fraction has been measured in isolation despite evidence that the lipid fraction may contain more microRNA. This study aimed to standardize isolation of microRNA and total RNA from all three fractions of human milk to determine the most appropriate sampling and analysis procedure for future studies. Three different methods from eight commercially available kits were tested for their efficacy in extracting total RNA and microRNA from the lipid, skim, and cell fractions of human milk. Each fraction yielded different concentrations of RNA and microRNA, with the highest quantities found in the cell and lipid fractions, and the lowest in skim milk. The column-based phenol-free method was the most efficient extraction method for all three milk fractions. Two microRNAs were expressed and validated in the three milk fractions by qPCR using the three recommended extraction kits for each fraction. High expression levels were identified in the skim and lipid milk factions for these microRNAs. These results suggest that careful consideration of both the human milk sample preparation and extraction protocols should be made prior to embarking upon research in this area. © 2015 The Authors. Journal of Cellular Biochemistry Published by Wiley Periodicals, Inc.

  6. TruSeq Stranded mRNA and Total RNA Sample Preparation Kits

    Science.gov (United States)

    Total RNA-Seq enabled by ribosomal RNA (rRNA) reduction is compatible with formalin-fixed paraffin embedded (FFPE) samples, which contain potentially critical biological information. The family of TruSeq Stranded Total RNA sample preparation kits provides a unique combination of unmatched data quality for both mRNA and whole-transcriptome analyses, robust interrogation of both standard and low-quality samples and workflows compatible with a wide range of study designs.

  7. Isolation of Microarray-Grade Total RNA, MicroRNA, and DNA from a Single PAXgene Blood RNA Tube

    DEFF Research Database (Denmark)

    Kruhøffer, Mogens; Andersen, Lars Dyrskjøt; Voss, Thorsten

    2007-01-01

    We have developed a procedure for isolation of microRNA and genomic DNA in addition to total RNA from whole blood stabilized in PAXgene Blood RNA tubes. The procedure is based on automatic extraction on a BioRobot MDx and includes isolation of DNA from a fraction of the stabilized blood......RNA was tested using spotted locked nucleic acid-based microarrays. We conclude that the yield and quality of total RNA, microRNA, and DNA from a single PAXgene blood RNA tube is sufficient for downstream microarray analysis....

  8. Extraction of total RNA in the developing chicken forebrain

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    Sayed Rasoul Zaker

    2014-01-01

    Full Text Available Background: Gene expression of Gama-Aminobutyric acid (GABA A receptor subunits may change during development. Procedures in molecular biology are required to understand the gene expression profile GABA A R in chicken. The outcome of the results depends on good-quality high-molecular-weight RNA. Several procedures can be used to isolate RNA from the brain of chicken; however, most of them are time-consuming and require disruption of cells or freeze and thaw in the presence of RNase inhibitors. The aim of this experiment was isolation of RNA from chicken embryonic brain tissues using appropriate RNA extraction kit. Materials and Methods: Fertilized eggs from Ross breed (Gallus gallus were incubated at 38°C and 60% relative humidity in a forced-draft incubator and were turned every 3 h. After 3, 7, 14 and 20 days of incubation, eggs were cooled on ice to induce deep anesthesia. Then whole brains were dissected out. As brains could not be excised in a reproducible way from earlier embryos (embryonic days 4 and 6, whole heads were collected. Chicken embryos between day 7 to 20 and 1 day after birth were decapitated, and their brains removed. Samples were immediately inserted into lysis buffer and stored at −70°C. Total RNA was isolated and a contaminating genomic deoxyribonucleic acid (DNA was digested. RNA quality was checked using gel electrophoresis. Results: We obtained 52 mg/ml to 745 mg/ml with A260/280 1.7-2.2. Only high-quality RNA, with no signs of degradation, was used for further experiments. Conclusion: In conclusion, protocol was found to be suitable for the isolation of total RNA from embryonic chicken cells.

  9. Human Milk MicroRNA and Total RNA Differ Depending on Milk Fractionation

    OpenAIRE

    Alsaweed, Mohammed; Hepworth, Anna R.; Lef?vre, Christophe; Hartmann, Peter E.; Geddes, Donna T.; Hassiotou, Foteini

    2015-01-01

    ABSTRACT MicroRNA have been recently discovered in human milk signifying potentially important functions for both the lactating breast and the infant. Whilst human milk microRNA have started to be explored, little data exist on the evaluation of sample processing, and analysis to ensure that a full spectrum of microRNA can be obtained. Human milk comprises three main fractions: cells, skim milk, and lipids. Typically, the skim milk fraction has been measured in isolation despite evidence that...

  10. Evaluation of commercially available RNA amplification kits for RNA sequencing using very low input amounts of total RNA.

    Science.gov (United States)

    Shanker, Savita; Paulson, Ariel; Edenberg, Howard J; Peak, Allison; Perera, Anoja; Alekseyev, Yuriy O; Beckloff, Nicholas; Bivens, Nathan J; Donnelly, Robert; Gillaspy, Allison F; Grove, Deborah; Gu, Weikuan; Jafari, Nadereh; Kerley-Hamilton, Joanna S; Lyons, Robert H; Tepper, Clifford; Nicolet, Charles M

    2015-04-01

    This article includes supplemental data. Please visit http://www.fasebj.org to obtain this information.Multiple recent publications on RNA sequencing (RNA-seq) have demonstrated the power of next-generation sequencing technologies in whole-transcriptome analysis. Vendor-specific protocols used for RNA library construction often require at least 100 ng total RNA. However, under certain conditions, much less RNA is available for library construction. In these cases, effective transcriptome profiling requires amplification of subnanogram amounts of RNA. Several commercial RNA amplification kits are available for amplification prior to library construction for next-generation sequencing, but these kits have not been comprehensively field evaluated for accuracy and performance of RNA-seq for picogram amounts of RNA. To address this, 4 types of amplification kits were tested with 3 different concentrations, from 5 ng to 50 pg, of a commercially available RNA. Kits were tested at multiple sites to assess reproducibility and ease of use. The human total reference RNA used was spiked with a control pool of RNA molecules in order to further evaluate quantitative recovery of input material. Additional control data sets were generated from libraries constructed following polyA selection or ribosomal depletion using established kits and protocols. cDNA was collected from the different sites, and libraries were synthesized at a single site using established protocols. Sequencing runs were carried out on the Illumina platform. Numerous metrics were compared among the kits and dilutions used. Overall, no single kit appeared to meet all the challenges of small input material. However, it is encouraging that excellent data can be recovered with even the 50 pg input total RNA.

  11. Structure and Function of Caliciviral RNA Polymerases

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    Ji-Hye Lee

    2017-11-01

    Full Text Available Caliciviruses are a leading agent of human and animal gastroenteritis and respiratory tract infections, which are growing concerns in immunocompromised individuals. However, no vaccines or therapeutics are yet available. Since the rapid rate of genetic evolution of caliciviruses is mainly due to the error-prone nature of RNA-dependent RNA polymerase (RdRp, this article focuses on recent studies of the structures and functions of RdRp from caliciviruses. It also provides recent advances in the interactions of RdRp with virion protein genome-linked (VPg and RNA and the structural and functional features of its precursor.

  12. Retroviral RNA Dimerization: From Structure to Functions

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    Noé Dubois

    2018-03-01

    Full Text Available The genome of the retroviruses is a dimer composed by two homologous copies of genomic RNA (gRNA molecules of positive polarity. The dimerization process allows two gRNA molecules to be non-covalently linked together through intermolecular base-pairing. This step is critical for the viral life cycle and is highly conserved among retroviruses with the exception of spumaretroviruses. Furthermore, packaging of two gRNA copies into viral particles presents an important evolutionary advantage for immune system evasion and drug resistance. Recent studies reported RNA switches models regulating not only gRNA dimerization, but also translation and packaging, and a spatio-temporal characterization of viral gRNA dimerization within cells are now at hand. This review summarizes our current understanding on the structural features of the dimerization signals for a variety of retroviruses (HIVs, MLV, RSV, BLV, MMTV, MPMV…, the mechanisms of RNA dimer formation and functional implications in the retroviral cycle.

  13. Ribosomal RNA gene functioning in avian oogenesis.

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    Koshel, Elena; Galkina, Svetlana; Saifitdinova, Alsu; Dyomin, Alexandr; Deryusheva, Svetlana; Gaginskaya, Elena

    2016-12-01

    Despite long-term exploration into ribosomal RNA gene functioning during the oogenesis of various organisms, many intriguing problems remain unsolved. In this review, we describe nucleolus organizer region (NOR) activity in avian oocytes. Whereas oocytes from an adult avian ovary never reveal the formation of the nucleolus in the germinal vesicle (GV), an ovary from juvenile birds possesses both nucleolus-containing and non-nucleolus-containing oocytes. The evolutionary diversity of oocyte NOR functioning and the potential non-rRNA-related functions of the nucleolus in oocytes are also discussed.

  14. Totally optimal decision trees for Boolean functions

    KAUST Repository

    Chikalov, Igor

    2016-07-28

    We study decision trees which are totally optimal relative to different sets of complexity parameters for Boolean functions. A totally optimal tree is an optimal tree relative to each parameter from the set simultaneously. We consider the parameters characterizing both time (in the worst- and average-case) and space complexity of decision trees, i.e., depth, total path length (average depth), and number of nodes. We have created tools based on extensions of dynamic programming to study totally optimal trees. These tools are applicable to both exact and approximate decision trees, and allow us to make multi-stage optimization of decision trees relative to different parameters and to count the number of optimal trees. Based on the experimental results we have formulated the following hypotheses (and subsequently proved): for almost all Boolean functions there exist totally optimal decision trees (i) relative to the depth and number of nodes, and (ii) relative to the depth and average depth.

  15. Structure and function of echinoderm telomerase RNA

    OpenAIRE

    Podlevsky, Joshua D.; Li, Yang; Chen, Julian J.-L.

    2016-01-01

    Telomerase is a ribonucleoprotein (RNP) enzyme that requires an integral telomerase RNA (TR) subunit, in addition to the catalytic telomerase reverse transcriptase (TERT), for enzymatic function. The secondary structures of TRs from the three major groups of species, ciliates, fungi, and vertebrates, have been studied extensively and demonstrate dramatic diversity. Herein, we report the first comprehensive secondary structure of TR from echinoderms—marine invertebrates closely related to vert...

  16. Natural RNA circles function as efficient microRNA sponges

    DEFF Research Database (Denmark)

    Hansen, Thomas B; Jensen, Trine I; Clausen, Bettina H

    2013-01-01

    MicroRNAs (miRNAs) are important post-transcriptional regulators of gene expression that act by direct base pairing to target sites within untranslated regions of messenger RNAs. Recently, miRNA activity has been shown to be affected by the presence of miRNA sponge transcripts, the so......-called competing endogenous RNA in humans and target mimicry in plants. We previously identified a highly expressed circular RNA (circRNA) in human and mouse brain. Here we show that this circRNA acts as a miR-7 sponge; we term this circular transcript ciRS-7 (circular RNA sponge for miR-7). ciRS-7 contains more...... of miR-7 targets. In the mouse brain, we observe overlapping co-expression of ciRS-7 and miR-7, particularly in neocortical and hippocampal neurons, suggesting a high degree of endogenous interaction. We further show that the testis-specific circRNA, sex-determining region Y (Sry), serves as a miR-138...

  17. Total Variation Depth for Functional Data

    KAUST Repository

    Huang, Huang

    2016-11-15

    There has been extensive work on data depth-based methods for robust multivariate data analysis. Recent developments have moved to infinite-dimensional objects such as functional data. In this work, we propose a new notion of depth, the total variation depth, for functional data. As a measure of depth, its properties are studied theoretically, and the associated outlier detection performance is investigated through simulations. Compared to magnitude outliers, shape outliers are often masked among the rest of samples and harder to identify. We show that the proposed total variation depth has many desirable features and is well suited for outlier detection. In particular, we propose to decompose the total variation depth into two components that are associated with shape and magnitude outlyingness, respectively. This decomposition allows us to develop an effective procedure for outlier detection and useful visualization tools, while naturally accounting for the correlation in functional data. Finally, the proposed methodology is demonstrated using real datasets of curves, images, and video frames.

  18. Toward a more efficient isolation of total RNA from loquat (Eriobotrya japonica Lindl.) tissues

    OpenAIRE

    Martinez Fuentes, Amparo; Mesejo Conejos, Carlos; Agustí Fonfría, Manuel; Reig Valor, Carmina

    2015-01-01

    [EN] Loquat (Eriobotrya japonica Lindl.) tissues contain considerable amounts of polyphenolic compounds and polysaccharides which can copurify with RNA. This can render the RNA unsuitable for DNA synthesis and makes the isolation of high quality RNA from loquat tissues particularly difficult. Materials and methods. Total RNA was extracted from leaves, buds and fruit of loquat by adapting the method described by Song et al. in 2011. This involved improvements both to the extraction protocol an...

  19. Isolated total RNA and protein are preserved after thawing for more than twenty-four hours

    Science.gov (United States)

    de Oliveira, Ivone Braga; Ramos, Débora Rothstein; Lopes, Karen Lucasechi; de Souza, Regiane Machado; Heimann, Joel Claudio; Furukawa, Luzia Naôko Shinohara

    2012-01-01

    OBJECTIVE: The preservation of biological samples at a low temperature is important for later biochemical and/or histological analyses. However, the molecular viability of thawed samples has not been studied sufficiently in depth. The present study was undertaken to evaluate the viability of intact tissues, tissue homogenates, and isolated total RNA after defrosting for more than twenty-four hours. METHODS: The molecular viability of the thawed samples (n = 82) was assessed using the A260/A280 ratio, the RNA concentration, the RNA integrity, the level of intact mRNA determined by reverse transcriptase polymerase chain reaction, the protein level determined by Western blotting, and an examination of the histological structure. RESULTS: The integrity of the total RNA was not preserved in the thawed intact tissue, but the RNA integrity and level of mRNA were perfectly preserved in isolated defrosted samples of total RNA. Additionally, the level of β-actin protein was preserved in both thawed intact tissue and homogenates. CONCLUSION: Isolated total RNA does not undergo degradation due to thawing for at least 24 hours, and it is recommended to isolate the total RNA as soon as possible after tissue collection. Moreover, the protein level is preserved in defrosted tissues. PMID:22473407

  20. Total RNA Sequencing Analysis of DCIS Progressing to Invasive Breast Cancer

    Science.gov (United States)

    2015-09-01

    Assay Kits respectively on the Qubit 2.0 Fluorometer (Life Technologies). The BioRad Experion Automated Electrophoresis System RNA kit was used to...AWARD NUMBER: W81XWH-14-1-0080 TITLE: Total RNA Sequencing Analysis of DCIS Progressing to Invasive Breast Cancer. PRINCIPAL INVESTIGATOR...Aug 2015 4. TITLE AND SUBTITLE Total RNA Sequencing Analysis of DCIS Progressing to Invasive Breast Cancer. 5a. CONTRACT NUMBER 5b. GRANT

  1. Construction of sized eukaryotic cDNA libraries using low input of total environmental metatranscriptomic RNA.

    Science.gov (United States)

    Yadav, Rajiv Kumar; Barbi, Florian; Ziller, Antoine; Luis, Patricia; Marmeisse, Roland; Reddy, M Sudhakara; Fraissinet-Tachet, Laurence

    2014-09-03

    Construction of high quality cDNA libraries from the usually low amounts of eukaryotic mRNA extracted from environmental samples is essential in functional metatranscriptomics for the selection of functional, full-length genes encoding proteins of interest. Many of the inserts in libraries constructed by standard methods are represented by truncated cDNAs due to premature stoppage of reverse transcriptase activity and preferential cloning of short cDNAs. We report here a simple and cost effective technique for preparation of sized eukaryotic cDNA libraries from as low as three microgram of total soil RNA dominated by ribosomal and bacterial RNA. cDNAs synthesized by a template switching approach were size-fractionated by two dimensional agarose gel electrophoresis prior to PCR amplification and cloning. Effective size selection was demonstrated by PCR amplification of conserved gene families specific of each size class. Libraries of more than one million independent inserts whose sizes ranged between one and four kb were thus produced. Up to 80% of the insert sequences were homologous to eukaryotic gene sequences present in public databases. A simple and cost effective technique has been developed to construct sized eukaryotic cDNA libraries from environmental samples. This technique will facilitate expression cloning of environmental eukaryotic genes and contribute to a better understanding of basic biological and/or ecological processes carried out by eukaryotic microbial communities.

  2. Biochemistry and Function of the RNA Exosomes

    DEFF Research Database (Denmark)

    Lubas, Michal Szymon; Chlebowski, Aleksander; Dziembowski, Andrzej

    2012-01-01

    Discovery of the evolutionary conserved RNA exosome was a milestone in RNA biology. First identified as an activity essential for the processing of ribosomal RNA, the exosome has since proved to be central for RNA processing and degradation in both the nucleus and the cytoplasm of eukaryotic cell...

  3. MicroRNA function in Drosophila melanogaster.

    Science.gov (United States)

    Carthew, Richard W; Agbu, Pamela; Giri, Ritika

    2017-05-01

    Over the last decade, microRNAs have emerged as critical regulators in the expression and function of animal genomes. This review article discusses the relationship between microRNA-mediated regulation and the biology of the fruit fly Drosophila melanogaster. We focus on the roles that microRNAs play in tissue growth, germ cell development, hormone action, and the development and activity of the central nervous system. We also discuss the ways in which microRNAs affect robustness. Many gene regulatory networks are robust; they are relatively insensitive to the precise values of reaction constants and concentrations of molecules acting within the networks. MicroRNAs involved in robustness appear to be nonessential under uniform conditions used in conventional laboratory experiments. However, the robust functions of microRNAs can be revealed when environmental or genetic variation otherwise has an impact on developmental outcomes. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. Structure and function of echinoderm telomerase RNA.

    Science.gov (United States)

    Podlevsky, Joshua D; Li, Yang; Chen, Julian J-L

    2016-02-01

    Telomerase is a ribonucleoprotein (RNP) enzyme that requires an integral telomerase RNA (TR) subunit, in addition to the catalytic telomerase reverse transcriptase (TERT), for enzymatic function. The secondary structures of TRs from the three major groups of species, ciliates, fungi, and vertebrates, have been studied extensively and demonstrate dramatic diversity. Herein, we report the first comprehensive secondary structure of TR from echinoderms-marine invertebrates closely related to vertebrates-determined by phylogenetic comparative analysis of 16 TR sequences from three separate echinoderm classes. Similar to vertebrate TR, echinoderm TR contains the highly conserved template/pseudoknot and H/ACA domains. However, echinoderm TR lacks the ancestral CR4/5 structural domain found throughout vertebrate and fungal TRs. Instead, echinoderm TR contains a distinct simple helical region, termed eCR4/5, that is functionally equivalent to the CR4/5 domain. The urchin and brittle star eCR4/5 domains bind specifically to their respective TERT proteins and stimulate telomerase activity. Distinct from vertebrate telomerase, the echinoderm TR template/pseudoknot domain with the TERT protein is sufficient to reconstitute significant telomerase activity. This gain-of-function of the echinoderm template/pseudoknot domain for conferring telomerase activity presumably facilitated the rapid structural evolution of the eCR4/5 domain throughout the echinoderm lineage. Additionally, echinoderm TR utilizes the template-adjacent P1.1 helix as a physical template boundary element to prevent nontelomeric DNA synthesis, a mechanism used by ciliate and fungal TRs. Thus, the chimeric and eccentric structural features of echinoderm TR provide unparalleled insights into the rapid evolution of telomerase RNP structure and function. © 2016 Podlevsky et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  5. Functions of microRNA in response to cocaine stimulation.

    Science.gov (United States)

    Xu, L-F; Wang, J; Lv, F B; Song, Q

    2013-12-04

    MicroRNAs (miRNAs) are a type of non-protein-coding single-stranded RNA, which are typically 20-25 nt in length. miRNAs play important roles in various biological processes, including development, cell proliferation, differentiation, and apoptosis. We aimed to detect the miRNA response to cocaine stimulations and their target genes. Using the miRNA expression data GSE21901 downloaded from the Gene Expression Omnibus database, we screened out the differentially expressed miRNA after short-term (1 h) and longer-term (6 h) cocaine stimulations based on the fold change >1.2. Target genes of differentially expressed miRNAs were retrieved from TargetScan database with the context score -0.3. Functional annotation enrichment analysis was performed for all the target genes with DAVID. A total of 121 differentially expressed miRNAs between the 1-h treatment and the control samples, 58 between the 6-h treatment and the control samples, and 69 between the 1-h and the 6-h treatment samples. Among them, miR-212 results of particular interest, since its expression level was constantly elevated responding to cocaine treatment. After functional and pathway annotations of target genes, we proved that miR-212 was a critical element in cocaine-addiction, because of its involvement in regulating several important cell cycle events. The results may pave the way for further understanding the regulatory mechanisms of cocaine-response in human bodies.

  6. Small catalytic RNA: Structure, function and application

    Energy Technology Data Exchange (ETDEWEB)

    Monforte, Joseph Albert [Univ. of California, Berkeley, CA (United States)

    1991-04-01

    We have utilized a combination of photochemical cross-linking techniques and site-directed mutagenesis to obtain secondary and tertiary structure information for the self-cleaving, self-ligating subsequence of RNA from the negative strand of Satellite Tobacco Ringspot Virus. We have found that the helical regions fold about a hinge to promoting four different possible tertiary interactions, creating a molecular of similar shape to a paperclip. A model suggesting that the ``paperclip`` and ``hammerhead`` RNAs share a similar three dimensional structure is proposed. We have used a self-cleaving RNA molecule related to a subsequence of plant viroids, a ``hammerhead,`` to study the length-dependent folding of RNA produced during transcription by RNA polymerase. We have used this method to determine the length of RNA sequestered within elongating E. coli and T7 RNA polymerase complexes. The data show that for E. coli RNA polymerase 121±s are sequestered within the ternary complex, which is consistent with the presence of an RNA-DNA hybrid within the transcription bubble, as proposed by others. The result for T7 RNA polymerase differs from E. coli RNA polymerase, with only 10{plus_minus}1 nucleotides sequestered within the ternary complex, setting a new upper limit for the minimum RNA-DNA required for a stable elongating complex. Comparisons between E. coli and T7 RNA polymerase are made. The relevance of the results to models or transcription termination, abortive initiation, and initiation to elongation mode transitions are discussed.

  7. Isolation of High-Quality Total RNA from Chinese Fir (Cunninghamia lanceolata (Lamb. Hook.

    Directory of Open Access Journals (Sweden)

    Zhihui Ma

    Full Text Available RNA isolation with RNA in a high quantity is a basic analytical method in plant genetics, molecular biology and related physiological investigations. To understand the genetic and molecular biology of Chinese fir, sufficient high-quality total RNA must be obtained for cDNA library construction and other downstream molecular applications. However, extracting RNA from Chinese fir is difficult and often requires the modification of existing protocols. Chinese fir tissues containing large amounts of polysaccharides and polyphenol compounds and are one of the most difficult plant tissues for RNA isolation. Therefore, we developed a simple method for extracting high-quality RNA from Chinese fir tissues. RNA isolations were performed within two hours, RNA quality was measured for yield and purity. Total RNA obtained from this procedure was successfully used for cDNA library construction, RT-PCR and transcriptome sequencing. It was proven that extracted RNA was intact and suitable for downstream molecular applications, including RT-PCR and qPCR, and other downstream molecular applications. Thus, this protocol represents a simple, efficient, and low-cost method.

  8. Identification of Subtype Specific miRNA-mRNA Functional Regulatory Modules in Matched miRNA-mRNA Expression Data: Multiple Myeloma as a Case

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    Yunpeng Zhang

    2015-01-01

    Full Text Available Identification of miRNA-mRNA modules is an important step to elucidate their combinatorial effect on the pathogenesis and mechanisms underlying complex diseases. Current identification methods primarily are based upon miRNA-target information and matched miRNA and mRNA expression profiles. However, for heterogeneous diseases, the miRNA-mRNA regulatory mechanisms may differ between subtypes, leading to differences in clinical behavior. In order to explore the pathogenesis of each subtype, it is important to identify subtype specific miRNA-mRNA modules. In this study, we integrated the Ping-Pong algorithm and multiobjective genetic algorithm to identify subtype specific miRNA-mRNA functional regulatory modules (MFRMs through integrative analysis of three biological data sets: GO biological processes, miRNA target information, and matched miRNA and mRNA expression data. We applied our method on a heterogeneous disease, multiple myeloma (MM, to identify MM subtype specific MFRMs. The constructed miRNA-mRNA regulatory networks provide modular outlook at subtype specific miRNA-mRNA interactions. Furthermore, clustering analysis demonstrated that heterogeneous MFRMs were able to separate corresponding MM subtypes. These subtype specific MFRMs may aid in the further elucidation of the pathogenesis of each subtype and may serve to guide MM subtype diagnosis and treatment.

  9. Drosha regulates gene expression independently of RNA cleavage function

    DEFF Research Database (Denmark)

    Gromak, Natalia; Dienstbier, Martin; Macias, Sara

    2013-01-01

    Drosha is the main RNase III-like enzyme involved in the process of microRNA (miRNA) biogenesis in the nucleus. Using whole-genome ChIP-on-chip analysis, we demonstrate that, in addition to miRNA sequences, Drosha specifically binds promoter-proximal regions of many human genes in a transcription......-terminal protein-interaction domain, which associates with the RNA-binding protein CBP80 and RNA Polymerase II. Consequently, we uncover a previously unsuspected RNA cleavage-independent function of Drosha in the regulation of human gene expression....

  10. Approaching TERRA Firma: Genomic Functions of Telomeric Noncoding RNA.

    Science.gov (United States)

    Roake, Caitlin M; Artandi, Steven E

    2017-06-29

    Functions of the telomeric repeat-containing RNA (TERRA), the long noncoding RNA (lncRNA) transcribed from telomeres, have eluded researchers. In this issue of Cell, Graf el al. and Chu et al. uncover new regulatory roles for TERRA at the telomere and at distant genomic sites. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Small catalytic RNA: Structure, function and application

    Energy Technology Data Exchange (ETDEWEB)

    Monforte, J.A.

    1991-04-01

    We have utilized a combination of photochemical cross-linking techniques and site-directed mutagenesis to obtain secondary and tertiary structure information for the self-cleaving, self-ligating subsequence of RNA from the negative strand of Satellite Tobacco Ringspot Virus. We have found that the helical regions fold about a hinge to promoting four different possible tertiary interactions, creating a molecular of similar shape to a paperclip. A model suggesting that the paperclip'' and hammerhead'' RNAs share a similar three dimensional structure is proposed. We have used a self-cleaving RNA molecule related to a subsequence of plant viroids, a hammerhead,'' to study the length-dependent folding of RNA produced during transcription by RNA polymerase. We have used this method to determine the length of RNA sequestered within elongating E. coli and T7 RNA polymerase complexes. The data show that for E. coli RNA polymerase 12{plus minus}1 nucleotides are sequestered within the ternary complex, which is consistent with the presence of an RNA-DNA hybrid within the transcription bubble, as proposed by others. The result for T7 RNA polymerase differs from E. coli RNA polymerase, with only 10{plus minus}1 nucleotides sequestered within the ternary complex, setting a new upper limit for the minimum RNA-DNA required for a stable elongating complex. Comparisons between E. coli and T7 RNA polymerase are made. The relevance of the results to models or transcription termination, abortive initiation, and initiation to elongation mode transitions are discussed.

  12. An Improved Quick Method for the Isolation of Total RNA from Cotton ...

    African Journals Online (AJOL)

    David PANG

    2011-11-02

    Nov 2, 2011 ... However, genetic and functional genomics in plants including cotton requires, isolating the RNA from a large number of plant tissue samples or a mapping population. The objective of this study was to overcome these problems encountered in RNA extraction and develop a quick as well as a simple method ...

  13. Evaluating Methods for Isolating Total RNA and Predicting the Success of Sequencing Phylogenetically Diverse Plant Transcriptomes

    Science.gov (United States)

    Bruskiewich, Richard; Burris, Jason N.; Carrigan, Charlotte T.; Chase, Mark W.; Clarke, Neil D.; Covshoff, Sarah; dePamphilis, Claude W.; Edger, Patrick P.; Goh, Falicia; Graham, Sean; Greiner, Stephan; Hibberd, Julian M.; Jordon-Thaden, Ingrid; Kutchan, Toni M.; Leebens-Mack, James; Melkonian, Michael; Miles, Nicholas; Myburg, Henrietta; Patterson, Jordan; Pires, J. Chris; Ralph, Paula; Rolf, Megan; Sage, Rowan F.; Soltis, Douglas; Soltis, Pamela; Stevenson, Dennis; Stewart, C. Neal; Surek, Barbara; Thomsen, Christina J. M.; Villarreal, Juan Carlos; Wu, Xiaolei; Zhang, Yong; Deyholos, Michael K.; Wong, Gane Ka-Shu

    2012-01-01

    Next-generation sequencing plays a central role in the characterization and quantification of transcriptomes. Although numerous metrics are purported to quantify the quality of RNA, there have been no large-scale empirical evaluations of the major determinants of sequencing success. We used a combination of existing and newly developed methods to isolate total RNA from 1115 samples from 695 plant species in 324 families, which represents >900 million years of phylogenetic diversity from green algae through flowering plants, including many plants of economic importance. We then sequenced 629 of these samples on Illumina GAIIx and HiSeq platforms and performed a large comparative analysis to identify predictors of RNA quality and the diversity of putative genes (scaffolds) expressed within samples. Tissue types (e.g., leaf vs. flower) varied in RNA quality, sequencing depth and the number of scaffolds. Tissue age also influenced RNA quality but not the number of scaffolds ≥1000 bp. Overall, 36% of the variation in the number of scaffolds was explained by metrics of RNA integrity (RIN score), RNA purity (OD 260/230), sequencing platform (GAIIx vs HiSeq) and the amount of total RNA used for sequencing. However, our results show that the most commonly used measures of RNA quality (e.g., RIN) are weak predictors of the number of scaffolds because Illumina sequencing is robust to variation in RNA quality. These results provide novel insight into the methods that are most important in isolating high quality RNA for sequencing and assembling plant transcriptomes. The methods and recommendations provided here could increase the efficiency and decrease the cost of RNA sequencing for individual labs and genome centers. PMID:23185583

  14. [Effective method for isolation of total RNA from Fagopyrum cymosum callus].

    Science.gov (United States)

    Sui, Shunzhao; Ma, Jing; Li, Yandong; Zhu, Qinlong; Li, Mingyang

    2011-03-01

    To establish a method for isolation of the total RNA from Fagopyrum cymosum callus. The improved method combining that of CTAB extraction with the LiCl precipitation was used to isolate the total RNA from the four F. cymosum callus. The quality of the RNA was detected by UV spectrophotometric analysis, 0.8% non-denaturing agarose gel electrophoresis and RNA reverse transcription. The bands of 28S and 18S could be seen clearly by agarose gel electrophoresis, and the value of A260/A280 was between 1.9 and 2.0. The cDNA which was reverse-transcribed by the total RNA showed a wide length rage of 500 bp-5 kb. The RNA extracted by this method meets the requirement of reverse transcription-polymerase chain reaction (RT-PCR), construction of cDNA libraries, et al. This improved method can be used to isolate the total RNA from F. cymosum callus with the advantage of simpleness, efficiency and low cost.

  15. Functional characterization of the Drosophila MRP (mitochondrial RNA processing) RNA gene.

    Science.gov (United States)

    Schneider, Mary D; Bains, Anupinder K; Rajendra, T K; Dominski, Zbigniew; Matera, A Gregory; Simmonds, Andrew J

    2010-11-01

    MRP RNA is a noncoding RNA component of RNase mitochondrial RNA processing (MRP), a multi-protein eukaryotic endoribonuclease reported to function in multiple cellular processes, including ribosomal RNA processing, mitochondrial DNA replication, and cell cycle regulation. A recent study predicted a potential Drosophila ortholog of MRP RNA (CR33682) by computer-based genome analysis. We have confirmed the expression of this gene and characterized the phenotype associated with this locus. Flies with mutations that specifically affect MRP RNA show defects in growth and development that begin in the early larval period and end in larval death during the second instar stage. We present several lines of evidence demonstrating a role for Drosophila MRP RNA in rRNA processing. The nuclear fraction of Drosophila MRP RNA localizes to the nucleolus. Further, a mutant strain shows defects in rRNA processing that include a defect in 5.8S rRNA processing, typical of MRP RNA mutants in other species, as well as defects in early stages of rRNA processing.

  16. Efficient isolation of total RNA from antibiotic-producing bacterium Amycolatopsis mediterranei.

    Science.gov (United States)

    Yao, Yufeng; Zhang, Weiwen; Jiao, Ruishen; Zhao, Guoping; Jiang, Weihong

    2002-10-01

    RNA extraction from antibiotic-producing actinomycetes can be a difficult and time-consuming process due to their special peptidoglycans cell wall composition and the short life of RNA. Hence, the rapidity of cellular lysis and complete inhibition of RNase are of particular importance for isolating intact RNA of high quality. The genus of Amycolatopsis mediterranei produces many clinically important antibiotics, such as rifamycin and vancomycin; however, the available methods for bacterial RNA isolation did not work very well with this genus. In this report, we described a new method for RNA isolation using the combination of LiCl, urea and guanidinium thiocyanate to disrupt the cell wall of Amycolatopsis. Compared with earlier published RNA isolation methods, the method gave higher yields of pure and intact RNA. About 1 microg total RNA free of DNA contamination can be obtained from 1 mg wet weight of A. mediterranei. The integrity of the RNA was demonstrated by formaldehyde agarose gel electrophoresis and Northern blot analyses.

  17. Synthetic RNA Controllers for Programming Mammalian Cell Fate and Function

    Science.gov (United States)

    2015-11-04

    Final report for “Synthetic RNA controllers for programming mammalian cell fate and function” Principal Investigator: Christina D. Smolke...SUBTITLE Synthetic RNA controllers for programming mammalian cell fate and function 5a. CONTRACT NUMBER 5b. GRANT NUMBER 5c. PROGRAM ELEMENT NUMBER...Form 298 (Rev. 8-98) Prescribed by ANSI Std Z39-18   2 Synthetic RNA controllers for programming mammalian cell fate and function Task 1

  18. Renal function after elective total hip replacement

    DEFF Research Database (Denmark)

    Perregaard, Helene; Damholt, Mette B; Solgaard, Søren

    2016-01-01

    and the prevalence of chronic kidney disease (CKD) in an elective population of orthopedic patients undergoing primary total hip replacement, hypothesizing that chronic kidney disease predisposes to AKI. Patients and methods - This was a single-center, population-based, retrospective, registry-based cohort study...... involving all primary elective total hip replacements performed from January 2003 through December 2012. Patient demographics and creatinine values were registered. We evaluated the presence of CKD and AKI according to the international guidelines for kidney disease (KDIGO Acute Kidney Injury Workgroup 2013...... ). Results - 3,416 patients were included (2,064 females (60%)). AKI (according to KDIGO criteria) was seen in 75 patients (2.2%, 95% CI: 1.7-2.7) in the course of primary total hip replacement. Of these, 26 had pre-existing CKD of class 3-5. Pre-existing CKD of class 3-5, indicating moderately to severely...

  19. High-quality total RNA isolation from melon (Cucumis melo L. fruits rich in polysaccharides

    Directory of Open Access Journals (Sweden)

    Gabrielle Silveira de Campos

    2017-08-01

    Full Text Available Melon, a member of the family Cucurbitaceae, is the fourth most important fruit in the world market and, on a volume basis, is Brazil’s main fresh fruit export. Many molecular techniques used to understand the maturation of these fruits require high concentrations of highly purified RNA. However, melons are rich in polyphenolic compounds and polysaccharides, which interfere with RNA extraction. This study aimed to determine the most appropriate method for total RNA extraction from melon fruits. Six extraction buffers were tested: T1 guanidine thiocyanate/phenol/chloroform; T2 sodium azide/?-mercaptoethanol; T3 phenol/guanidine thiocyanate; T4 CTAB/PVP/?-mercaptoethanol; T5 SDS/sodium perchlorate/PVP/?-mercaptoethanol, and T6 sarkosyl/PVP/guanidine thiocyanate, using the AxyPrepTM Multisource Total RNA Miniprep Kit. The best method for extracting RNA from both mature and green fruit was based on the SDS/PVP/?-mercaptoethanol buffer, because it rapidly generated a high quality and quantity of material. In general, higher amounts of RNA were obtained from green than mature fruits, probably due to the lower concentration of polysaccharides and water. The purified material can be used as a template in molecular techniques, such as microarrays, RT-PCR, and in the construction of cDNA and RNA-seq data.

  20. Production Function Geometry with "Knightian" Total Product

    Science.gov (United States)

    Truett, Dale B.; Truett, Lila J.

    2007-01-01

    Authors of principles and price theory textbooks generally illustrate short-run production using a total product curve that displays first increasing and then diminishing marginal returns to employment of the variable input(s). Although it seems reasonable that a temporary range of increasing returns to variable inputs will likely occur as…

  1. Total scattering of disordered crystalline functional materials

    International Nuclear Information System (INIS)

    Shamoto, Shin-Ichi; Kodama, Katsuaki; Iikubo, Satoshi; Taguchi, Tomitsugu

    2009-01-01

    There are disorders in some modern functional materials. As an example, the crystalline phase of an optical recording material has low thermal conductivity but high electrical conductivity, simultaneously. This contradiction is a challenge to material scientists in designing good functional materials, which should have at least two types of crystallographic sites. One site limits thermal conductivity while the other site carries electrons or holes with high mobility. This problem exists with not only optical recording materials but also thermoelectric materials. The periodic boundary condition gets lost in the disordered parts. This therefore, makes atomic pair distribution function (PDF) analysis with a wide range of real space suitable for investigating the form and size of crystalline parts as well as disordered parts in the material. Pulsed neutron powder diffraction is one of the best tools for use in this new type of emerging research, together with synchrotron X-ray powder diffraction and electron diffraction.

  2. Functional RNA during Zika virus infection

    NARCIS (Netherlands)

    Göertz, Giel P.; Abbo, Sandra R.; Fros, Jelke J.; Pijlman, Gorben P.

    2017-01-01

    Zika virus (ZIKV; family Flaviviridae; genus Flavivirus) is a pathogenic mosquito-borne RNA virus that currently threatens human health in the Americas, large parts of Asia and occasionally elsewhere in the world. ZIKV infection is often asymptomatic but can cause severe symptoms including

  3. A Broad RNA Virus Survey Reveals Both miRNA Dependence and Functional Sequestration

    DEFF Research Database (Denmark)

    Scheel, Troels K H; Luna, Joseph M; Liniger, Matthias

    2016-01-01

    , critically depended on the interaction of cellular miR-17 and let-7 with the viral 3' UTR. Unlike canonical miRNA interactions, miR-17 and let-7 binding enhanced pestivirus translation and RNA stability. miR-17 sequestration by pestiviruses conferred reduced AGO binding and functional de...... immunoprecipitation (CLIP) of the Argonaute (AGO) proteins to characterize strengths and specificities of miRNA interactions in the context of 15 different RNA virus infections, including several clinically relevant pathogens. Notably, replication of pestiviruses, a major threat to milk and meat industries...

  4. Comparative evaluation of total RNA extraction methods in Theobroma cacao using shoot apical meristems.

    Science.gov (United States)

    Silva, D V; Branco, S M J; Holanda, I S A; Royaert, S; Motamayor, J C; Marelli, J P; Corrêa, R X

    2016-03-04

    Theobroma cacao is a species of great economic importance with its beans used for chocolate production. The tree has been a target of various molecular studies. It contains many polyphenols, which complicate the extraction of nucleic acids with the extraction protocols requiring a large amount of plant material. These issues, therefore, necessitate the optimization of the protocols. The aim of the present study was to evaluate different methods for extraction of total RNA from shoot apical meristems of T. cacao 'CCN 51' and to assess the influence of storage conditions for the meristems on the extraction. The study also aimed to identify the most efficient protocol for RNA extraction using a small amount of plant material. Four different protocols were evaluated for RNA extraction using one shoot apical meristem per sample. Among these protocols, one that was more efficient was then tested to extract RNA using four different numbers of shoot apical meristems, subjected to three different storage conditions. The best protocol was tested for cDNA amplification using reverse transcription-polymerase chain reaction; the cDNA quality was determined to be satisfactory for molecular analyses. The study revealed that with the best RNA extraction protocol, one shoot apical meristem was sufficient for extraction of high-quality total RNA. The results obtained might enable advances in genetic analyses and molecular studies using reduced amount of plant material.

  5. An improved method for the isolation of total RNA from Avicennia germinans leaves.

    Science.gov (United States)

    Gonzalez-Mendoza, Daniel; Moreno, Adriana Quiroz; Zapata-Perez, Omar

    2008-01-01

    Isolation of high-quality RNA of Avicennia germinans L. tissue is difficult due to high levels of phenols and other substances that interfere when using conventional procedures for the isolation. These substances not only decrease the yield but also the quality of RNA is almost poor. We present here a simple RNA protocol and fast methodology that effectively removes these contaminating substances without affecting the yield. The protocol developed is based on the SDS/phenol method with modifications including beta-mercaptoethanol to prevent oxidation of phenolic complexes, and phenol/chloroform extraction is introduced to remove proteins, genomic DNA, and secondary metabolites, and co-precipitated polysaccharides. Both A260/A230 and A260/A280 absorbance ratios of isolated RNA were around 2 and the yield was about 0.3 mg g(-1) fresh weight. Good-quality total RNA from leaves of Avicennia germinans could be easily isolated within 2 h by this protocol which avoided the limitation of plant materials and could provide total RNA for all kinds of further molecular studies.

  6. A method for the construction of equalized directional cDNA libraries from hydrolyzed total RNA.

    Science.gov (United States)

    Davis, Claytus; Barvish, Zeev; Gitelman, Inna

    2007-10-09

    The transcribed sequences of a cell, the transcriptome, represent the trans-acting fraction of the genetic information, yet eukaryotic cDNA libraries are typically made from only the poly-adenylated fraction. The non-coding or translated but non-polyadenylated RNAs are therefore not represented. The goal of this study was to develop a method that would more completely represent the transcriptome in a useful format, avoiding over-representation of some of the abundant, but low-complexity non-translated transcripts. We developed a combination of self-subtraction and directional cloning procedures for this purpose. Libraries were prepared from partially degraded (hydrolyzed) total RNA from three different species. A restriction endonuclease site was added to the 3' end during first-strand synthesis using a directional random-priming technique. The abundant non-polyadenylated rRNA and tRNA sequences were largely removed by using self-subtraction to equalize the representation of the various RNA species. Sequencing random clones from the libraries showed that 87% of clones were in the forward orientation with respect to known or predicted transcripts. 70% matched identified or predicted translated RNAs in the sequence databases. Abundant mRNAs were less frequent in the self-subtracted libraries compared to a non-subtracted mRNA library. 3% of the sequences were from known or hypothesized ncRNA loci, including five matches to miRNA loci. We describe a simple method for making high-quality, directional, random-primed, cDNA libraries from small amounts of degraded total RNA. This technique is advantageous in situations where a cDNA library with complete but equalized representation of transcribed sequences, whether polyadenylated or not, is desired.

  7. A method for the construction of equalized directional cDNA libraries from hydrolyzed total RNA

    Directory of Open Access Journals (Sweden)

    Gitelman Inna

    2007-10-01

    Full Text Available Abstract Background The transcribed sequences of a cell, the transcriptome, represent the trans-acting fraction of the genetic information, yet eukaryotic cDNA libraries are typically made from only the poly-adenylated fraction. The non-coding or translated but non-polyadenylated RNAs are therefore not represented. The goal of this study was to develop a method that would more completely represent the transcriptome in a useful format, avoiding over-representation of some of the abundant, but low-complexity non-translated transcripts. Results We developed a combination of self-subtraction and directional cloning procedures for this purpose. Libraries were prepared from partially degraded (hydrolyzed total RNA from three different species. A restriction endonuclease site was added to the 3' end during first-strand synthesis using a directional random-priming technique. The abundant non-polyadenylated rRNA and tRNA sequences were largely removed by using self-subtraction to equalize the representation of the various RNA species. Sequencing random clones from the libraries showed that 87% of clones were in the forward orientation with respect to known or predicted transcripts. 70% matched identified or predicted translated RNAs in the sequence databases. Abundant mRNAs were less frequent in the self-subtracted libraries compared to a non-subtracted mRNA library. 3% of the sequences were from known or hypothesized ncRNA loci, including five matches to miRNA loci. Conclusion We describe a simple method for making high-quality, directional, random-primed, cDNA libraries from small amounts of degraded total RNA. This technique is advantageous in situations where a cDNA library with complete but equalized representation of transcribed sequences, whether polyadenylated or not, is desired.

  8. Nuclear export of RNA: Different sizes, shapes and functions.

    Science.gov (United States)

    Williams, Tobias; Ngo, Linh H; Wickramasinghe, Vihandha O

    2018-03-01

    Export of protein-coding and non-coding RNA molecules from the nucleus to the cytoplasm is critical for gene expression. This necessitates the continuous transport of RNA species of different size, shape and function through nuclear pore complexes via export receptors and adaptor proteins. Here, we provide an overview of the major RNA export pathways in humans, highlighting the similarities and differences between each. Its importance is underscored by the growing appreciation that deregulation of RNA export pathways is associated with human diseases like cancer. Crown Copyright © 2017. Published by Elsevier Ltd. All rights reserved.

  9. A mini-scale hot borate method for the isolation of total RNA from a ...

    African Journals Online (AJOL)

    By modifying and scaling down a hot borate extraction method, a quick and easy method was developed to extract total RNA from ovaries of three genotypes ranging in ages from -1 to 10 days post anthesis (dpa) with an average yield of 600 μg/gfw (gram fresh weight) and ten dpa ovaries of 64 genotypes. The method was ...

  10. Functional Integration of mRNA Translational Control Programs

    Directory of Open Access Journals (Sweden)

    Melanie C. MacNicol

    2015-07-01

    Full Text Available Regulated mRNA translation plays a key role in control of cell cycle progression in a variety of physiological and pathological processes, including in the self-renewal and survival of stem cells and cancer stem cells. While targeting mRNA translation presents an attractive strategy for control of aberrant cell cycle progression, mRNA translation is an underdeveloped therapeutic target. Regulated mRNAs are typically controlled through interaction with multiple RNA binding proteins (RBPs but the mechanisms by which the functions of distinct RBPs bound to a common target mRNA are coordinated are poorly understood. The challenge now is to gain insight into these mechanisms of coordination and to identify the molecular mediators that integrate multiple, often conflicting, inputs. A first step includes the identification of altered mRNA ribonucleoprotein complex components that assemble on mRNAs bound by multiple, distinct RBPs compared to those recruited by individual RBPs. This review builds upon our knowledge of combinatorial control of mRNA translation during the maturation of oocytes from Xenopus laevis, to address molecular strategies that may mediate RBP diplomacy and conflict resolution for coordinated control of mRNA translational output. Continued study of regulated ribonucleoprotein complex dynamics promises valuable new insights into mRNA translational control and may suggest novel therapeutic strategies for the treatment of disease.

  11. The complexity, function and applications of RNA in circulation

    Directory of Open Access Journals (Sweden)

    Alton eEtheridge

    2013-06-01

    Full Text Available Blood carries a wide array of biomolecules, including nutrients, hormones and molecules that are secreted by cells for specific biological functions. The recent finding of stable RNA of both endogenous and exogenous origin in the circulation raises a number of questions and opens a broad, new field: exploring the origins, functions and applications of these extracellular RNA molecules. These findings raise many important questions, including: what are the mechanisms of export and cellular uptake, what is the nature and source of their stability, what molecules do they interact with in the blood, and what are the possible biological functions of the circulating RNA. This review summarizes some key recent developments in circulating RNA research and discusses some of the open questions in the field.

  12. Construction of sized eukaryotic cDNA libraries using low input of total environmental metatranscriptomic RNA

    OpenAIRE

    Yadav, Rajiv Kumar; Barbi, Florian; Ziller, Antoine; Luis, Patricia; Marmeisse, Roland; Reddy, M Sudhakara; Fraissinet-Tachet, Laurence

    2014-01-01

    Background: Construction of high quality cDNA libraries from the usually low amounts of eukaryotic mRNA extracted from environmental samples is essential in functional metatranscriptomics for the selection of functional, full-length genes encoding proteins of interest. Many of the inserts in libraries constructed by standard methods are represented by truncated cDNAs due to premature stoppage of reverse transcriptase activity and preferential cloning of short cDNAs. Results: We report here a ...

  13. Open Maximal Mucosa-Sparing Functional Total Laryngectomy

    Directory of Open Access Journals (Sweden)

    Pavel Dulguerov

    2017-10-01

    Full Text Available BackgroundTotal laryngectomy after (chemoradiotherapy is associated with a high incidence of fistula and therefore flaps are advocated. The description of a transoral robotic total laryngectomy prompted us to develop similar minimally invasive open approaches for functional total laryngectomy.MethodsA retrospective study of consecutive unselected patients with a dysfunctional larynx after (chemoradiation that underwent open maximal mucosal-sparing functional total laryngectomy (MMSTL between 2014 and 2016 is presented. The surgical technique is described, and the complications and functional outcome are reviewed.ResultsThe cohorts included 10 patients who underwent open MMSTL. No pedicled flap was used. Only one postoperative fistula was noted (10%. All patients resumed oral diet and experienced a functional tracheo-esophageal voice.ConclusionMMSTL could be used to perform functional total laryngectomy without a robot and with minimal incidence of complications.

  14. Human Enterovirus Nonstructural Protein 2CATPase Functions as Both an RNA Helicase and ATP-Independent RNA Chaperone

    Science.gov (United States)

    Xia, Hongjie; Wang, Peipei; Wang, Guang-Chuan; Yang, Jie; Sun, Xianlin; Wu, Wenzhe; Qiu, Yang; Shu, Ting; Zhao, Xiaolu; Yin, Lei; Qin, Cheng-Feng; Hu, Yuanyang; Zhou, Xi

    2015-01-01

    RNA helicases and chaperones are the two major classes of RNA remodeling proteins, which function to remodel RNA structures and/or RNA-protein interactions, and are required for all aspects of RNA metabolism. Although some virus-encoded RNA helicases/chaperones have been predicted or identified, their RNA remodeling activities in vitro and functions in the viral life cycle remain largely elusive. Enteroviruses are a large group of positive-stranded RNA viruses in the Picornaviridae family, which includes numerous important human pathogens. Herein, we report that the nonstructural protein 2CATPase of enterovirus 71 (EV71), which is the major causative pathogen of hand-foot-and-mouth disease and has been regarded as the most important neurotropic enterovirus after poliovirus eradication, functions not only as an RNA helicase that 3′-to-5′ unwinds RNA helices in an adenosine triphosphate (ATP)-dependent manner, but also as an RNA chaperone that destabilizes helices bidirectionally and facilitates strand annealing and complex RNA structure formation independently of ATP. We also determined that the helicase activity is based on the EV71 2CATPase middle domain, whereas the C-terminus is indispensable for its RNA chaperoning activity. By promoting RNA template recycling, 2CATPase facilitated EV71 RNA synthesis in vitro; when 2CATPase helicase activity was impaired, EV71 RNA replication and virion production were mostly abolished in cells, indicating that 2CATPase-mediated RNA remodeling plays a critical role in the enteroviral life cycle. Furthermore, the RNA helicase and chaperoning activities of 2CATPase are also conserved in coxsackie A virus 16 (CAV16), another important enterovirus. Altogether, our findings are the first to demonstrate the RNA helicase and chaperoning activities associated with enterovirus 2CATPase, and our study provides both in vitro and cellular evidence for their potential roles during viral RNA replication. These findings increase our

  15. Human Enterovirus Nonstructural Protein 2CATPase Functions as Both an RNA Helicase and ATP-Independent RNA Chaperone.

    Directory of Open Access Journals (Sweden)

    Hongjie Xia

    2015-07-01

    Full Text Available RNA helicases and chaperones are the two major classes of RNA remodeling proteins, which function to remodel RNA structures and/or RNA-protein interactions, and are required for all aspects of RNA metabolism. Although some virus-encoded RNA helicases/chaperones have been predicted or identified, their RNA remodeling activities in vitro and functions in the viral life cycle remain largely elusive. Enteroviruses are a large group of positive-stranded RNA viruses in the Picornaviridae family, which includes numerous important human pathogens. Herein, we report that the nonstructural protein 2CATPase of enterovirus 71 (EV71, which is the major causative pathogen of hand-foot-and-mouth disease and has been regarded as the most important neurotropic enterovirus after poliovirus eradication, functions not only as an RNA helicase that 3'-to-5' unwinds RNA helices in an adenosine triphosphate (ATP-dependent manner, but also as an RNA chaperone that destabilizes helices bidirectionally and facilitates strand annealing and complex RNA structure formation independently of ATP. We also determined that the helicase activity is based on the EV71 2CATPase middle domain, whereas the C-terminus is indispensable for its RNA chaperoning activity. By promoting RNA template recycling, 2CATPase facilitated EV71 RNA synthesis in vitro; when 2CATPase helicase activity was impaired, EV71 RNA replication and virion production were mostly abolished in cells, indicating that 2CATPase-mediated RNA remodeling plays a critical role in the enteroviral life cycle. Furthermore, the RNA helicase and chaperoning activities of 2CATPase are also conserved in coxsackie A virus 16 (CAV16, another important enterovirus. Altogether, our findings are the first to demonstrate the RNA helicase and chaperoning activities associated with enterovirus 2CATPase, and our study provides both in vitro and cellular evidence for their potential roles during viral RNA replication. These findings

  16. Hepatitis C virus RNA functionally sequesters miR-122

    DEFF Research Database (Denmark)

    Luna, Joseph M; Scheel, Troels K H; Danino, Tal

    2015-01-01

    Hepatitis C virus (HCV) uniquely requires the liver-specific microRNA-122 for replication, yet global effects on endogenous miRNA targets during infection are unexplored. Here, high-throughput sequencing and crosslinking immunoprecipitation (HITS-CLIP) experiments of human Argonaute (AGO) during...... HCV infection showed robust AGO binding on the HCV 5'UTR at known and predicted miR-122 sites. On the human transcriptome, we observed reduced AGO binding and functional mRNA de-repression of miR-122 targets during virus infection. This miR-122 "sponge" effect was relieved and redirected to miR-15...... targets by swapping the miRNA tropism of the virus. Single-cell expression data from reporters containing miR-122 sites showed significant de-repression during HCV infection depending on expression level and site number. We describe a quantitative mathematical model of HCV-induced miR-122 sequestration...

  17. How to quantify knee function after total knee arthroplasty?

    NARCIS (Netherlands)

    Boonstra, M.C.; Waal Malefijt, M.C. de; Verdonschot, N.J.J.

    2008-01-01

    Total knee arthroplasty (TKA) is being undertaken in a younger population than before and as a result the functional demands on the knee are likely to be increasing. As a consequence, it is important to define quantitative functional knee tests that can monitor any increase. A valuable functional

  18. MicroRNA functional network in pancreatic cancer: From biology to ...

    Indian Academy of Sciences (India)

    malignancy. [Wang J and Sen S 2011 MicroRNA functional network in pancreatic cancer: From biology to biomarkers of disease. J. Biosci. 36 481–491] ... aid in improving clinical management and therapeutic outcome for the patients. ..... 133a is a characteristic of pancreatic tissue and that a total of. 26 miRs are aberrantly ...

  19. Reconstruction and analysis of the lncRNA-miRNA-mRNA network based on competitive endogenous RNA reveal functional lncRNAs in rheumatoid arthritis.

    Science.gov (United States)

    Jiang, Hui; Ma, Rong; Zou, Shubiao; Wang, Yongzhong; Li, Zhuqing; Li, Weiping

    2017-06-01

    Rheumatoid arthritis (RA) is an autoimmune disease with an unknown etiology, occurring in approximately 1.0% of general population. More and more studies have suggested that long non-coding RNAs (lncRNAs) could play important roles in various biological processes and be associated with the pathogenesis of different kinds of diseases including RA. Although a large number of lncRNAs have been found, our knowledge of their function and physiological/pathological significance is still in its infancy. In order to reveal functional lncRNAs and identify the key lncRNAs in RA, we reconstructed a global triple network based on the competitive endogenous RNA (ceRNA) theory using the data from National Center for Biotechnology Information Gene Expression Omnibus and our previous paper. Meanwhile, Gene Ontology (GO) and pathway analysis were performed using Cytoscape plug-in BinGO and Database for Annotation, Visualization, and Integration Discovery (DAVID), respectively. We found that the lncRNA-miRNA-mRNA network was composed of 7 lncRNA nodes, 90 mRNA nodes, 24 miRNA nodes, and 301 edges. The functional assay showed that 147 GO terms and 23 pathways were enriched. In addition, three lncRNAs (S5645.1, XR_006437.1, J01878) were highly related to RA, and therefore, were selected as key lncRNAs. This study suggests that specific lncRNAs are associated with the development of RA, and three lncRNAs (S5645.1, XR_006437.1, J01878) could be used as potential diagnostic biomarkers and therapeutic targets.

  20. Structure of noncoding RNA is a determinant of function of RNA binding proteins in transcriptional regulation

    Directory of Open Access Journals (Sweden)

    Oyoshi Takanori

    2012-01-01

    Full Text Available Abstract The majority of the noncoding regions of mammalian genomes have been found to be transcribed to generate noncoding RNAs (ncRNAs, resulting in intense interest in their biological roles. During the past decade, numerous ncRNAs and aptamers have been identified as regulators of transcription. 6S RNA, first described as a ncRNA in E. coli, mimics an open promoter structure, which has a large bulge with two hairpin/stalk structures that regulate transcription through interactions with RNA polymerase. B2 RNA, which has stem-loops and unstructured single-stranded regions, represses transcription of mRNA in response to various stresses, including heat shock in mouse cells. The interaction of TLS (translocated in liposarcoma with CBP/p300 was induced by ncRNAs that bind to TLS, and this in turn results in inhibition of CBP/p300 histone acetyltransferase (HAT activity in human cells. Transcription regulator EWS (Ewing's sarcoma, which is highly related to TLS, and TLS specifically bind to G-quadruplex structures in vitro. The carboxy terminus containing the Arg-Gly-Gly (RGG repeat domains in these proteins are necessary for cis-repression of transcription activation and HAT activity by the N-terminal glutamine-rich domain. Especially, the RGG domain in the carboxy terminus of EWS is important for the G-quadruplex specific binding. Together, these data suggest that functions of EWS and TLS are modulated by specific structures of ncRNAs.

  1. Biosynthesis and functions of sulfur modifications in tRNA

    Directory of Open Access Journals (Sweden)

    Naoki eShigi

    2014-04-01

    Full Text Available Sulfur is an essential element for a variety of cellular constituents in all living organisms. In tRNA molecules, there are many sulfur-containing nucleosides, such as the derivatives of 2‑thiouridine (s2U, 4-thiouridine (s4U, 2-thiocytidine (s2C, and 2-methylthioadenosine (ms2A. Earlier studies established the functions of these modifications for accurate and efficient translation, including proper recognition of the codons in mRNA or stabilization of tRNA structure. In many cases, the biosynthesis of these sulfur modifications starts with cysteine desulfurases, which catalyze the generation of persulfide (an activated form of sulfur from cysteine. Many sulfur-carrier proteins are responsible for delivering this activated sulfur to each biosynthesis pathway. Finally, specific modification enzymes activate target tRNAs and then incorporate sulfur atoms. Intriguingly, the biosynthesis of 2-thiouridine in all domains of life is functionally and evolutionarily related to the ubiquitin-like post-translational modification system of cellular proteins in eukaryotes. This review summarizes the recent characterization of the biosynthesis of sulfur modifications in tRNA and the novel roles of this modification in cellular functions in various model organisms, with a special emphasis on 2-thiouridine derivatives. Each biosynthesis pathway of sulfur-containing molecules is mutually modulated via sulfur trafficking, and 2-thiouridine and codon usage bias have been proposed to control the translation of specific genes.

  2. KLE Translog production function and total factor productivity

    Czech Academy of Sciences Publication Activity Database

    Klacek, J.; Vošvrda, Miloslav; Schlosser, Š.

    2007-01-01

    Roč. 87, č. 4 (2007), s. 261-274 ISSN 0322-788X Institutional research plan: CEZ:AV0Z10750506 Keywords : total factor productivity * translog production function * aggregate production function * least squares method * ridge regression Subject RIV: AH - Economics

  3. Extraction of total RNA from leaves of Eucalyptus and other woody and herbaceous plants using sodium isoascorbate.

    Science.gov (United States)

    Suzuki, Y; Hibino, T; Kawazu, T; Wada, T; Kihara, T; Koyama, H

    2003-05-01

    Rapid extraction of total RNA from Eucalyptus leaves is difficult due to the high content of polyphenolics and polysaccharides. A rapid and simple method was developed by using an extraction buffer containing sodium isoascorbate at a concentration of 500 mM. This method consisted of one or two chloroform extractions, one acid guanidium-phenol-chloroform extraction, and isopropanol precipitation alone. The yields of the RNA fractions were 246-1750 micrograms/g fresh weight when leaves of Eucalyptus, five other woody plants, and four herbaceous plants were used as samples. The contamination of the RNA fractions by proteins and polysaccharides was very limited as judged spectrophotometrically. When the RNA fractions were subjected to agarose gel electrophoresis, intact rRNA bands were detected. The RNA fractions could be used for RT-PCR. These results indicate that our new method achieves a simple and rapid preparation of high-quality RNA from leaves of Eucalyptus and other plant species.

  4. Recovery of physical functioning after total hip arthroplasty

    OpenAIRE

    Heiberg, Kristi Elisabeth

    2014-01-01

    Purpose. The overall aim of this thesis was to examine recovery of physical functioning in patients with hip osteoarthritis (OA) during the first year after total hip arthroplasty (THA). The specific aims were 1) to examine the desires of a group of patients regarding improvements in physical functioning before they underwent THA and at three and 12 months after surgery, 2) to examine changes in physical functioning during the first year of recovery and examine which preoperative measures pre...

  5. Pelvic floor functional outcomes after total abdominal vs total laparoscopic hysterectomy for endometrial cancer.

    Science.gov (United States)

    Higgs, Peta; Janda, Monika; Asher, Rebecca; Gebski, Val; Forder, Peta; Obermair, Andreas

    2018-01-02

    Pelvic floor functioning is an important concern for women requiring a hysterectomy for endometrial cancer. The incidence of pelvic floor symptoms has not been reported in women who have undergone a hysterectomy for early-stage endometrial cancer. We sought to evaluate pelvic floor function in women who have had surgical treatment for early-stage endometrial cancer as part of the multinational Laparoscopic Approach to Cancer of the Endometrium trial and to compare patients' outcomes who had total abdominal vs total laparoscopic hysterectomy. A multinational, phase III, randomized noninferiority trial compared disease-free survival of patients who had total abdominal hysterectomy vs total laparoscopic hysterectomy. This substudy analyzes the results from a self-administered validated questionnaire on pelvic floor symptoms (Pelvic Floor Distress Inventory) administered preoperatively, and at follow-up visits 6, 18, 30, 42, and 54 months postoperatively. Overall, 381 patients with endometrial cancer were included in the analysis (total abdominal hysterectomy, n = 195; total laparoscopic hysterectomy, n = 186). At 6 months postsurgery both groups experienced an improvement in Pelvic Floor Distress Inventory scores compared to presurgical pelvic floor well-being (total abdominal hysterectomy: mean change -11.17; 95% confidence interval, -17.11 to -5.24; total laparoscopic hysterectomy: mean change -10.25; 95% confidence interval, -16.31 to -4.19). The magnitude of change from baseline in pelvic floor symptoms did not differ between both treatment groups up to 54 months postsurgery. These findings suggest that pelvic floor function in terms of urinary, bowel, and prolapse symptoms are unlikely to deteriorate following abdominal or laparoscopic hysterectomy and are reassuring for women undergoing hysterectomy for early-stage endometrial cancer. Crown Copyright © 2017. Published by Elsevier Inc. All rights reserved.

  6. Expression and Functional Role of Reprogramming-Related Long Noncoding RNA (lincRNA-ROR) in Glioma.

    Science.gov (United States)

    Feng, Shiyu; Yao, Jie; Chen, Yang; Geng, Peiliang; Zhang, Haibo; Ma, Xiaodong; Zhao, Jing; Yu, Xinguang

    2015-07-01

    The objective of the study was to investigate the expression and function of reprogramming-related long noncoding RNA (lincRNA-ROR) in glioma and glioma stem cells (GSCs). With real-time quantitative PCR, we analyzed lincRNA-ROR expression levels in 26 primary glioma patients and the expression correlation of lincRNA-ROR with SOX11 and KLF4. To explore its functional role, gain- and loss-of-function studies were performed to assess the effect of lincRNA-ROR on cell proliferation, expression rate of GSCs marker CD133, and glioma stem sphere-forming ability in vitro. We found that the lincRNA-ROR expression was significantly lower in glioma tissues than in adjacent normal tissues. Knockdown of lincRNA-ROR expression by small hairpin RNA (shRNA) significantly elevated the cell proliferation and enhanced the CD133 expression rate and glioma stem sphere-forming ability in U87 cells, while overexpression of lincRNA-ROR in U87 cells showed the opposite effect. Moreover, we found that the expression of lincRNA-ROR was negatively correlated with stem cell factor KLF4 and the "up- and down-regulation" of lincRNA-ROR resulted in inverse modulation of KLF4 messenger RNA (mRNA) expression. Our results suggest that the reprogramming-related lincRNA-ROR may serve as a novel tumor suppressor gene in glioma, which can inhibit the proliferation of cancer cell and self-renewal of GSCs, partly by inhibiting the KLF4 expression. Further research about lincRNA-ROR may provide a novel biomarker and therapeutic target of glioma for cancer clinic in future.

  7. Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs.

    Science.gov (United States)

    Hayashi, Tetsutaro; Ozaki, Haruka; Sasagawa, Yohei; Umeda, Mana; Danno, Hiroki; Nikaido, Itoshi

    2018-02-12

    Total RNA sequencing has been used to reveal poly(A) and non-poly(A) RNA expression, RNA processing and enhancer activity. To date, no method for full-length total RNA sequencing of single cells has been developed despite the potential of this technology for single-cell biology. Here we describe random displacement amplification sequencing (RamDA-seq), the first full-length total RNA-sequencing method for single cells. Compared with other methods, RamDA-seq shows high sensitivity to non-poly(A) RNA and near-complete full-length transcript coverage. Using RamDA-seq with differentiation time course samples of mouse embryonic stem cells, we reveal hundreds of dynamically regulated non-poly(A) transcripts, including histone transcripts and long noncoding RNA Neat1. Moreover, RamDA-seq profiles recursive splicing in >300-kb introns. RamDA-seq also detects enhancer RNAs and their cell type-specific activity in single cells. Taken together, we demonstrate that RamDA-seq could help investigate the dynamics of gene expression, RNA-processing events and transcriptional regulation in single cells.

  8. NUMERICAL SIMULATION OF DIGITAL VLSI TOTAL DOSE FUNCTIONAL FAILURES

    Directory of Open Access Journals (Sweden)

    O. A. Kalashnikov

    2016-10-01

    Full Text Available The technique for numerical simulation of digital VLSI total dose failures is presented, based on fuzzy logic sets theory. It assumes transfer from boolean logic model of a VLSI with values {0,1} to fuzzy model with continuous interval [0,1], and from boolean logic functions to continuous minimax functions. The technique is realized as a calculation system and allows effective estimating of digital VLSI radiation behavior without experimental investigation.

  9. Intragraft interleukin 2 mRNA expression during acute cellular rejection and left ventricular total wall thickness after heart transplantation

    NARCIS (Netherlands)

    H.A. de Groot-Kruseman; C.C. Baan (Carla); E.M. Hagman; W.M. Mol (Wendy); H.G.M. Niesters (Bert); P.E. Zondervan (Pieter); W. Weimar (Willem); A.H.M.M. Balk (Aggie); A.W.P.M. Maat (Alex)

    2002-01-01

    textabstractOBJECTIVE: To assess whether diastolic graft function is influenced by intragraft interleukin 2 (IL-2) messenger RNA (mRNA) expression in rejecting cardiac allografts. DESIGN: 16 recipients of cardiac allografts were monitored during the first three months after

  10. Intragraft interleukin 2 mRNA expression during acute cellular rejection and left ventricular total wall thickness after heart transplantation

    NARCIS (Netherlands)

    de Groot-Kruseman, H A; Baan, C C; Hagman, E M; Mol, W M; Niesters, H G; Maat, A P; Zondervan, P E; Weimar, W; Balk, A H

    OBJECTIVE: To assess whether diastolic graft function is influenced by intragraft interleukin 2 (IL-2) messenger RNA (mRNA) expression in rejecting cardiac allografts. DESIGN: 16 recipients of cardiac allografts were monitored during the first three months after transplantation. The presence of IL-2

  11. RNA Secondary Structure Modulates FMRP’s Bi-Functional Role in the MicroRNA Pathway

    Directory of Open Access Journals (Sweden)

    Phillip Kenny

    2016-06-01

    Full Text Available MicroRNAs act by post-transcriptionally regulating the gene expression of 30%–60% of mammalian genomes. MicroRNAs are key regulators in all cellular processes, though the mechanism by which the cell activates or represses microRNA-mediated translational regulation is poorly understood. In this review, we discuss the RNA binding protein Fragile X Mental Retardation Protein (FMRP and its role in microRNA-mediated translational regulation. Historically, FMRP is known to function as a translational suppressor. However, emerging data suggests that FMRP has both an agonistic and antagonistic role in regulating microRNA-mediated translational suppression. This bi-functional role is dependent on FMRP’s interaction with the RNA helicase Moloney leukemia virus 10 (MOV10, which modifies the structural landscape of bound mRNA, therefore facilitating or inhibiting its association with the RNA-Induced Silencing Complex.

  12. A locally adapted functional outcome measurement score for total ...

    African Journals Online (AJOL)

    Background: Functional outcome scores are often used to measure results of Total Hip Replacement (THR). Most current scoring systems were designed for use in Europe or North America and seem not optimally suited for a general West African setting. We introduce a cross-cultural adaptation of the Lequesne index as a ...

  13. Communication, functional disorders and lifestyle changes after total laryngectomy

    NARCIS (Netherlands)

    Ackerstaff, A. H.; Hilgers, F. J.; Aaronson, N. K.; Balm, A. J.

    1994-01-01

    Functional changes after total laryngectomy, including voice quality, hyposmia and dysgeusia, nasal discharge, swallowing and smoking habits were studied by means of a structured interview with 63 laryngectomized patients. Eighty per cent of the patients reported that they were satisfied with the

  14. Efficient extraction of small and large RNAs in bacteria for excellent total RNA sequencing and comprehensive transcriptome analysis.

    Science.gov (United States)

    Heera, Rajandas; Sivachandran, Parimannan; Chinni, Suresh V; Mason, Joanne; Croft, Larry; Ravichandran, Manickam; Yin, Lee Su

    2015-12-08

    Next-generation transcriptome sequencing (RNA-Seq) has become the standard practice for studying gene splicing, mutations and changes in gene expression to obtain valuable, accurate biological conclusions. However, obtaining good sequencing coverage and depth to study these is impeded by the difficulties of obtaining high quality total RNA with minimal genomic DNA contamination. With this in mind, we evaluated the performance of Phenol-free total RNA purification kit (Amresco) in comparison with TRI Reagent (MRC) and RNeasy Mini (Qiagen) for the extraction of total RNA of Pseudomonas aeruginosa which was grown in glucose-supplemented (control) and polyethylene-supplemented (growth-limiting condition) minimal medium. All three extraction methods were coupled with an in-house DNase I treatment before the yield, integrity and size distribution of the purified RNA were assessed. RNA samples extracted with the best extraction kit were then sequenced using the Illumina HiSeq 2000 platform. TRI Reagent gave the lowest yield enriched with small RNAs (sRNAs), while RNeasy gave moderate yield of good quality RNA with trace amounts of sRNAs. The Phenol-free kit, on the other hand, gave the highest yield and the best quality RNA (RIN value of 9.85 ± 0.3) with good amounts of sRNAs. Subsequent bioinformatic analysis of the sequencing data revealed that 5435 coding genes, 452 sRNAs and 7 potential novel intergenic sRNAs were detected, indicating excellent sequencing coverage across RNA size ranges. In addition, detection of low abundance transcripts and consistency of their expression profiles across replicates from the same conditions demonstrated the reproducibility of the RNA extraction technique. Amresco's Phenol-free Total RNA purification kit coupled with DNase I treatment yielded the highest quality RNAs containing good ratios of high and low molecular weight transcripts with minimal genomic DNA. These RNA extracts gave excellent non-biased sequencing coverage useful

  15. Total Correlation Function Integrals and Isothermal Compressibilities from Molecular Simulations

    DEFF Research Database (Denmark)

    Wedberg, Rasmus; Peters, Günther H.j.; Abildskov, Jens

    2008-01-01

    Generation of thermodynamic data, here compressed liquid density and isothermal compressibility data, using molecular dynamics simulations is investigated. Five normal alkane systems are simulated at three different state points. We compare two main approaches to isothermal compressibilities: (1......) fluctuation solution theory analysis of trajectories obtained from simulations to yield total correlation function integrals; and (2) the more commonly used fluctuation formula. The results show that the two approaches yield consistent values and consistent uncertainties. Also, the computations converge...

  16. Essential nontranslational functions of tRNA synthetases.

    Science.gov (United States)

    Guo, Min; Schimmel, Paul

    2013-03-01

    Nontranslational functions of vertebrate aminoacyl tRNA synthetases (aaRSs), which catalyze the production of aminoacyl-tRNAs for protein synthesis, have recently been discovered. Although these new functions were thought to be 'moonlighting activities', many are as critical for cellular homeostasis as their activity in translation. New roles have been associated with their cytoplasmic forms as well as with nuclear and secreted extracellular forms that affect pathways for cardiovascular development and the immune response and mTOR, IFN-γ and p53 signaling. The associations of aaRSs with autoimmune disorders, cancers and neurological disorders further highlight nontranslational functions of these proteins. New architecture elaborations of the aaRSs accompany their functional expansion in higher organisms and have been associated with the nontranslational functions for several aaRSs. Although a general understanding of how these functions developed is limited, the expropriation of aaRSs for essential nontranslational functions may have been initiated by co-opting the amino acid-binding site for another purpose.

  17. Essential Non-Translational Functions of tRNA Synthetases

    Science.gov (United States)

    Guo, Min; Schimmel, Paul

    2013-01-01

    Nontranslational functions of vertebrate aminoacyl tRNA synthetases (aaRSs), which catalyze the production of aminoacyl-tRNAs for protein synthesis, have recently been discovered. While these new functions were thought to be ‘moonlighting activities’, many are as critical for cellular homeostasis as the activity in translation. New roles have been associated with cytoplasmic forms as well as with nuclear and secreted extracellular forms that impact pathways for cardiovascular development, the immune response, and mTOR, IFN-γ and p53 signaling. The associations of aaRSs with autoimmune disorders, cancers and neurological disorders further highlight nontranslational functions of these proteins. Novel architecture elaborations of the aaRSs accompany their functional expansion in higher organisms and have been associated with the nontranslational functions for several aaRSs. While a general understanding of how these functions developed is limited, the expropriation of aaRSs for essential nontranslational functions may have been initiated by co-opting the amino acid binding site for another purpose. PMID:23416400

  18. Sexual Function before and after Total Hip Replacement: Narrative Review

    Directory of Open Access Journals (Sweden)

    Rotem Meiri, BPT

    2014-12-01

    Conclusions: The ability to move comfortably is included among the many physical and psychosocial factors influencing sexual functioning. Practitioners should be encouraged to question their THR patients about sexual concerns and to provide counseling related to physical and functional aspects of sexual activity. Rehabilitation that focuses specifically on activities of daily living of sex should include sexual counseling, therapeutic exercise, and advice regarding sexual positions. Rehabilitation provided by physical therapists may help decrease pain, and facilitate greater self‐awareness, self‐confidence, and improved body image, all of which encourage and affirm optimal sexual health. Meiri R, Rosenbaum TY, and Kalichman L. Sexual function before and after total hip replacement: Narrative review. Sex Med 2014;2:159–167.

  19. De novo RNA-Seq and functional annotation of Sarcoptes scabiei canis.

    Science.gov (United States)

    Hu, Li; Zhao, YaE; Yang, YuanJun; Niu, DongLing; Wang, RuiLing; Cheng, Juan; Yang, Fan

    2016-07-01

    The transcriptomic data of Sarcoptes is still lacking in the public database due to the difficulty in extracting high-quality RNA from tiny mites with thick chitin. In this study, total RNA was extracted from live Sarcoptes mites for quality assessment, RNA-Seq, functional annotation, and coding region (CD) prediction and verification. The results showed that the sample JMQ-lngm was qualified for cDNA library construction. Firstly, Agilent 2100 detection showed that the RNA baseline was smooth and the 18S peak was single. Second, the Illumina platform generated 65.78M clean reads and 20,826 unigenes with 35.43M were assembled, occupying 62.98 % of the 56.26M genome. In total, 15,034 unigenes were annotated in seven functional databases. Finally, 13,122 CDs were detected in the 20,826 unigenes, of which 70 complete CDs were matched with Sarcoptes manually in non-redundant nucleotide (NT). Three CDs with indels ≥10 bp were verified. Those results indicated that peritrophin sequences of JMQ-lngm missed 35 bp during the assembly; the pressure-sensitive sodium channel sequences of all the six Sarcoptes scabiei canis isolates were confirmed to be 90 bp shorter than that of a Sarcoptes scabiei hominis isolate; three introns remained in PH chlorine ion channel gating sequences of JMQ-lngm. Moreover, the allergen gene prediction for JMQ-lngm indicated that 61 unigenes were matched with 19 allergen genes of Dermatophagoides, of which Der 1, Der 3, Der 8, and Der 10 had been confirmed in NT. In conclusion, this study successfully completed the RNA-Seq and functional annotation of S. s. canis for the first time, which provides molecular data for future studies on the identification and pathogenic genes of Sarcoptidae.

  20. Establishment of the total RNA extraction system for lily bulbs with ...

    African Journals Online (AJOL)

    NaCl concentration of both CTAB and SDS protocols was increased in order to get rid of the polysaccharides in scales. The resulting RNA is of high-quality, as judged by agarose gel electrophoresis and UV-spectroscopic analysis. Furthermore, the middle scales obtain the highest RNA yield followed by the exterior and the ...

  1. Functional rehabilitation after total hip arthroplasty with uncemented prosthesis

    Directory of Open Access Journals (Sweden)

    Nicolae-Bogdan Negru-Aman

    2011-12-01

    Full Text Available Arthritis is a disease that acts irreversibly on joint surfaces with significant consequences especially in the third quarter of life. About 7% of our population is affected by arthritis localized at a certain level and stage of development, and this percentage is expected to reach a much higher value in future years. Aim: The aim of this study is to propose a rehabilitation program for functional recovery after total hip arthroplasty and a set of recommendations for post surgery period. Material and methods: There were included 13 subjects (10 female and 3 male aged between 51 and 78 years old, ready for total hip arthroplasty with uncemented prosthesis. The subjects were selected in Orthopaedics and Traumatology department of Military Clinical Emergency Hospital ”Dr. Victor Popescu” Timisoara. The evaluations regarding programs’ efficiency were made using 3 questionnaires (Oxford Hip Score, Harris Hip Score, Outcome Hip Score and goniometry. The subjects were tested initially before the surgery and 3 times after (at one month, at 3 months and at 6 months. Results: The results showed a remarcable increase for the scores of majority, in both questionnaire and goniometry values. Conclusions: functional rehabilitation exercises proved to be particularly important in regaining independence, control of the prosthetic leg, pain reduction and functional and social reintegration.

  2. Exploring Connectivity in Sequence Space of Functional RNA

    Science.gov (United States)

    Wei, Chenyu; Pohorille, Andrzej; Popovic, Milena; Ditzler, Mark

    2017-01-01

    Emergence of replicable genetic molecules was one of the marking points in the origin of life, evolution of which can be conceptualized as a walk through the space of all possible sequences. A theoretical concept of fitness landscape helps to understand evolutionary processes through assigning a value of fitness to each genotype. Then, evolution of a phenotype is viewed as a series of consecutive, single-point mutations. Natural selection biases evolution toward peaks of high fitness and away from valleys of low fitness. whereas neutral drift occurs in the sequence space without direction as mutations are introduced at random. Large networks of neutral or near-neutral mutations on a fitness landscape, especially for sufficiently long genomes, are possible or even inevitable. Their detection in experiments, however, has been elusive. Although a few near-neutral evolutionary pathways have been found, recent experimental evidence indicates landscapes consist of largely isolated islands. The generality of these results, however, is not clear, as the genome length or the fraction of functional molecules in the genotypic space might have been insufficient for the emergence of large, neutral networks. Thorough investigation on the structure of the fitness landscape is essential to understand the mechanisms of evolution of early genomes. RNA molecules are commonly assumed to play the pivotal role in the origin of genetic systems. They are widely believed to be early, if not the earliest, genetic and catalytic molecules, with abundant biochemical activities as aptamers and ribozymes, i.e. RNA molecules capable, respectively, to bind small molecules or catalyze chemical reactions. Here, we present results of our recent studies on the structure of the sequence space of RNA ligase ribozymes selected through in vitro evolution. Several hundred thousands of sequences active to a different degree were obtained by way of deep sequencing. Analysis of these sequences revealed

  3. miRNA-mediated functional changes through co-regulating function related genes.

    Directory of Open Access Journals (Sweden)

    Jie He

    Full Text Available BACKGROUND: MicroRNAs play important roles in various biological processes involving fairly complex mechanism. Analysis of genome-wide miRNA microarray demonstrate that a single miRNA can regulate hundreds of genes, but the regulative extent on most individual genes is surprisingly mild so that it is difficult to understand how a miRNA provokes detectable functional changes with such mild regulation. RESULTS: To explore the internal mechanism of miRNA-mediated regulation, we re-analyzed the data collected from genome-wide miRNA microarray with bioinformatics assay, and found that the transfection of miR-181b and miR-34a in Hela and HCT-116 tumor cells regulated large numbers of genes, among which, the genes related to cell growth and cell death demonstrated high Enrichment scores, suggesting that these miRNAs may be important in cell growth and cell death. MiR-181b induced changes in protein expression of most genes that were seemingly related to enhancing cell growth and decreasing cell death, while miR-34a mediated contrary changes of gene expression. Cell growth assays further confirmed this finding. In further study on miR-20b-mediated osteogenesis in hMSCs, miR-20b was found to enhance osteogenesis by activating BMPs/Runx2 signaling pathway in several stages by co-repressing of PPARγ, Bambi and Crim1. CONCLUSIONS: With its multi-target characteristics, miR-181b, miR-34a and miR-20b provoked detectable functional changes by co-regulating functionally-related gene groups or several genes in the same signaling pathway, and thus mild regulation from individual miRNA targeting genes could have contributed to an additive effect. This might also be one of the modes of miRNA-mediated gene regulation.

  4. Deploying RNA and DNA with Functionalized Carbon Nanotubes

    Science.gov (United States)

    Alidori, Simone; Asqiriba, Karim; Londero, Pablo; Bergkvist, Magnus; Leona, Marco; Scheinberg, David A.; McDevitt, Michael R.

    2013-01-01

    Carbon nanotubes internalize into cells and are potential molecular platforms for siRNA and DNA delivery. A comprehensive understanding of the identity and stability of ammoniumfunctionalized carbon nanotube (f-CNT)-based nucleic acid constructs is critical to deploying them in vivo as gene delivery vehicles. This work explored the capability of f-CNT to bind single- and double-strand oligonucleotides by determining the thermodynamics and kinetics of assembly and the stoichiometric composition in aqueous solution. Surprisingly, the binding affinity of f-CNT and short oligonucleotide sequences was in the nanomolar range, kinetics of complexation were extremely rapid, and from one to five sequences were loaded per nanotube platform. Mechanistic evidence for an assembly process that involved electrostatic, hydrogen-bonding and π-stacking bonding interactions was obtained by varying nanotube functionalities, oligonucleotides, and reaction conditions. 31P-NMR and spectrophotometric fluorescence emission data described the conditions required to assemble and stably bind a DNA or RNA cargo for delivery in vivo and the amount of oligonucleotide that could be transported. The soluble oligonucleic acid-f-CNT supramolecular assemblies were suitable for use in vivo. Importantly, key evidence in support of an elegant mechanism by which the bound nucleic acid material can be ‘off-loaded’ from the f-CNT was discovered. PMID:23626864

  5. Parathyroid function following total thyroidectomy using energy devices.

    Science.gov (United States)

    Ciftci, Fatih; Sakalli, Erdal; Abdurrahman, Ibrahim; Guler, Burak

    2016-07-01

    LigaSure precise (LP) and harmonic scalpel (HS) are two energy-based devices used in thyroidectomy surgery. We aimed to compare the effect of these two devices in patients who had undergone total thyroidectomy, by highlighting the post-operative parathyroid function. A total of 201 consecutive patients for whom total thyroidectomy had been planned were prospectively classified into two groups. There were 104 patients in LP group and 97 patients in HS group. Hypoparathyroidism was followed up by serially measuring the levels of intact parathyroid hormone (iPTH) and serum calcium. The early iPTH level was 29 (8-50) pg/mL in group LP, and 17 (4-43) pg/mL in group HS. The early iPTH level was significantly lower in the HS group (p parathyroid hormone levels impaired in the HS group more than LP group in the early period, these levels were in the normal limits in both groups. These levels were not significant between groups in the long time period.

  6. SmD1 Modulates the miRNA Pathway Independently of Its Pre-mRNA Splicing Function.

    Directory of Open Access Journals (Sweden)

    Xiao-Peng Xiong

    2015-08-01

    Full Text Available microRNAs (miRNAs are a class of endogenous regulatory RNAs that play a key role in myriad biological processes. Upon transcription, primary miRNA transcripts are sequentially processed by Drosha and Dicer ribonucleases into ~22-24 nt miRNAs. Subsequently, miRNAs are incorporated into the RNA-induced silencing complexes (RISCs that contain Argonaute (AGO family proteins and guide RISC to target RNAs via complementary base pairing, leading to post-transcriptional gene silencing by a combination of translation inhibition and mRNA destabilization. Select pre-mRNA splicing factors have been implicated in small RNA-mediated gene silencing pathways in fission yeast, worms, flies and mammals, but the underlying molecular mechanisms are not well understood. Here, we show that SmD1, a core component of the Drosophila small nuclear ribonucleoprotein particle (snRNP implicated in splicing, is required for miRNA biogenesis and function. SmD1 interacts with both the microprocessor component Pasha and pri-miRNAs, and is indispensable for optimal miRNA biogenesis. Depletion of SmD1 impairs the assembly and function of the miRISC without significantly affecting the expression of major canonical miRNA pathway components. Moreover, SmD1 physically and functionally associates with components of the miRISC, including AGO1 and GW182. Notably, miRNA defects resulting from SmD1 silencing can be uncoupled from defects in pre-mRNA splicing, and the miRNA and splicing machineries are physically and functionally distinct entities. Finally, photoactivatable-ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP analysis identifies numerous SmD1-binding events across the transcriptome and reveals direct SmD1-miRNA interactions. Our study suggests that SmD1 plays a direct role in miRNA-mediated gene silencing independently of its pre-mRNA splicing activity and indicates that the dual roles of splicing factors in post-transcriptional gene regulation may be

  7. Total Variation Regularization for Functions with Values in a Manifold

    KAUST Repository

    Lellmann, Jan

    2013-12-01

    While total variation is among the most popular regularizers for variational problems, its extension to functions with values in a manifold is an open problem. In this paper, we propose the first algorithm to solve such problems which applies to arbitrary Riemannian manifolds. The key idea is to reformulate the variational problem as a multilabel optimization problem with an infinite number of labels. This leads to a hard optimization problem which can be approximately solved using convex relaxation techniques. The framework can be easily adapted to different manifolds including spheres and three-dimensional rotations, and allows to obtain accurate solutions even with a relatively coarse discretization. With numerous examples we demonstrate that the proposed framework can be applied to variational models that incorporate chromaticity values, normal fields, or camera trajectories. © 2013 IEEE.

  8. Isolation of biologically functional RNA during programmed death of a colonial ascidian.

    Science.gov (United States)

    Chang, W T; Lauzon, R J

    1995-01-01

    The blastogenic (asexual) cycle of the colonial ascidian Botryllus schlosseri (Tunicata, Ascidiaceae) concludes in a cyclical phase of programmed cell and zooid death called takeover, in which all asexually derived adults die synchronously by apoptosis. The characterization of developmentally regulated genes whose expression patterns are selectively modulated during this process could pave the way to understand how this model organism dies. However, isolation of biologically functional RNA in this and other colonial ascidians with conventional phenol/chloroform-based procedures is hampered by extensive contamination of RNA preparations by pigments. Upon cell lysis, pigments that normally reside within specialized cells in the mantle wall of the adult are released and tightly associate with nucleic acids. Here, we report on the usefulness of a single-step RNA isolation method in which acid guanidinium isothiocyanate is used as an extraction medium, followed by preparative cesium chloride ultracentrifugation. This procedure successfully isolated biologically active, high-purity total RNA (OD260/OD280 = 1.9-2.1) from Botryllus colonies during takeover, as well as other species of colonial ascidians (Diplosoma macdonaldii, Botrylloides diegense) irrespective of pigmentation. Northern blot analysis performed with a 32P-labeled tunicate actin probe detected two polyadenylated transcripts of 1.5 and 1.7 kilobases in length from both growth phase and takeover colonies. Two-dimensional protein gel assays from in vitro translated mRNA preparations further revealed that specific transcripts were up-regulated during takeover, while others were repressed or down-regulated. Growth phase and takeover-specific cDNA libraries were constructed from pooled poly(A)+ RNA with a complexity of 1.0 x 10(7) and 1.2 x 10(7) recombinants respectively per 100 ng of cDNA before amplification.(ABSTRACT TRUNCATED AT 250 WORDS)

  9. Non-Watson-Crick RNA synthesis suited to origin functions.

    Science.gov (United States)

    Puthenvedu, Deepa; Majerfeld, Irene; Yarus, Michael

    2018-01-01

    A templated RNA synthesis is characterized in which G 5' pp 5' G accelerates synthesis of A 5' pp 5' A from pA and chemically activated ImpA precursors. Similar acceleration is not observable in the presence of UppU, CppC, AppG, AppA, or pG alone. Thus, it seems likely that AppA is templated by GppG via a form or forms of G:A base-pairing. AppA also appears, more slowly, via a previously known untemplated second-order chemical route. Such AppA synthesis requires only ordinary near-neutral solutions containing monovalent and divalent salts, and rates are only slightly sensitive to variation in pH. Templated synthesis rates are first order in pA, ImpA, and template GppG; thus third order overall. Therefore, this reaction resembles cross-templating of AppA on poly(U), but is notably slower and less sensitive to temperature. Viewing AppA as a coenzyme analog, GppG templating provides a simpler molecular route, termed para-templating, to encoded chemical functions. Para-templating can also arise from a single, localized nucleobase geosynthetic event which yields purines. It requires only a single backbone-forming chemistry. Thus it may have appeared earlier and served as evolutionary precursor for more complex forms of encoded genetic expression. © 2018 Puthenvedu et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  10. Functional gene silencing mediated by chitosan/siRNA nanocomplexes

    Energy Technology Data Exchange (ETDEWEB)

    Ji, A M; Su, D; Che, O; Li, W S; Sun, L; Zhang, Z Y; Xu, F [Department of Pharmaceutical Science, Zhujiang Hospital, Southern Medical University, Guangzhou 510282 (China); Yang, B, E-mail: andrewfxu1998@gmail.co [Department of Chemistry, Indiana University-Bloomington, Bloomington, IN 47405 (United States)

    2009-10-07

    Chitosan/siRNA nanoparticles to knock down FHL2 gene expression were reported in this work. The physicochemical properties such as particle size, surface charge, morphology and complex stability of chitosan nanoparticle-incorporated siRNA were evaluated. Nanoparticles which were formulated with chitosan/siRNA exhibited irregular, lamellar and dendritic structures with a hydrodynamic radius size of about 148 nm and net positive charges with zeta-potential value of 58.5 mV. The knockdown effect of the chitosan/siRNA nanoparticles on gene expression in FHL2 over-expressed human colorectal cancer Lovo cells was investigated. The result showed that FHL2 siRNA formulated within chitosan nanoparticles could knock down about 69.6% FHL2 gene expression, which is very similar to the 68.8% reduced gene expression when siRNA was transfected with liposome Lipofectamine. Western analysis further showed significant FHL-2 protein expression reduced by the chitosan/siRNA nanoparticles. The results also showed that blocking FHL2 expression by siRNA could also inhibit the growth and proliferation of human colorectal cancer Lovo cells. The current results demonstrated that chitosan-based siRNA nanoparticles were a very efficient delivery system for siRNA in vivo as previously reported.

  11. Functional gene silencing mediated by chitosan/siRNA nanocomplexes

    Science.gov (United States)

    Ji, A. M.; Su, D.; Che, O.; Li, W. S.; Sun, L.; Zhang, Z. Y.; Yang, B.; Xu, F.

    2009-10-01

    Chitosan/siRNA nanoparticles to knock down FHL2 gene expression were reported in this work. The physicochemical properties such as particle size, surface charge, morphology and complex stability of chitosan nanoparticle-incorporated siRNA were evaluated. Nanoparticles which were formulated with chitosan/siRNA exhibited irregular, lamellar and dendritic structures with a hydrodynamic radius size of about 148 nm and net positive charges with zeta-potential value of 58.5 mV. The knockdown effect of the chitosan/siRNA nanoparticles on gene expression in FHL2 over-expressed human colorectal cancer Lovo cells was investigated. The result showed that FHL2 siRNA formulated within chitosan nanoparticles could knock down about 69.6% FHL2 gene expression, which is very similar to the 68.8% reduced gene expression when siRNA was transfected with liposome Lipofectamine. Western analysis further showed significant FHL-2 protein expression reduced by the chitosan/siRNA nanoparticles. The results also showed that blocking FHL2 expression by siRNA could also inhibit the growth and proliferation of human colorectal cancer Lovo cells. The current results demonstrated that chitosan-based siRNA nanoparticles were a very efficient delivery system for siRNA in vivo as previously reported.

  12. Functional gene silencing mediated by chitosan/siRNA nanocomplexes

    International Nuclear Information System (INIS)

    Ji, A M; Su, D; Che, O; Li, W S; Sun, L; Zhang, Z Y; Xu, F; Yang, B

    2009-01-01

    Chitosan/siRNA nanoparticles to knock down FHL2 gene expression were reported in this work. The physicochemical properties such as particle size, surface charge, morphology and complex stability of chitosan nanoparticle-incorporated siRNA were evaluated. Nanoparticles which were formulated with chitosan/siRNA exhibited irregular, lamellar and dendritic structures with a hydrodynamic radius size of about 148 nm and net positive charges with zeta-potential value of 58.5 mV. The knockdown effect of the chitosan/siRNA nanoparticles on gene expression in FHL2 over-expressed human colorectal cancer Lovo cells was investigated. The result showed that FHL2 siRNA formulated within chitosan nanoparticles could knock down about 69.6% FHL2 gene expression, which is very similar to the 68.8% reduced gene expression when siRNA was transfected with liposome Lipofectamine. Western analysis further showed significant FHL-2 protein expression reduced by the chitosan/siRNA nanoparticles. The results also showed that blocking FHL2 expression by siRNA could also inhibit the growth and proliferation of human colorectal cancer Lovo cells. The current results demonstrated that chitosan-based siRNA nanoparticles were a very efficient delivery system for siRNA in vivo as previously reported.

  13. The identification and functional annotation of RNA structures conserved in vertebrates

    DEFF Research Database (Denmark)

    Seemann, Ernst Stefan; Mirza, Aashiq Hussain; Hansen, Claus

    2017-01-01

    Structured elements of RNA molecules are essential in, e.g., RNA stabilization, localization and protein interaction, and their conservation across species suggests a common functional role. We computationally screened vertebrate genomes for Conserved RNA Structures (CRSs), leveraging structure-b...

  14. Functional long non-coding RNA transcription in Schizosaccharomyces pombe

    OpenAIRE

    Ard, Ryan Anthony

    2016-01-01

    Eukaryotic genomes are pervasively transcribed and frequently generate long noncoding RNAs (lncRNAs). However, most lncRNAs remain uncharacterized. In this work, a set of positionally conserved intergenic lncRNAs in the fission yeast Schizosaccharomyces pombe genome are selected for further analysis. Deleting one of these lncRNA genes (ncRNA.1343) exhibited a clear phenotype: increased drug sensitivity. Further analyses revealed that deleting ncRNA.1343 also disrupted a prev...

  15. Dynamic foot function changes following total knee replacement surgery.

    Science.gov (United States)

    Levinger, Pazit; Menz, Hylton B; Morrow, Adam D; Bartlett, John R; Feller, Julian A; Fotoohabadi, Mohammad R; Bergman, Neil R

    2012-12-01

    Individuals with knee osteoarthritis (OA) have flatter/more pronated feet than those without OA, but it is unclear whether altered foot posture and function are a cause or consequence of knee OA. The purpose of this study was to examine whether changes in foot posture and function occur after realignment of the knee following total knee replacement (TKR). Nineteen patients with predominantly medial compartment knee OA were tested prior to and 12 months after TKR. The Foot Posture Index (FPI) and Arch Index (AI) were measured as well as motion of the tibia, rearfoot and forefoot using a 3D motion analysis system incorporating a multisegment foot model. There were no significant changes in FPI or AI following TKR, however gait analysis revealed significant increases in tibial external rotation (-18.7 ± 7.0° vs -22.5 ± 8.7°, p=0.002), tibial transverse plane range of motion (-9.1 ± 4.6° vs -11.4 ± 6.1°, p=0.0028) and rearfoot range of motion in the frontal plane (8.6 ± 2.6° vs 10.4 ± 2.7°, p=0.002), and a decrease in rearfoot transverse plane range of motion (8.7 ± 5.3° vs 5.9 ± 4.1°, p=0.038) following the procedure. TKR produces no change in static foot posture, but results in significant changes in rearfoot kinematics during gait. These findings suggest that rearfoot motion compensates for changes in the alignment of the knee, highlighting the ability of the foot to accommodate for proximal skeletal malalignment. Copyright © 2012 Elsevier B.V. All rights reserved.

  16. RNA-Mediated Regulation of HMGA1 Function

    Directory of Open Access Journals (Sweden)

    Arndt G. Benecke

    2015-05-01

    Full Text Available The high mobility group protein A1 (HMGA1 is a master regulator of chromatin structure mediating its major gene regulatory activity by direct interactions with A/T-rich DNA sequences located in the promoter and enhancer regions of a large variety of genes. HMGA1 DNA-binding through three AT-hook motifs results in an open chromatin structure and subsequently leads to changes in gene expression. Apart from its significant expression during development, HMGA1 is over-expressed in virtually every cancer, where HMGA1 expression levels correlate with tumor malignancy. The exogenous overexpression of HMGA1 can lead to malignant cell transformation, assigning the protein a key role during cancerogenesis. Recent studies have unveiled highly specific competitive interactions of HMGA1 with cellular and viral RNAs also through an AT-hook domain of the protein, significantly impacting the HMGA1-dependent gene expression. In this review, we discuss the structure and function of HMGA1-RNA complexes during transcription and epigenomic regulation and their implications in HMGA1-related diseases.

  17. Construction of microRNA functional families by a mixture model of position weight matrices

    Directory of Open Access Journals (Sweden)

    Je-Keun Rhee

    2013-10-01

    Full Text Available MicroRNAs (miRNAs are small regulatory molecules that repress the translational processes of their target genes by binding to their 3′ untranslated regions (3′ UTRs. Because the target genes are predominantly determined by their sequence complementarity to the miRNA seed regions (nucleotides 2–7 which are evolutionarily conserved, it is inferred that the target relationships and functions of the miRNA family members are conserved across many species. Therefore, detecting the relevant miRNA families with confidence would help to clarify the conserved miRNA functions, and elucidate miRNA-mediated biological processes. We present a mixture model of position weight matrices for constructing miRNA functional families. This model systematically finds not only evolutionarily conserved miRNA family members but also functionally related miRNAs, as it simultaneously generates position weight matrices representing the conserved sequences. Using mammalian miRNA sequences, in our experiments, we identified potential miRNA groups characterized by similar sequence patterns that have common functions. We validated our results using score measures and by the analysis of the conserved targets. Our method would provide a way to comprehensively identify conserved miRNA functions.

  18. Simultaneous characterization of cellular RNA structure and function with in-cell SHAPE-Seq.

    Science.gov (United States)

    Watters, Kyle E; Abbott, Timothy R; Lucks, Julius B

    2016-01-29

    Many non-coding RNAs form structures that interact with cellular machinery to control gene expression. A central goal of molecular and synthetic biology is to uncover design principles linking RNA structure to function to understand and engineer this relationship. Here we report a simple, high-throughput method called in-cell SHAPE-Seq that combines in-cell probing of RNA structure with a measurement of gene expression to simultaneously characterize RNA structure and function in bacterial cells. We use in-cell SHAPE-Seq to study the structure-function relationship of two RNA mechanisms that regulate translation in Escherichia coli. We find that nucleotides that participate in RNA-RNA interactions are highly accessible when their binding partner is absent and that changes in RNA structure due to RNA-RNA interactions can be quantitatively correlated to changes in gene expression. We also characterize the cellular structures of three endogenously expressed non-coding RNAs: 5S rRNA, RNase P and the btuB riboswitch. Finally, a comparison between in-cell and in vitro folded RNA structures revealed remarkable similarities for synthetic RNAs, but significant differences for RNAs that participate in complex cellular interactions. Thus, in-cell SHAPE-Seq represents an easily approachable tool for biologists and engineers to uncover relationships between sequence, structure and function of RNAs in the cell. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  19. Total cellular HIV-1 DNA decreases after switching to raltegravir-based regimens in patients with suppressed HIV-1 RNA.

    Science.gov (United States)

    Rossetti, Barbara; Meini, Genny; Bianco, Claudia; Lamonica, Silvia; Mondi, Annalisa; Belmonti, Simone; Fanti, Iuri; Ciccarelli, Nicoletta; Di Giambenedetto, Simona; Zazzi, Maurizio; De Luca, Andrea

    2017-06-01

    The integrase inhibitor raltegravir has been used to intensify antiretroviral therapy in patients with undetectable plasma HIV-1RNA, resulting in variable perturbation of HIV-1 nucleic acids levels in peripheral blood. We aimed at monitoring residual plasma HIV-1RNA and total cellular HIV-1DNA in virologically suppressed patients switching to raltegravir-based regimens. Fifty-eight subjects on protease inhibitor (PI) or nonnucleoside reverse transcriptase inhibitor (NNRTI)-based regimens, with plasma HIV-1RNA levels 200cells/μl for ≥12 months were enrolled. Thirty-four patients were from the treatment simplification RASTA randomized study switching standard therapy to a raltegravir-based regimen (RASTA group), while 24 continued a PI or NNRTI based-regimen (controls). Residual plasma HIV-1RNA (5-40copies/mL) and HIV-1DNA were assessed at 0, 24 and 48 weeks. At week 0 (W0), HIV-1DNA was detected in all patients while at W48 it was detectable in 82.4% of the RASTA group vs 100% of controls (p=0.03). There was a significant decline of HIV-1DNA at W48 in the RASTA group (mean change from baseline -0.21 [95% CI -0.41; -0.01] log 10 copies/10 6 CD4; p=0.03) but not in controls. Ultrasensitive HIV-1RNA was detectable at baseline in 50% of RASTA group vs 67% of controls and at W48 in 32.4% vs 42%, respectively. No differences were found between HIV-1RNA levels at baseline and W48 within and between groups. Switching successful therapy to raltegravir-based regimens may be associated with a decrease of the HIV-1 reservoir, as measured by peripheral blood cellular HIV-1DNA levels. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. A new total distal radioulnar joint prosthesis: functional outcome.

    Science.gov (United States)

    Schuurman, Arnold H; Teunis, Teun

    2010-10-01

    To present the evolution of design and the short-term functional outcome of our distal radioulnar joint (DRUJ) prosthesis. This total DRUJ prosthesis differs from others in that it consists of 2 parts and attains bony fixation by its hydroxyapatite coating. Nineteen patients received a DRUJ prosthesis after a failed Darrach procedure (n = 10), Sauvé-Kapandji procedure (n = 7), trauma (n = 1), or DRUJ synovitis (n = 1). Indications for the placement were decreased grip, decreased forearm motion, and pain due to ulnar impingement syndrome and instability of the distal ulna. Seven prostheses were removed, 5 due to loosening, 1 due to continuing pain, and 1 at the request of the patient. The 5 prostheses that loosened were an intermediate prototype no longer in use. In 12 remaining cases, range of motion, grip strength, and pinch strength were measured, and patients completed the Disabilities of the Arm, Shoulder, and Hand (DASH) questionnaire. Pain was assessed with the visual analog scale (range, 0-10). A paired t-test was performed to assess the significance of the difference between preoperative and postoperative measurements. Statistically significant improvements were seen in forearm pronation, from an average of 79° to 88°; grip strength, from an average of 10 kg to an average of 16 kg; and visual analog scale score, decreased from a mean of 5.3 to a mean of 3.5. The distal ulna was clinically stable in all 12 patients who retained the prosthesis. The intermediate prototype had a high failure rate, 5 out of 5. The early results for the current prosthesis prototype show clinical improvement. Based on these results, we conclude that this prosthesis offers a new treatment option for ulnar instability after distal ulnar resection. Therapeutic IV. Copyright © 2010. Published by Elsevier Inc.

  1. A functional pseudoknot in 16S ribosomal RNA.

    OpenAIRE

    Powers, T; Noller, H F

    1991-01-01

    Several lines of evidence indicate that the universally conserved 530 loop of 16S ribosomal RNA plays a crucial role in translation, related to the binding of tRNA to the ribosomal A site. Based upon limited phylogenetic sequence variation, Woese and Gutell (1989) have proposed that residues 524-526 in the 530 hairpin loop are base paired with residues 505-507 in an adjoining bulge loop, suggesting that this region of 16S rRNA folds into a pseudoknot structure. Here, we demonstrate that Watso...

  2. RNA Polymerase III Output Is Functionally Linked to tRNA Dimethyl-G26 Modification.

    Directory of Open Access Journals (Sweden)

    Aneeshkumar G Arimbasseri

    2015-12-01

    Full Text Available Control of the differential abundance or activity of tRNAs can be important determinants of gene regulation. RNA polymerase (RNAP III synthesizes all tRNAs in eukaryotes and it derepression is associated with cancer. Maf1 is a conserved general repressor of RNAP III under the control of the target of rapamycin (TOR that acts to integrate transcriptional output and protein synthetic demand toward metabolic economy. Studies in budding yeast have indicated that the global tRNA gene activation that occurs with derepression of RNAP III via maf1-deletion is accompanied by a paradoxical loss of tRNA-mediated nonsense suppressor activity, manifested as an antisuppression phenotype, by an unknown mechanism. We show that maf1-antisuppression also occurs in the fission yeast S. pombe amidst general activation of RNAP III. We used tRNA-HydroSeq to document that little changes occurred in the relative levels of different tRNAs in maf1Δ cells. By contrast, the efficiency of N2,N2-dimethyl G26 (m(22G26 modification on certain tRNAs was decreased in response to maf1-deletion and associated with antisuppression, and was validated by other methods. Over-expression of Trm1, which produces m(22G26, reversed maf1-antisuppression. A model that emerges is that competition by increased tRNA levels in maf1Δ cells leads to m(22G26 hypomodification due to limiting Trm1, reducing the activity of suppressor-tRNASerUCA and accounting for antisuppression. Consistent with this, we show that RNAP III mutations associated with hypomyelinating leukodystrophy decrease tRNA transcription, increase m(22G26 efficiency and reverse antisuppression. Extending this more broadly, we show that a decrease in tRNA synthesis by treatment with rapamycin leads to increased m(22G26 modification and that this response is conserved among highly divergent yeasts and human cells.

  3. The MRB1 complex functions in kinetoplastid RNA

    Czech Academy of Sciences Publication Activity Database

    Acestor, N.; Panigrahi, A. K.; Jason, C.; Zíková, Alena; Stuart, K. D.

    2009-01-01

    Roč. 15, č. 2 (2009), s. 277-286 ISSN 1355-8382 Keywords : Trypanosoma * RNA editing * mitochondria * RNAi * protein complex Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.198, year: 2009

  4. Influence of bilateral sequential total knee arthroplasty on functional recovery

    Directory of Open Access Journals (Sweden)

    Rajesh N Maniar

    2013-01-01

    Conclusion: The early functional recovery of bilateral TKA patient lags behind that of unilateral TKA patient for the first 5 days, becomes equal by the 14 th day and remains equal till 1 year after surgery. Bilateral TKA patients regain their preoperative functional status by 6 weeks against 3 months for unilateral TKA. The operative status of the contralateral knee makes no difference to early functional recovery after unilateral TKA. With bilateral TKA, there is no difference in pain and ROM parameters.

  5. Structural and functional characterisation of Aichi virus RNA dependent RNA polymerase

    Czech Academy of Sciences Publication Activity Database

    Dubánková, Anna; Humpolíčková, Jana; Šilhán, Jan; Bäumlová, Adriana; Chalupská, Dominika; Klíma, Martin; Bouřa, Evžen

    2017-01-01

    Roč. 15, č. 1 (2017), s. 7-8 ISSN 2336-7202. [Mezioborové setkání mladých biologů, biochemiků a chemiků /17./. 30.05.2017-01.06.2017, Milovy] Institutional support: RVO:61388963 Keywords : Aichi virus * RNA replication Subject RIV: CE - Biochemistry

  6. A long noncoding RNA controls muscle differentiation by functioning as a competing endogenous RNA.

    KAUST Repository

    Cesana, Marcella

    2011-10-01

    Recently, a new regulatory circuitry has been identified in which RNAs can crosstalk with each other by competing for shared microRNAs. Such competing endogenous RNAs (ceRNAs) regulate the distribution of miRNA molecules on their targets and thereby impose an additional level of post-transcriptional regulation. Here we identify a muscle-specific long noncoding RNA, linc-MD1, which governs the time of muscle differentiation by acting as a ceRNA in mouse and human myoblasts. Downregulation or overexpression of linc-MD1 correlate with retardation or anticipation of the muscle differentiation program, respectively. We show that linc-MD1 "sponges" miR-133 and miR-133 [corrected] to regulate the expression of MAML1 and MEF2C, transcription factors that activate muscle-specific gene expression. Finally, we demonstrate that linc-MD1 exerts the same control over differentiation timing in human myoblasts, and that its levels are strongly reduced in Duchenne muscle cells. We conclude that the ceRNA network plays an important role in muscle differentiation.

  7. Pre-mRNA Splicing in Plants: In Vivo Functions of RNA-Binding Proteins Implicated in the Splicing Process

    Directory of Open Access Journals (Sweden)

    Katja Meyer

    2015-07-01

    Full Text Available Alternative pre-messenger RNA splicing in higher plants emerges as an important layer of regulation upon exposure to exogenous and endogenous cues. Accordingly, mutants defective in RNA-binding proteins predicted to function in the splicing process show severe phenotypic alterations. Among those are developmental defects, impaired responses to pathogen threat or abiotic stress factors, and misregulation of the circadian timing system. A suite of splicing factors has been identified in the model plant Arabidopsis thaliana. Here we summarize recent insights on how defects in these splicing factors impair plant performance.

  8. Functional characterization of endogenous siRNA target genes in Caenorhabditis elegans

    Directory of Open Access Journals (Sweden)

    Heikkinen Liisa

    2008-06-01

    Full Text Available Abstract Background Small interfering RNA (siRNA molecules mediate sequence specific silencing in RNA interference (RNAi, a gene regulatory phenomenon observed in almost all organisms. Large scale sequencing of small RNA libraries obtained from C. elegans has revealed that a broad spectrum of siRNAs is endogenously transcribed from genomic sequences. The biological role and molecular diversity of C. elegans endogenous siRNA (endo-siRNA molecules, nonetheless, remain poorly understood. In order to gain insight into their biological function, we annotated two large libraries of endo-siRNA sequences, identified their cognate targets, and performed gene ontology analysis to identify enriched functional categories. Results Systematic trends in categorization of target genes according to the specific length of siRNA sequences were observed: 18- to 22-mer siRNAs were associated with genes required for embryonic development; 23-mers were associated uniquely with post-embryonic development; 24–26-mers were associated with phosphorus metabolism or protein modification. Moreover, we observe that some argonaute related genes associate with siRNAs with multiple reads. Sequence frequency graphs suggest that different lengths of siRNAs share similarities in overall sequence structure: the 5' end begins with G, while the body predominates with U and C. Conclusion These results suggest that the lengths of endogenous siRNA molecules are consequential to their biological functions since the gene ontology categories for their cognate mRNA targets vary depending upon their lengths.

  9. Deep sequencing of RNA from immune cell-derived vesicles uncovers the selective incorporation of small non-coding RNA biotypes with potential regulatory functions.

    Science.gov (United States)

    Nolte-'t Hoen, Esther N M; Buermans, Henk P J; Waasdorp, Maaike; Stoorvogel, Willem; Wauben, Marca H M; 't Hoen, Peter A C

    2012-10-01

    Cells release RNA-carrying vesicles and membrane-free RNA/protein complexes into the extracellular milieu. Horizontal vesicle-mediated transfer of such shuttle RNA between cells allows dissemination of genetically encoded messages, which may modify the function of target cells. Other studies used array analysis to establish the presence of microRNAs and mRNA in cell-derived vesicles from many sources. Here, we used an unbiased approach by deep sequencing of small RNA released by immune cells. We found a large variety of small non-coding RNA species representing pervasive transcripts or RNA cleavage products overlapping with protein coding regions, repeat sequences or structural RNAs. Many of these RNAs were enriched relative to cellular RNA, indicating that cells destine specific RNAs for extracellular release. Among the most abundant small RNAs in shuttle RNA were sequences derived from vault RNA, Y-RNA and specific tRNAs. Many of the highly abundant small non-coding transcripts in shuttle RNA are evolutionary well-conserved and have previously been associated to gene regulatory functions. These findings allude to a wider range of biological effects that could be mediated by shuttle RNA than previously expected. Moreover, the data present leads for unraveling how cells modify the function of other cells via transfer of specific non-coding RNA species.

  10. HIGH ECCENTRICITY EOQ TOTAL COST FUNCTION YIELDS JIT RESULTs

    Directory of Open Access Journals (Sweden)

    Willian Roach

    2010-06-01

    Full Text Available No estoque de bens perecíveis, o custo de armazenamento H é muito maior do que o previsto na fórmula clássica do lote econômico do pedido (EOQ. Para bens perecíveis, a função custo total no EOQ é um pico e não uma reta horizontal. Esta forma pontiaguda leva o modelo EOQ a produzir entregas just in time (JIT - resultados semelhantes. O efeito pontiagudo (excentricidade da curva de custo total do lote econômico EOQ depende apenas do custo de armazenamento (H e não da demanda anual (D ou do custo do pedido (S. D e S determinam o nível (altura da curva de custo total do estoque (TC, mas não a forma.

  11. Optimization of ribosome structure and function by rRNA base modification.

    Directory of Open Access Journals (Sweden)

    Jennifer L Baxter-Roshek

    2007-01-01

    Full Text Available Translating mRNA sequences into functional proteins is a fundamental process necessary for the viability of organisms throughout all kingdoms of life. The ribosome carries out this process with a delicate balance between speed and accuracy. This work investigates how ribosome structure and function are affected by rRNA base modification. The prevailing view is that rRNA base modifications serve to fine tune ribosome structure and function.To test this hypothesis, yeast strains deficient in rRNA modifications in the ribosomal peptidyltransferase center were monitored for changes in and translational fidelity. These studies revealed allele-specific sensitivity to translational inhibitors, changes in reading frame maintenance, nonsense suppression and aa-tRNA selection. Ribosomes isolated from two mutants with the most pronounced phenotypic changes had increased affinities for aa-tRNA, and surprisingly, increased rates of peptidyltransfer as monitored by the puromycin assay. rRNA chemical analyses of one of these mutants identified structural changes in five specific bases associated with the ribosomal A-site.Together, the data suggest that modification of these bases fine tune the structure of the A-site region of the large subunit so as to assure correct positioning of critical rRNA bases involved in aa-tRNA accommodation into the PTC, of the eEF-1A.aa-tRNA.GTP ternary complex with the GTPase associated center, and of the aa-tRNA in the A-site. These findings represent a direct demonstration in support of the prevailing hypothesis that rRNA modifications serve to optimize rRNA structure for production of accurate and efficient ribosomes.

  12. Sequence composition similarities with the 7SL RNA are highly predictive of functional genomic features

    OpenAIRE

    Paquet, Yanick; Anderson, Alan

    2010-01-01

    Transposable elements derived from the 7SL RNA gene, such as Alu elements in primates, have had remarkable success in several mammalian lineages. The results presented here show a broad spectrum of functions for genomic segments that display sequence composition similarities with the 7SL RNA gene. Using thoroughly documented loci, we report that DNaseI-hypersensitive sites can be singled out in large genomic sequences by an assessment of sequence composition similarities with the 7SL RNA gene...

  13. Noncoding Subgenomic Flavivirus RNA: Multiple Functions in West Nile Virus Pathogenesis and Modulation of Host Responses

    Directory of Open Access Journals (Sweden)

    Justin A. Roby

    2014-01-01

    Full Text Available Flaviviruses are a large group of positive strand RNA viruses transmitted by arthropods that include many human pathogens such as West Nile virus (WNV, Japanese encephalitis virus (JEV, yellow fever virus, dengue virus, and tick-borne encephalitis virus. All members in this genus tested so far are shown to produce a unique subgenomic flavivirus RNA (sfRNA derived from the 3' untranslated region (UTR. sfRNA is a product of incomplete degradation of genomic RNA by the cell 5'–3' exoribonuclease XRN1 which stalls at highly ordered secondary RNA structures at the beginning of the 3'UTR. Generation of sfRNA results in inhibition of XRN1 activity leading to an increase in stability of many cellular mRNAs. Mutant WNV deficient in sfRNA generation was highly attenuated displaying a marked decrease in cytopathicity in cells and pathogenicity in mice. sfRNA has also been shown to inhibit the antiviral activity of IFN-α/β by yet unknown mechanism and of the RNAi pathway by likely serving as a decoy substrate for Dicer. Thus, sfRNA is involved in modulating multiple cellular pathways to facilitate viral pathogenicity; however the overlying mechanism linking all these multiple functions of sfRNA remains to be elucidated.

  14. a locally adapted functional outcome measurement score for total

    African Journals Online (AJOL)

    Preceding the yearly operating program, a preoperative grouped outpatient clinic is run. At this time patients selected for THR filled out the questionnaire, possibly a second time. Patients who had completed the questionnaire twice were asked whether their pain or functional impairment had changed since the previous.

  15. Deep sequencing of RNA from immune cell-derived vesicles uncovers the selective incorporation of small non-coding RNA biotypes with potential regulatory functions.

    NARCIS (Netherlands)

    Nolte-'t Hoen, E.N.M.|info:eu-repo/dai/nl/261632175; Buermans, H.P.; Waasdorp, M.; Stoorvogel, W.|info:eu-repo/dai/nl/074352385; Wauben, M.H.M.|info:eu-repo/dai/nl/112675735; `t Hoen, P.A.C.

    2012-01-01

    Cells release RNA-carrying vesicles and membrane-free RNA/protein complexes into the extracellular milieu. Horizontal vesicle-mediated transfer of such shuttle RNA between cells allows dissemination of genetically encoded messages, which may modify the function of target cells. Other studies used

  16. Structural and functional basis for RNA cleavage by Ire1

    Directory of Open Access Journals (Sweden)

    Stroud Robert M

    2011-07-01

    Full Text Available Abstract Background The unfolded protein response (UPR controls the protein folding capacity of the endoplasmic reticulum (ER. Central to this signaling pathway is the ER-resident bifunctional transmembrane kinase/endoribonuclease Ire1. The endoribonuclease (RNase domain of Ire1 initiates a non-conventional mRNA splicing reaction, leading to the production of a transcription factor that controls UPR target genes. The mRNA splicing reaction is an obligatory step of Ire1 signaling, yet its mechanism has remained poorly understood due to the absence of substrate-bound crystal structures of Ire1, the lack of structural similarity between Ire1 and other RNases, and a scarcity of quantitative enzymological data. Here, we experimentally define the active site of Ire1 RNase and quantitatively evaluate the contribution of the key active site residues to catalysis. Results This analysis and two new crystal structures suggest that Ire1 RNase uses histidine H1061 and tyrosine Y1043 as the general acid-general base pair contributing ≥ 7.6 kcal/mol and 1.4 kcal/mol to transition state stabilization, respectively, and asparagine N1057 and arginine R1056 for coordination of the scissile phosphate. Investigation of the stem-loop recognition revealed that additionally to the stem-loops derived from the classic Ire1 substrates HAC1 and Xbp1 mRNA, Ire1 can site-specifically and rapidly cleave anticodon stem-loop (ASL of unmodified tRNAPhe, extending known substrate specificity of Ire1 RNase. Conclusions Our data define the catalytic center of Ire1 RNase and suggest a mechanism of RNA cleavage: each RNase monomer apparently contains a separate catalytic apparatus for RNA cleavage, whereas two RNase subunits contribute to RNA stem-loop docking. Conservation of the key residues among Ire1 homologues suggests that the mechanism elucidated here for yeast Ire1 applies to Ire1 in metazoan cells, and to the only known Ire1 homologue RNase L.

  17. DIANA-microT web server v5.0: service integration into miRNA functional analysis workflows.

    Science.gov (United States)

    Paraskevopoulou, Maria D; Georgakilas, Georgios; Kostoulas, Nikos; Vlachos, Ioannis S; Vergoulis, Thanasis; Reczko, Martin; Filippidis, Christos; Dalamagas, Theodore; Hatzigeorgiou, A G

    2013-07-01

    MicroRNAs (miRNAs) are small endogenous RNA molecules that regulate gene expression through mRNA degradation and/or translation repression, affecting many biological processes. DIANA-microT web server (http://www.microrna.gr/webServer) is dedicated to miRNA target prediction/functional analysis, and it is being widely used from the scientific community, since its initial launch in 2009. DIANA-microT v5.0, the new version of the microT server, has been significantly enhanced with an improved target prediction algorithm, DIANA-microT-CDS. It has been updated to incorporate miRBase version 18 and Ensembl version 69. The in silico-predicted miRNA-gene interactions in Homo sapiens, Mus musculus, Drosophila melanogaster and Caenorhabditis elegans exceed 11 million in total. The web server was completely redesigned, to host a series of sophisticated workflows, which can be used directly from the on-line web interface, enabling users without the necessary bioinformatics infrastructure to perform advanced multi-step functional miRNA analyses. For instance, one available pipeline performs miRNA target prediction using different thresholds and meta-analysis statistics, followed by pathway enrichment analysis. DIANA-microT web server v5.0 also supports a complete integration with the Taverna Workflow Management System (WMS), using the in-house developed DIANA-Taverna Plug-in. This plug-in provides ready-to-use modules for miRNA target prediction and functional analysis, which can be used to form advanced high-throughput analysis pipelines.

  18. FARNA: knowledgebase of inferred functions of non-coding RNA transcripts

    KAUST Repository

    Alam, Tanvir

    2016-10-12

    Non-coding RNA (ncRNA) genes play a major role in control of heterogeneous cellular behavior. Yet, their functions are largely uncharacterized. Current available databases lack in-depth information of ncRNA functions across spectrum of various cells/tissues. Here, we present FARNA, a knowledgebase of inferred functions of 10,289 human ncRNA transcripts (2,734 microRNA and 7,555 long ncRNA) in 119 tissues and 177 primary cells of human. Since transcription factors (TFs) and TF co-factors (TcoFs) are crucial components of regulatory machinery for activation of gene transcription, cellular processes and diseases in which TFs and TcoFs are involved suggest functions of the transcripts they regulate. In FARNA, functions of a transcript are inferred from TFs and TcoFs whose genes co-express with the transcript controlled by these TFs and TcoFs in a considered cell/tissue. Transcripts were annotated using statistically enriched GO terms, pathways and diseases across cells/tissues based on guilt-by-association principle. Expression profiles across cells/tissues based on Cap Analysis of Gene Expression (CAGE) are provided. FARNA, having the most comprehensive function annotation of considered ncRNAs across widest spectrum of human cells/tissues, has a potential to greatly contribute to our understanding of ncRNA roles and their regulatory mechanisms in human. FARNA can be accessed at: http://cbrc.kaust.edu.sa/farna

  19. The human cap-binding complex is functionally connected to the nuclear RNA exosome

    DEFF Research Database (Denmark)

    Andersen, Peter Refsing; Domanski, Michal; Kristiansen, Maiken Søndergaard

    2013-01-01

    of combinatorial depletion of CBCN and exosome components underscore the functional relevance of CBC-exosome bridging at the level of target RNA. Specifically, CBCA suppresses read-through products of several RNA families by promoting their transcriptional termination. We suggest that the RNP 5' cap links...

  20. RNA

    African Journals Online (AJOL)

    SARAH

    30 nov. 2013 ... RÉSUMÉ. Objectif : La présente étude est conduite dans les régions de Maradi et Zinder situées dans le Centre-Sud du. Niger où la pratique de la régénération naturelle assistée des ligneux dans les champs (RNA) a permis de reverdir plus de 5 millions d'hectares. Le but de ce travail est d'évaluer ...

  1. Functional genomic analysis of human mitochondrial RNA processing.

    Science.gov (United States)

    Wolf, Ashley R; Mootha, Vamsi K

    2014-05-08

    Both strands of human mtDNA are transcribed in continuous, multigenic units that are cleaved into the mature rRNAs, tRNAs, and mRNAs required for respiratory chain biogenesis. We sought to systematically identify nuclear-encoded proteins that contribute to processing of mtRNAs within the organelle. First, we devised and validated a multiplex MitoString assay that quantitates 27 mature and precursor mtDNA transcripts. Second, we applied MitoString profiling to evaluate the impact of silencing each of 107 mitochondrial-localized, predicted RNA-binding proteins. With the resulting data set, we rediscovered the roles of recently identified RNA-processing enzymes, detected unanticipated roles of known disease genes in RNA processing, and identified new regulatory factors. We demonstrate that one such factor, FASTKD4, modulates the half-lives of a subset of mt-mRNAs and associates with mtRNAs in vivo. MitoString profiling may be useful for diagnosing and deciphering the pathogenesis of mtDNA disorders. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  2. Functional Genomic Analysis of Human Mitochondrial RNA Processing

    Directory of Open Access Journals (Sweden)

    Ashley R. Wolf

    2014-05-01

    Full Text Available Both strands of human mtDNA are transcribed in continuous, multigenic units that are cleaved into the mature rRNAs, tRNAs, and mRNAs required for respiratory chain biogenesis. We sought to systematically identify nuclear-encoded proteins that contribute to processing of mtRNAs within the organelle. First, we devised and validated a multiplex MitoString assay that quantitates 27 mature and precursor mtDNA transcripts. Second, we applied MitoString profiling to evaluate the impact of silencing each of 107 mitochondrial-localized, predicted RNA-binding proteins. With the resulting data set, we rediscovered the roles of recently identified RNA-processing enzymes, detected unanticipated roles of known disease genes in RNA processing, and identified new regulatory factors. We demonstrate that one such factor, FASTKD4, modulates the half-lives of a subset of mt-mRNAs and associates with mtRNAs in vivo. MitoString profiling may be useful for diagnosing and deciphering the pathogenesis of mtDNA disorders.

  3. MicroRNA functions in osteogenesis and dysfunctions in osteoporosis.

    Science.gov (United States)

    van Wijnen, Andre J; van de Peppel, Jeroen; van Leeuwen, Johannes P; Lian, Jane B; Stein, Gary S; Westendorf, Jennifer J; Oursler, Merry-Jo; Im, Hee-Jeong; Taipaleenmäki, Hanna; Hesse, Eric; Riester, Scott; Kakar, Sanjeev

    2013-06-01

    MicroRNAs (miRNAs) are critical post-transcriptional regulators of gene expression that control osteoblast mediated bone formation and osteoclast-related bone remodeling. Deregulation of miRNA mediated mechanisms is emerging as an important pathological factor in bone degeneration (eg, osteoporosis) and other bone-related diseases. MiRNAs are intriguing regulatory molecules that are networked with cell signaling pathways and intricate transcriptional programs through ingenuous circuits with remarkably simple logic. This overview examines key principles by which miRNAs control differentiation of osteoblasts as they evolve from mesenchymal stromal cells during osteogenesis, or of osteoclasts as they originate from monocytic precursors in the hematopoietic lineage during osteoclastogenesis. Of particular note are miRNAs that are temporally upregulated during osteoblastogenesis (eg, miR-218) or osteoclastogenesis (eg, miR-148a). Each miRNA stimulates differentiation by suppressing inhibitory signaling pathways ('double-negative' regulation). The excitement surrounding miRNAs in bone biology stems from the prominent effects that individual miRNAs can have on biological transitions during differentiation of skeletal cells and correlations of miRNA dysfunction with bone diseases. MiRNAs have significant clinical potential which is reflected by their versatility as disease-specific biomarkers and their promise as therapeutic agents to ameliorate or reverse bone tissue degeneration.

  4. The RNA helicase DDX1 is involved in restricted HIV-1 Rev function in human astrocytes

    International Nuclear Information System (INIS)

    Fang Jianhua; Acheampong, Edward; Dave, Rajnish; Wang Fengxiang; Mukhtar, Muhammad; Pomerantz, Roger J.

    2005-01-01

    Productive infection by human immunodeficiency virus type I (HIV-1) in the central nervous system (CNS) involves mainly macrophages and microglial cells. A frequency of less than 10% of human astrocytes is estimated to be infectable with HIV-1. Nonetheless, this relatively low percentage of infected astrocytes, but associated with a large total number of astrocytic cells in the CNS, makes human astrocytes a critical part in the analyses of potential HIV-1 reservoirs in vivo. Investigations in astrocytic cell lines and primary human fetal astrocytes revealed that limited HIV-1 replication in these cells resulted from low-level viral entry, transcription, viral protein processing, and virion maturation. Of note, a low ratio of unspliced versus spliced HIV-1-specific RNA was also investigated, as Rev appeared to act aberrantly in astrocytes, via loss of nuclear and/or nucleolar localization and diminished Rev-mediated function. Host cellular machinery enabling Rev function has become critical for elucidation of diminished Rev activity, especially for those factors leading to RNA metabolism. We have recently identified a DEAD-box protein, DDX1, as a Rev cellular co-factor and now have explored its potential importance in astrocytes. Cells were infected with HIV-1 pseudotyped with envelope glycoproteins of amphotropic murine leukemia viruses (MLV). Semi-quantitative reverse transcriptase-polymerase chain reactions (RT-PCR) for unspliced, singly-spliced, and multiply-spliced RNA clearly showed a lower ratio of unspliced/singly-spliced over multiply-spliced HIV-1-specific RNA in human astrocytes as compared to Rev-permissive, non-glial control cells. As well, the cellular localization of Rev in astrocytes was cytoplasmically dominant as compared to that of Rev-permissive, non-glial controls. This endogenous level of DDX1 expression in astrocytes was demonstrated directly to lead to a shift of Rev sub-cellular distribution dominance from nuclear and/or nucleolar to

  5. The structure and function of tRNA genes of higher eukaryotes.

    Science.gov (United States)

    Kubli, E

    1981-01-15

    The most recent findings concerning the structure and function of tRNA genes of higher eukaryotes are discussed in an exemplary way. The tRNA genes of higher organisms are either dispersed or clustered at different sites of the genome. Clusters contain tRNA genes oriented in both directions and on both strands of the DNA with spacers of various length inbetween. Some genes contain intervening sequences close to the 3' side of the anticodon. The primary transcription product possesses a 5' leader and a 3' trailer sequence which are removed by several maturation steps in a strict temporal and spacial order. Internal transcription control regions (promotors) are located at the 5' and 3' ends of the mature tRNA coding section of the tRNA gene. External sequences modulating the efficiency of the expression are present at the immediate 5' ends of the genes. Transfer RNA genes are located nonrandomly in the nucleosomes.

  6. Regulation of Trypanosoma brucei Total and Polysomal mRNA during Development within Its Mammalian Host.

    Directory of Open Access Journals (Sweden)

    Paul Capewell

    Full Text Available The gene expression of Trypanosoma brucei has been examined extensively in the blood of mammalian hosts and in forms found in the midgut of its arthropod vector, the tsetse fly. However, trypanosomes also undergo development within the mammalian bloodstream as they progress from morphologically 'slender forms' to transmissible 'stumpy forms' through morphological intermediates. This transition is temporally progressive within the first wave of parasitaemia such that gene expression can be monitored in relatively pure slender and stumpy populations as well as during the progression between these extremes. The development also represents the progression of cells from translationally active forms adapted for proliferation in the host to translationally quiescent forms, adapted for transmission. We have used metabolic labelling to quantitate translational activity in slender forms, stumpy forms and in forms undergoing early differentiation to procyclic forms in vitro. Thereafter we have examined the cohort of total mRNAs that are enriched throughout development in the mammalian bloodstream (slender, intermediate and stumpy forms, irrespective of strain, revealing those that exhibit consistent developmental regulation rather than sample specific changes. Transcripts that cosediment with polysomes in stumpy forms and slender forms have also been enriched to identify transcripts that escape translational repression prior to transmission. Combined, the expression and polysomal association of transcripts as trypanosomes undergo development in the mammalian bloodstream have been defined, providing a resource for trypanosome researchers. This facilitates the identification of those that undergo developmental regulation in the bloodstream and therefore those likely to have a role in the survival and capacity for transmission of stumpy forms.

  7. MicroRNA functional network in pancreatic cancer: From biology to ...

    Indian Academy of Sciences (India)

    2011-06-07

    Jun 7, 2011 ... Cellular pathways; genetic network; microRNA; pancreatic cancer; tumorigenic transformation; 3' untranslated region ... components of the complex functional pathway networks controlling important cellular processes, such as proliferation, development, differentiation, stress response' and apoptosis.

  8. An integrated miRNA functional screening and target validation method for organ morphogenesis

    OpenAIRE

    Rebustini, Ivan T.; Vlahos, Maryann; Packer, Trevor; Kukuruzinska, Maria A.; Maas, Richard L.

    2016-01-01

    The relative ease of identifying microRNAs and their increasing recognition as important regulators of organogenesis motivate the development of methods to efficiently assess microRNA function during organ morphogenesis. In this context, embryonic organ explants provide a reliable and reproducible system that recapitulates some of the important early morphogenetic processes during organ development. Here we present a method to target microRNA function in explanted mouse embryonic organs. Our ...

  9. RNA-directed DNA methylation: Mechanisms and functions

    KAUST Repository

    Mahfouz, Magdy M.

    2010-07-01

    Epigenetic RNA based gene silencing mechanisms play a major role in genome stability and control of gene expression. Transcriptional gene silencing via RNA-directed DNA methylation (RdDM) guides the epigenetic regulation of the genome in response to disease states, growth, developmental and stress signals. RdDM machinery is composed of proteins that produce and modify 24-nt- long siRNAs, recruit the RdDM complex to genomic targets, methylate DNA and remodel chromatin. The final DNA methylation pattern is determined by either DNA methyltransferase alone or by the combined action of DNA methyltransferases and demethylases. The dynamic interaction between RdDM and demethylases may render the plant epigenome plastic to growth, developmental, and environmental cues. The epigenome plasticity may allow the plant genome to assume many epigenomes and to have the right epigenome at the right time in response to intracellular or extracellular stimuli. This review discusses recent advances in RdDM research and considers future perspectives.

  10. Protein functional features are reflected in the patterns of mRNA translation speed.

    Science.gov (United States)

    López, Daniel; Pazos, Florencio

    2015-07-09

    The degeneracy of the genetic code makes it possible for the same amino acid string to be coded by different messenger RNA (mRNA) sequences. These "synonymous mRNAs" may differ largely in a number of aspects related to their overall translational efficiency, such as secondary structure content and availability of the encoded transfer RNAs (tRNAs). Consequently, they may render different yields of the translated polypeptides. These mRNA features related to translation efficiency are also playing a role locally, resulting in a non-uniform translation speed along the mRNA, which has been previously related to some protein structural features and also used to explain some dramatic effects of "silent" single-nucleotide-polymorphisms (SNPs). In this work we perform the first large scale analysis of the relationship between three experimental proxies of mRNA local translation efficiency and the local features of the corresponding encoded proteins. We found that a number of protein functional and structural features are reflected in the patterns of ribosome occupancy, secondary structure and tRNA availability along the mRNA. One or more of these proxies of translation speed have distinctive patterns around the mRNA regions coding for certain protein local features. In some cases the three patterns follow a similar trend. We also show specific examples where these patterns of translation speed point to the protein's important structural and functional features. This support the idea that the genome not only codes the protein functional features as sequences of amino acids, but also as subtle patterns of mRNA properties which, probably through local effects on the translation speed, have some consequence on the final polypeptide. These results open the possibility of predicting a protein's functional regions based on a single genomic sequence, and have implications for heterologous protein expression and fine-tuning protein function.

  11. Biochemical and genetic functional dissection of the P38 viral suppressor of RNA silencing.

    Science.gov (United States)

    Iki, Taichiro; Tschopp, Marie-Aude; Voinnet, Olivier

    2017-05-01

    Phytoviruses encode viral suppressors of RNA silencing (VSRs) to counteract the plant antiviral silencing response, which relies on virus-derived small interfering (si)RNAs processed by Dicer RNaseIII enzymes and subsequently loaded into ARGONAUTE (AGO) effector proteins. Here, a tobacco cell-free system was engineered to recapitulate the key steps of antiviral RNA silencing and, in particular, the most upstream double-stranded (ds)RNA processing reaction, not kinetically investigated thus far in the context of plant VSR studies. Comparative biochemical analyses of distinct VSRs in the reconstituted assay showed that in all cases tested, VSR interactions with siRNA duplexes inhibited the loading, but not the activity, of antiviral AGO1 and AGO2. Turnip crinkle virus P38 displayed the additional and unique property to bind both synthetic and RNA-dependent-RNA-polymerase-generated long dsRNAs, and inhibited the processing into siRNAs. Single amino acid substitutions in P38 could dissociate dsRNA-processing from AGO-loading inhibition in vitro and in vivo, illustrating dual-inhibitory strategies discriminatively deployed within a single viral protein, which, we further show, are bona fide suppressor functions that evolved independently of the conserved coat protein function of P38. © 2017 Iki et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  12. Integrative Analysis of miRNA and mRNA Paired Expression Profiling of Primary Fibroblast Derived from Diabetic Foot Ulcers Reveals Multiple Impaired Cellular Functions

    Science.gov (United States)

    Liang, Liang; Stone, Rivka C.; Stojadinovic, Olivera; Ramirez, Horacio; Pastar, Irena; Maione, Anna G.; Smith, Avi; Yanez, Vanessa; Veves, Aristides; Kirsner, Robert S.; Garlick, Jonathan A.; Tomic-Canic, Marjana

    2017-01-01

    Diabetic foot ulcers (DFUs) are one of the major complications of diabetes. Its molecular pathology remains poorly understood, impeding the development of effective treatments. Although it has been established that multiple cell types, including fibroblasts, keratinocytes, macrophages and endothelial cells, all contribute to inhibition of healing, less is known regarding contributions of individual cell type. Thus, we generated primary fibroblasts from non-healing DFUs and evaluated their cellular and molecular properties in comparison to non-diabetic foot fibroblasts (NFFs). Specifically, we analyzed both micro-RNA and mRNA expression profiles of primary DFU fibroblasts. Paired genomic analyses identified a total of 331 reciprocal miRNA-mRNA pairs including 21 miRNAs (FC>2.0) along with 239 predicted target genes (FC>1.5) that are significantly and differentially expressed. Of these, we focused on three miRNAs (miR-21-5p, miR-34a-5p, miR-145-5p) that were induced in DFU fibroblasts as most differentially regulated. The involvement of these microRNAs in wound healing was investigated by testing the expression of their downstream targets as well as by quantifying cellular behaviors in prospectively collected and generated cell lines from 15 patients (7 DFUF and 8 NFF samples). We found large number of downstream targets of miR-21-5p, miR-34a-5p, miR-145-5p to be coordinately regulated in mRNA profiles, which was confirmed by qPCR. Pathway analysis on paired miRNA-mRNA profiles predicted inhibition of cell movement and cell proliferation, as well as activation of cell differentiation and senescence in DFU fibroblasts, which was confirmed by cellular assays. We concluded that induction of miR-21-5p, miR-34a-5p, miR-145-5p in DFU dermal fibroblasts plays an important role in impairing multiple cellular functions, thus contributing to overall inhibition of healing in DFUs. PMID:27607190

  13. A common function for mRNA 5' and 3' ends in translation initiation in yeast.

    Science.gov (United States)

    Tarun, S Z; Sachs, A B

    1995-12-01

    The mRNA poly(A) tail and its associated poly(A) binding protein (Pab1p) are ubiquitous in eukaryotes. The function of the poly(A) tail is to stabilize mRNA and to stimulate its translation. The development of a poly(A)- and cap-dependent yeast in vitro translation system has allowed us to understand how poly(A) stimulates translation. We find that Pab1p but not the cap binding protein eIF-4E is required for poly(A) tail-dependent translation, and that the Pab1p-poly(A) tail complex functions to recruit the 40S ribosomal subunit to the mRNA. These data introduce a new step into the pathway of translation initiation and merge the translational functions of the two ends of mRNA.

  14. Obstacles and opportunities in the functional analysis of extracellular vesicle RNA – an ISEV position paper

    Science.gov (United States)

    Mateescu, Bogdan; Kowal, Emma J. K.; van Balkom, Bas W. M.; Bartel, Sabine; Bhattacharyya, Suvendra N.; Buzás, Edit I.; Buck, Amy H.; de Candia, Paola; Chow, Franklin W. N.; Das, Saumya; Driedonks, Tom A. P.; Fernández-Messina, Lola; Haderk, Franziska; Hill, Andrew F.; Jones, Jennifer C.; Van Keuren-Jensen, Kendall R.; Lai, Charles P.; Lässer, Cecilia; Liegro, Italia di; Lunavat, Taral R.; Lorenowicz, Magdalena J.; Maas, Sybren L. N.; Mäger, Imre; Mittelbrunn, Maria; Momma, Stefan; Mukherjee, Kamalika; Nawaz, Muhammed; Pegtel, D. Michiel; Pfaffl, Michael W.; Schiffelers, Raymond M.; Tahara, Hidetoshi; Théry, Clotilde; Tosar, Juan Pablo; Wauben, Marca H. M.; Witwer, Kenneth W.; Nolte-‘t Hoen, Esther N. M.

    2017-01-01

    ABSTRACT The release of RNA-containing extracellular vesicles (EV) into the extracellular milieu has been demonstrated in a multitude of different in vitro cell systems and in a variety of body fluids. RNA-containing EV are in the limelight for their capacity to communicate genetically encoded messages to other cells, their suitability as candidate biomarkers for diseases, and their use as therapeutic agents. Although EV-RNA has attracted enormous interest from basic researchers, clinicians, and industry, we currently have limited knowledge on which mechanisms drive and regulate RNA incorporation into EV and on how RNA-encoded messages affect signalling processes in EV-targeted cells. Moreover, EV-RNA research faces various technical challenges, such as standardisation of EV isolation methods, optimisation of methodologies to isolate and characterise minute quantities of RNA found in EV, and development of approaches to demonstrate functional transfer of EV-RNA in vivo. These topics were discussed at the 2015 EV-RNA workshop of the International Society for Extracellular Vesicles. This position paper was written by the participants of the workshop not only to give an overview of the current state of knowledge in the field, but also to clarify that our incomplete knowledge – of the nature of EV(-RNA)s and of how to effectively and reliably study them – currently prohibits the implementation of gold standards in EV-RNA research. In addition, this paper creates awareness of possibilities and limitations of currently used strategies to investigate EV-RNA and calls for caution in interpretation of the obtained data. PMID:28326170

  15. DIANA-microT web server: elucidating microRNA functions through target prediction.

    Science.gov (United States)

    Maragkakis, M; Reczko, M; Simossis, V A; Alexiou, P; Papadopoulos, G L; Dalamagas, T; Giannopoulos, G; Goumas, G; Koukis, E; Kourtis, K; Vergoulis, T; Koziris, N; Sellis, T; Tsanakas, P; Hatzigeorgiou, A G

    2009-07-01

    Computational microRNA (miRNA) target prediction is one of the key means for deciphering the role of miRNAs in development and disease. Here, we present the DIANA-microT web server as the user interface to the DIANA-microT 3.0 miRNA target prediction algorithm. The web server provides extensive information for predicted miRNA:target gene interactions with a user-friendly interface, providing extensive connectivity to online biological resources. Target gene and miRNA functions may be elucidated through automated bibliographic searches and functional information is accessible through Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. The web server offers links to nomenclature, sequence and protein databases, and users are facilitated by being able to search for targeted genes using different nomenclatures or functional features, such as the genes possible involvement in biological pathways. The target prediction algorithm supports parameters calculated individually for each miRNA:target gene interaction and provides a signal-to-noise ratio and a precision score that helps in the evaluation of the significance of the predicted results. Using a set of miRNA targets recently identified through the pSILAC method, the performance of several computational target prediction programs was assessed. DIANA-microT 3.0 achieved there with 66% the highest ratio of correctly predicted targets over all predicted targets. The DIANA-microT web server is freely available at www.microrna.gr/microT.

  16. Host ESCRT proteins are required for bromovirus RNA replication compartment assembly and function.

    Directory of Open Access Journals (Sweden)

    Arturo Diaz

    2015-03-01

    Full Text Available Positive-strand RNA viruses genome replication invariably is associated with vesicles or other rearranged cellular membranes. Brome mosaic virus (BMV RNA replication occurs on perinuclear endoplasmic reticulum (ER membranes in ~70 nm vesicular invaginations (spherules. BMV RNA replication vesicles show multiple parallels with membrane-enveloped, budding retrovirus virions, whose envelopment and release depend on the host ESCRT (endosomal sorting complexes required for transport membrane-remodeling machinery. We now find that deleting components of the ESCRT pathway results in at least two distinct BMV phenotypes. One group of genes regulate RNA replication and the frequency of viral replication complex formation, but had no effect on spherule size, while a second group of genes regulate RNA replication in a way or ways independent of spherule formation. In particular, deleting SNF7 inhibits BMV RNA replication > 25-fold and abolishes detectable BMV spherule formation, even though the BMV RNA replication proteins accumulate and localize normally on perinuclear ER membranes. Moreover, BMV ESCRT recruitment and spherule assembly depend on different sets of protein-protein interactions from those used by multivesicular body vesicles, HIV-1 virion budding, or tomato bushy stunt virus (TBSV spherule formation. These and other data demonstrate that BMV requires cellular ESCRT components for proper formation and function of its vesicular RNA replication compartments. The results highlight growing but diverse interactions of ESCRT factors with many viruses and viral processes, and potential value of the ESCRT pathway as a target for broad-spectrum antiviral resistance.

  17. Human 2'-phosphodiesterase localizes to the mitochondrial matrix with a putative function in mitochondrial RNA turnover

    DEFF Research Database (Denmark)

    Poulsen, Jesper Buchhave; Andersen, Kasper Røjkjær; Kjær, Karina Hansen

    2011-01-01

    . Interestingly, 2′-PDE shares both functionally and structurally characteristics with the CCR4-type exonuclease–endonuclease–phosphatase family of deadenylases. Here we show that 2′-PDE locates to the mitochondrial matrix of human cells, and comprise an active 3′–5′ exoribonuclease exhibiting a preference...... for oligo-adenosine RNA like canonical cytoplasmic deadenylases. Furthermore, we document a marked negative association between 2′-PDE and mitochondrial mRNA levels following siRNA-directed knockdown and plasmid-mediated overexpression, respectively. The results indicate that 2′-PDE, apart from playing...

  18. Total rRNA-Seq Analysis Gives Insight into Bacterial, Fungal, Protozoal and Archaeal Communities in the Rumen Using an Optimized RNA Isolation Method

    Directory of Open Access Journals (Sweden)

    Chijioke O. Elekwachi

    2017-09-01

    Full Text Available Advances in high throughput, next generation sequencing technologies have allowed an in-depth examination of biological environments and phenomena, and are particularly useful for culture-independent microbial community studies. Recently the use of RNA for metatranscriptomic studies has been used to elucidate the role of active microbes in the environment. Extraction of RNA of appropriate quality is critical in these experiments and TRIzol reagent is often used for maintaining stability of RNA molecules during extraction. However, for studies using rumen content there is no consensus on (1 the amount of rumen digesta to use or (2 the amount of TRIzol reagent to be used in RNA extraction procedures. This study evaluated the effect of using various quantities of ground rumen digesta and of TRIzol reagent on the yield and quality of extracted RNA. It also investigated the possibility of using lower masses of solid-phase rumen digesta and lower amounts of TRIzol reagent than is used currently, for extraction of RNA for metatranscriptomic studies. We found that high quality RNA could be isolated from 2 g of ground rumen digesta sample, whilst using 0.6 g of ground matter for RNA extraction and using 3 mL (a 5:1 TRIzol : extraction mass ratio of TRIzol reagent. This represents a significant savings in the cost of RNA isolation. These lower masses and volumes were then applied in the RNA-Seq analysis of solid-phase rumen samples obtained from 6 Angus X Hereford beef heifers which had been fed a high forage diet (comprised of barley straw in a forage-to-concentrate ratio of 70:30 for 102 days. A bioinformatics analysis pipeline was developed in-house that generated relative abundance values of archaea, protozoa, fungi and bacteria in the rumen and also allowed the extraction of individual rRNA variable regions that could be analyzed in downstream molecular ecology programs. The average relative abundances of rRNA transcripts of archaea, bacteria

  19. Computational and experimental studies of reassociating RNA/DNA hybrids containing split functionalities.

    Science.gov (United States)

    Afonin, Kirill A; Bindewald, Eckart; Kireeva, Maria; Shapiro, Bruce A

    2015-01-01

    Recently, we developed a novel technique based on RNA/DNA hybrid reassociation that allows conditional activation of different split functionalities inside diseased cells and in vivo. We further expanded this idea to permit simultaneous activation of multiple different functions in a fully controllable fashion. In this chapter, we discuss some novel computational approaches and experimental techniques aimed at the characterization, design, and production of reassociating RNA/DNA hybrids containing split functionalities. We also briefly describe several experimental techniques that can be used to test these hybrids in vitro and in vivo. 2015 Published by Elsevier Inc.

  20. Nuclear stability and transcriptional directionality separate functionally distinct RNA species

    DEFF Research Database (Denmark)

    Andersson, Robin; Andersen, Peter Refsing; Valen, Eivind

    2014-01-01

    Mammalian genomes are pervasively transcribed, yielding a complex transcriptome with high variability in composition and cellular abundance. Although recent efforts have identified thousands of new long non-coding (lnc) RNAs and demonstrated a complex transcriptional repertoire produced by protei...... a vast majority of unstable transcripts. The predictive power of the approach promises to streamline the functional analysis of known and novel RNAs....

  1. Crystal structure analysis reveals functional flexibility in the selenocysteine-specific tRNA from mouse.

    Directory of Open Access Journals (Sweden)

    Oleg M Ganichkin

    Full Text Available Selenocysteine tRNAs (tRNA(Sec exhibit a number of unique identity elements that are recognized specifically by proteins of the selenocysteine biosynthetic pathways and decoding machineries. Presently, these identity elements and the mechanisms by which they are interpreted by tRNA(Sec-interacting factors are incompletely understood.We applied rational mutagenesis to obtain well diffracting crystals of murine tRNA(Sec. tRNA(Sec lacking the single-stranded 3'-acceptor end ((ΔGCCARNA(Sec yielded a crystal structure at 2.0 Å resolution. The global structure of (ΔGCCARNA(Sec resembles the structure of human tRNA(Sec determined at 3.1 Å resolution. Structural comparisons revealed flexible regions in tRNA(Sec used for induced fit binding to selenophosphate synthetase. Water molecules located in the present structure were involved in the stabilization of two alternative conformations of the anticodon stem-loop. Modeling of a 2'-O-methylated ribose at position U34 of the anticodon loop as found in a sub-population of tRNA(Secin vivo showed how this modification favors an anticodon loop conformation that is functional during decoding on the ribosome. Soaking of crystals in Mn(2+-containing buffer revealed eight potential divalent metal ion binding sites but the located metal ions did not significantly stabilize specific structural features of tRNA(Sec.We provide the most highly resolved structure of a tRNA(Sec molecule to date and assessed the influence of water molecules and metal ions on the molecule's conformation and dynamics. Our results suggest how conformational changes of tRNA(Sec support its interaction with proteins.

  2. Cardiac Disease Status Dictates Functional mRNA Targeting Profiles of Individual MicroRNAs.

    Science.gov (United States)

    Matkovich, Scot J; Dorn, Gerald W; Grossenheider, Tiffani C; Hecker, Peter A

    2015-12-01

    MicroRNAs are key players in cardiac stress responses, but the mRNAs, whose abundance and translational potential are primarily affected by changes in cardiac microRNAs, are not well defined. Stimulus-induced, large-scale alterations in the cardiac transcriptome, together with consideration of the law of mass action, further suggest that the mRNAs most substantively targeted by individual microRNAs will vary between unstressed and stressed conditions. To test the hypothesis that microRNA target profiles differ in health and disease, we traced the fate of empirically determined miR-133a and miR-378 targets in mouse hearts undergoing pressure overload hypertrophy. Ago2 immunoprecipitation with RNA sequencing (RNA-induced silencing complex sequencing) was used for unbiased definition of microRNA-dependent and microRNA-independent alterations occurring among ≈13 000 mRNAs in response to transverse aortic constriction (TAC). Of 37 direct targets of miR-133a defined in unstressed hearts (fold change ≥25%, false discovery rate the effect of TAC on microRNA direct target selection resulted in widespread alterations of signaling function. Numerous microRNA-mediated regulatory events occurring exclusively during pressure overload revealed signaling networks that may be responsive to the endogenous decreases in miR-133a during TAC. Pressure overload-mediated changes in overall cardiac RNA content alter microRNA targeting profiles, reinforcing the need to define microRNA targets in tissue-, cell-, and status-specific contexts. © 2015 American Heart Association, Inc.

  3. EWS and FUS bind a subset of transcribed genes encoding proteins enriched in RNA regulatory functions

    DEFF Research Database (Denmark)

    Luo, Yonglun; Friis, Jenny Blechingberg; Fernandes, Ana Miguel

    2015-01-01

    Background FUS (TLS) and EWS (EWSR1) belong to the FET-protein family of RNA and DNA binding proteins. FUS and EWS are structurally and functionally related and participate in transcriptional regulation and RNA processing. FUS and EWS are identified in translocation generated cancer fusion proteins...... and involved in the human neurological diseases amyotrophic lateral sclerosis and fronto-temporal lobar degeneration. Results To determine the gene regulatory functions of FUS and EWS at the level of chromatin, we have performed chromatin immunoprecipitation followed by next generation sequencing (Ch......IP-seq). Our results show that FUS and EWS bind to a subset of actively transcribed genes, that binding often is downstream the poly(A)-signal, and that binding overlaps with RNA polymerase II. Functional examinations of selected target genes identified that FUS and EWS can regulate gene expression...

  4. Biogenesis and function of tRNA fragments during sperm maturation and fertilization in mammals.

    Science.gov (United States)

    Sharma, Upasna; Conine, Colin C; Shea, Jeremy M; Boskovic, Ana; Derr, Alan G; Bing, Xin Y; Belleannee, Clemence; Kucukural, Alper; Serra, Ryan W; Sun, Fengyun; Song, Lina; Carone, Benjamin R; Ricci, Emiliano P; Li, Xin Z; Fauquier, Lucas; Moore, Melissa J; Sullivan, Robert; Mello, Craig C; Garber, Manuel; Rando, Oliver J

    2016-01-22

    Several recent studies link parental environments to phenotypes in subsequent generations. In this work, we investigate the mechanism by which paternal diet affects offspring metabolism. Protein restriction in mice affects small RNA (sRNA) levels in mature sperm, with decreased let-7 levels and increased amounts of 5' fragments of glycine transfer RNAs (tRNAs). In testicular sperm, tRNA fragments are scarce but increase in abundance as sperm mature in the epididymis. Epididymosomes (vesicles that fuse with sperm during epididymal transit) carry RNA payloads matching those of mature sperm and can deliver RNAs to immature sperm in vitro. Functionally, tRNA-glycine-GCC fragments repress genes associated with the endogenous retroelement MERVL, in both embryonic stem cells and embryos. Our results shed light on sRNA biogenesis and its dietary regulation during posttesticular sperm maturation, and they also link tRNA fragments to regulation of endogenous retroelements active in the preimplantation embryo. Copyright © 2016, American Association for the Advancement of Science.

  5. Isolation and cloning of microRNAs from recalcitrant plant tissues with small amounts of total RNA: a step-by step approach.

    Science.gov (United States)

    Yew, Chee Wei; Kumar, S Vijay

    2012-02-01

    MicroRNAs (miRNAs) are small RNAs (sRNAs) with approximately 21-24 nucleotides in length. They regulate the expression of target genes through the mechanism of RNA silencing. Conventional isolation and cloning of miRNAs methods are usually technical demanding and inefficient. These limitations include the requirement for high amounts of starting total RNA, inefficient ligation of linkers, high amount of PCR artifacts and bias in the formation of short miRNA-concatamers. Here we describe in detail a method that uses 80 μg of total RNA as the starting material. Enhancement of the ligation of sRNAs and linkers with the use of polyethylene glycol (PEG8000) was described. PCR artifacts from the amplification of reverse-transcribed sRNAs were greatly decreased by using lower concentrations of primers and reducing the number of amplification cycles. Large concatamers with up to 1 kb in size with around 20 sRNAs/concatamer were obtained by using an optimized reaction condition. This protocol provide researchers with a rapid, efficient and cost-effective method for the construction of miRNA profiles from plant tissues containing low amounts of total RNA, such as fruit flesh and senescent leaves.

  6. Conserved TRAM Domain Functions as an Archaeal Cold Shock Protein via RNA Chaperone Activity

    Directory of Open Access Journals (Sweden)

    Bo Zhang

    2017-08-01

    Full Text Available Cold shock proteins (Csps enable organisms to acclimate to and survive in cold environments and the bacterial CspA family exerts the cold protection via its RNA chaperone activity. However, most Archaea do not contain orthologs to the bacterial csp. TRAM, a conserved domain among RNA modification proteins ubiquitously distributed in organisms, occurs as an individual protein in most archaeal phyla and has a structural similarity to Csp proteins, yet its biological functions remain unknown. Through physiological and biochemical studies on four TRAM proteins from a cold adaptive archaeon Methanolobus psychrophilus R15, this work demonstrated that TRAM is an archaeal Csp and exhibits RNA chaperone activity. Three TRAM encoding genes (Mpsy_0643, Mpsy_3043, and Mpsy_3066 exhibited remarkable cold-shock induced transcription and were preferentially translated at lower temperature (18°C, while the fourth (Mpsy_2002 was constitutively expressed. They were all able to complement the cspABGE mutant of Escherichia coli BX04 that does not grow in cold temperatures and showed transcriptional antitermination. TRAM3066 (gene product of Mpsy_3066 and TRAM2002 (gene product of Mpsy_2002 displayed sequence-non-specific RNA but not DNA binding activity, and TRAM3066 assisted RNases in degradation of structured RNA, thus validating the RNA chaperone activity of TRAMs. Given the chaperone activity, TRAM is predicted to function beyond a Csp.

  7. A collection of target mimics for comprehensive analysis of microRNA function in Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Marco Todesco

    2010-07-01

    Full Text Available Many targets of plant microRNAs (miRNAs are thought to play important roles in plant physiology and development. However, because plant miRNAs are typically encoded by medium-size gene families, it has often been difficult to assess their precise function. We report the generation of a large-scale collection of knockdowns for Arabidopsis thaliana miRNA families; this has been achieved using artificial miRNA target mimics, a recently developed technique fashioned on an endogenous mechanism of miRNA regulation. Morphological defects in the aerial part were observed for approximately 20% of analyzed families, all of which are deeply conserved in land plants. In addition, we find that non-cleavable mimic sites can confer translational regulation in cis. Phenotypes of plants expressing target mimics directed against miRNAs involved in development were in several cases consistent with previous reports on plants expressing miRNA-resistant forms of individual target genes, indicating that a limited number of targets mediates most effects of these miRNAs. That less conserved miRNAs rarely had obvious effects on plant morphology suggests that most of them do not affect fundamental aspects of development. In addition to insight into modes of miRNA action, this study provides an important resource for the study of miRNA function in plants.

  8. Construction and analysis of cardiac hypertrophy-associated lncRNA-mRNA network based on competitive endogenous RNA reveal functional lncRNAs in cardiac hypertrophy.

    Science.gov (United States)

    Song, Chao; Zhang, Jian; Liu, Yan; Pan, Hao; Qi, Han-Ping; Cao, Yong-Gang; Zhao, Jian-Mei; Li, Shang; Guo, Jing; Sun, Hong-Li; Li, Chun-Quan

    2016-03-08

    Cardiac hypertrophy (CH) could increase cardiac after-load and lead to heart failure. Recent studies have suggested that long non-coding RNA (lncRNA) played a crucial role in the process of the cardiac hypertrophy, such as Mhrt, TERMINATOR. Some studies have further found a new interacting mechanism, competitive endogenous RNA (ceRNA), of which lncRNA could interact with micro-RNAs (miRNA) and indirectly interact with mRNAs through competing interactions. However, the mechanism of ceRNA regulated by lncRNA in the CH remained unclear. In our study, we generated a global triple network containing mRNA, miRNA and lncRNA, and extracted a CH related lncRNA-mRNA network (CHLMN) through integrating the data from starbase, miRanda database and gene expression profile. Based on the ceRNA mechanism, we analyzed the characters of CHLMN and found that 3 lncRNAs (SLC26A4-AS1, RP11-344E13.3 and MAGI1-IT1) were high related to CH. We further performed cluster module analysis and random walk with restart for the CHLMN, finally 14 lncRNAs had been discovered as the potential CH related disease genes. Our results showed that lncRNA played an important role in the CH and could shed new light to the understanding underlying mechanisms of the CH.

  9. The MYC mRNA 3'-UTR couples RNA polymerase II function to glutamine and ribonucleotide levels.

    Science.gov (United States)

    Dejure, Francesca R; Royla, Nadine; Herold, Steffi; Kalb, Jacqueline; Walz, Susanne; Ade, Carsten P; Mastrobuoni, Guido; Vanselow, Jens T; Schlosser, Andreas; Wolf, Elmar; Kempa, Stefan; Eilers, Martin

    2017-07-03

    Deregulated expression of MYC enhances glutamine utilization and renders cell survival dependent on glutamine, inducing "glutamine addiction". Surprisingly, colon cancer cells that express high levels of MYC due to WNT pathway mutations are not glutamine-addicted but undergo a reversible cell cycle arrest upon glutamine deprivation. We show here that glutamine deprivation suppresses translation of endogenous MYC via the 3'-UTR of the MYC mRNA, enabling escape from apoptosis. This regulation is mediated by glutamine-dependent changes in adenosine-nucleotide levels. Glutamine deprivation causes a global reduction in promoter association of RNA polymerase II (RNAPII) and slows transcriptional elongation. While activation of MYC restores binding of MYC and RNAPII function on most promoters, restoration of elongation is imperfect and activation of MYC in the absence of glutamine causes stalling of RNAPII on multiple genes, correlating with R-loop formation. Stalling of RNAPII and R-loop formation can cause DNA damage, arguing that the MYC 3'-UTR is critical for maintaining genome stability when ribonucleotide levels are low. © 2017 The Authors.

  10. Cross disease analysis of co-functional microRNA pairs on a reconstructed network of disease-gene-microRNA tripartite.

    Science.gov (United States)

    Peng, Hui; Lan, Chaowang; Zheng, Yi; Hutvagner, Gyorgy; Tao, Dacheng; Li, Jinyan

    2017-03-24

    MicroRNAs always function cooperatively in their regulation of gene expression. Dysfunctions of these co-functional microRNAs can play significant roles in disease development. We are interested in those multi-disease associated co-functional microRNAs that regulate their common dysfunctional target genes cooperatively in the development of multiple diseases. The research is potentially useful for human disease studies at the transcriptional level and for the study of multi-purpose microRNA therapeutics. We designed a computational method to detect multi-disease associated co-functional microRNA pairs and conducted cross disease analysis on a reconstructed disease-gene-microRNA (DGR) tripartite network. The construction of the DGR tripartite network is by the integration of newly predicted disease-microRNA associations with those relationships of diseases, microRNAs and genes maintained by existing databases. The prediction method uses a set of reliable negative samples of disease-microRNA association and a pre-computed kernel matrix instead of kernel functions. From this reconstructed DGR tripartite network, multi-disease associated co-functional microRNA pairs are detected together with their common dysfunctional target genes and ranked by a novel scoring method. We also conducted proof-of-concept case studies on cancer-related co-functional microRNA pairs as well as on non-cancer disease-related microRNA pairs. With the prioritization of the co-functional microRNAs that relate to a series of diseases, we found that the co-function phenomenon is not unusual. We also confirmed that the regulation of the microRNAs for the development of cancers is more complex and have more unique properties than those of non-cancer diseases.

  11. Functional and evolutionary significance of human microRNA seed region mutations.

    Directory of Open Access Journals (Sweden)

    Christopher G Hill

    Full Text Available MicroRNAs have emerged in recent years as important regulators of cell function in both normal and diseased cells. MiRNAs coordinately regulate large suites of target genes by mRNA degradation and/or translational inhibition. The mRNA target specificities of miRNAs in animals are primarily encoded within a 7 nt "seed region" mapping to positions 2-8 at the molecule's 5' end. We here combine computational analyses with experimental studies to explore the functional significance of sequence variation within the seed region of human miRNAs. The results indicate that a substitution of even a single nucleotide within the seed region changes the spectrum of mRNA targets by >50%. The high functional cost of even single nucleotide changes within seed regions is consistent with their high sequence conservation among miRNA families both within and between species and suggests processes that may underlie the evolution of miRNA regulatory control.

  12. A resource for functional profiling of noncoding RNA in the yeastSaccharomyces cerevisiae.

    Science.gov (United States)

    Parker, Steven; Fraczek, Marcin G; Wu, Jian; Shamsah, Sara; Manousaki, Alkisti; Dungrattanalert, Kobchai; de Almeida, Rogerio Alves; Estrada-Rivadeneyra, Diego; Omara, Walid; Delneri, Daniela; O'Keefe, Raymond T

    2017-08-01

    Eukaryotic genomes are extensively transcribed, generating many different RNAs with no known function. We have constructed 1502 molecular barcoded ncRNA gene deletion strains encompassing 443 ncRNAs in the yeast Saccharomyces cerevisiae as tools for ncRNA functional analysis. This resource includes deletions of small nuclear RNAs (snRNAs), transfer RNAs (tRNAs), small nucleolar RNAs (snoRNAs), and other annotated ncRNAs as well as the more recently identified stable unannotated transcripts (SUTs) and cryptic unstable transcripts (CUTs) whose functions are largely unknown. Specifically, deletions have been constructed for ncRNAs found in the intergenic regions, not overlapping genes or their promoters (i.e., at least 200 bp minimum distance from the closest gene start codon). The deletion strains carry molecular barcodes designed to be complementary with the protein gene deletion collection enabling parallel analysis experiments. These strains will be useful for the numerous genomic and molecular techniques that utilize deletion strains, including genome-wide phenotypic screens under different growth conditions, pooled chemogenomic screens with drugs or chemicals, synthetic genetic array analysis to uncover novel genetic interactions, and synthetic dosage lethality screens to analyze gene dosage. Overall, we created a valuable resource for the RNA community and for future ncRNA research. © 2017 Parker et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  13. Modulation of 16S rRNA function by ribosomal protein S12.

    Science.gov (United States)

    Vila-Sanjurjo, Anton; Lu, Ying; Aragonez, Jamie L; Starkweather, Rebekah E; Sasikumar, Manoj; O'Connor, Michael

    2007-01-01

    Ribosomal protein S12 is a critical component of the decoding center of the 30S ribosomal subunit and is involved in both tRNA selection and the response to streptomycin. We have investigated the interplay between S12 and some of the surrounding 16S rRNA residues by examining the phenotypes of double-mutant ribosomes in strains of Escherichia coli carrying deletions in all chromosomal rrn operons and expressing total rRNA from a single plasmid-borne rrn operon. We show that the combination of S12 and otherwise benign mutations at positions C1409-G1491 in 16S rRNA severely compromises cell growth while the level and range of aminoglycoside resistances conferred by the G1491U/C substitutions is markedly increased by a mutant S12 protein. The G1491U/C mutations in addition confer resistance to the unrelated antibiotic, capreomycin. S12 also interacts with the 912 region of 16S rRNA. Genetic selection of suppressors of streptomycin dependence caused by mutations at proline 90 in S12 yielded a C912U substitution in 16S rRNA. The C912U mutation on its own confers resistance to streptomycin and restricts miscoding, properties that distinguish it from a majority of the previously described error-promoting ram mutants that also reverse streptomycin dependence.

  14. Genome-wide exploration of miRNA function in mammalian muscle cell differentiation.

    Science.gov (United States)

    Polesskaya, Anna; Degerny, Cindy; Pinna, Guillaume; Maury, Yves; Kratassiouk, Gueorgui; Mouly, Vincent; Morozova, Nadya; Kropp, Jeremie; Frandsen, Niels; Harel-Bellan, Annick

    2013-01-01

    MiRNAs impact on the control of cell fate by regulating gene expression at the post-transcriptional level. Here, using mammalian muscle differentiation as a model and a phenotypic loss-of-function screen, we explored the function of miRNAs at the genome-wide level. We found that the depletion of a high number of miRNAs (63) impacted on differentiation of human muscle precursors, underscoring the importance of this post-transcriptional mechanism of gene regulation. Interestingly, a comparison with miRNA expression profiles revealed that most of the hit miRNAs did not show any significant variations of expression during differentiation. These constitutively expressed miRNAs might be required for basic and/or essential cell function, or else might be regulated at the post-transcriptional level. MiRNA inhibition yielded a variety of phenotypes, reflecting the widespread miRNA involvement in differentiation. Using a functional screen (the STarS--Suppressor Target Screen--approach, i. e. concomitant knockdown of miRNAs and of candidate target proteins), we discovered miRNA protein targets that are previously uncharacterized controllers of muscle-cell terminal differentiation. Our results provide a strategy for functional annotation of the human miRnome.

  15. A second essential function of the Est1-binding arm of yeast telomerase RNA.

    Science.gov (United States)

    Lebo, Kevin J; Niederer, Rachel O; Zappulla, David C

    2015-05-01

    The enzymatic ribonucleoprotein telomerase maintains telomeres in many eukaryotes, including humans, and plays a central role in aging and cancer. Saccharomyces cerevisiae telomerase RNA, TLC1, is a flexible scaffold that tethers telomerase holoenzyme protein subunits to the complex. Here we test the hypothesis that a lengthy conserved region of the Est1-binding TLC1 arm contributes more than simply Est1-binding function. We separated Est1 binding from potential other functions by tethering TLC1 to Est1 via a heterologous RNA-protein binding module. We find that Est1-tethering rescues in vivo function of telomerase RNA alleles missing nucleotides specifically required for Est1 binding, but not those missing the entire conserved region. Notably, however, telomerase function is restored for this condition by expressing the arm of TLC1 in trans. Mutational analysis shows that the Second Essential Est1-arm Domain (SEED) maps to an internal loop of the arm, which SHAPE chemical mapping and 3D modeling suggest could be regulated by conformational change. Finally, we find that the SEED has an essential, Est1-independent role in telomerase function after telomerase recruitment to the telomere. The SEED may be required for establishing telomere extendibility or promoting telomerase RNP holoenzyme activity. © 2015 Lebo et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  16. Simple peptides derived from the ribosomal core potentiate RNA polymerase ribozyme function

    Science.gov (United States)

    Tagami, Shunsuke; Attwater, James; Holliger, Philipp

    2017-04-01

    The emergence of functional interactions between nucleic acids and polypeptides was a key transition in the origin of life and remains at the heart of all biology. However, how and why simple non-coded peptides could have become critical for RNA function is unclear. Here, we show that putative ancient peptide segments from the cores of both ribosomal subunits enhance RNA polymerase ribozyme (RPR) function, as do derived homopolymeric peptides comprising lysine or the non-proteinogenic lysine analogues ornithine or, to a lesser extent, diaminobutyric acid, irrespective of chirality or chiral purity. Lysine decapeptides enhance RPR function by promoting holoenzyme assembly through primer-template docking, accelerate RPR evolution, and allow RPR-catalysed RNA synthesis at near physiological (≥1 mM) Mg2+ concentrations, enabling templated RNA synthesis within membranous protocells. Our results outline how compositionally simple, mixed-chirality peptides may have augmented the functional potential of early RNAs and promoted the emergence of the first protocells.

  17. Structure-function studies of nucleocytoplasmic transport of retroviral genomic RNA by mRNA export factor TAP

    Science.gov (United States)

    Teplova, Marianna; Wohlbold, Lara; Khin, Nyan W.; Izaurralde, Elisa; Patel, Dinshaw J.

    2011-01-01

    Messenger RNA export is mediated by the TAP-p15 heterodimer, which belongs to the family of NTF2-like export receptors. TAP-p15 heterodimers also bind to the constitutive transport element (CTE) present in simian type D retroviral RNAs, and mediate export of viral unspliced RNAs to the host cytoplasm. We have solved the crystal structure of the RNA recognition and leucine-rich repeat motifs of TAP bound to one symmetrical-half of CTE RNA. L-shaped conformations of protein and RNA are involved in a mutual molecular embrace on complex formation. We have monitored the impact of structure-guided mutations on binding affinities in vitro and transport assays in vivo. Our studies define the principles by which CTE RNA subverts the mRNA export receptor TAP, thereby facilitating nuclear export of viral genomic RNAs, and more generally, provide insights on cargo RNA recognition by mRNA export receptors. PMID:21822283

  18. Carboxylesterase 1 gene duplication and mRNA expression in adipose tissue are linked to obesity and metabolic function

    DEFF Research Database (Denmark)

    Friedrichsen, Martin; Poulsen, Pernille; Wojtaszewski, Jørgen

    2013-01-01

    involved in the control of mRNA expression. Here, we investigated mRNA expression level in adipose tissue and its association with measures of adiposity and metabolic function in a population of elderly twins. Furthermore, the heritability of mRNA expression level in adipose tissue and the effect of gene...

  19. Several RNase T2 enzymes function in induced tRNA and rRNA turnover in the ciliate Tetrahymena

    DEFF Research Database (Denmark)

    Andersen, Kasper Langebjerg; Collins, Kathleen

    2012-01-01

    RNA fragments continued to accumulate, with only a minor change in fragment profile in one strain. We therefore generated strains lacking pairwise combinations of the top three candidates for Rnt2 tRNases. Each of these strains showed a distinct starvation-specific profile of tRNA and rRNA fragment accumulation...

  20. Chromatin Dynamics and the RNA Exosome Function in Concert to Regulate Transcriptional Homeostasis

    Directory of Open Access Journals (Sweden)

    Mayuri Rege

    2015-11-01

    Full Text Available The histone variant H2A.Z is a hallmark of nucleosomes flanking promoters of protein-coding genes and is often found in nucleosomes that carry lysine 56-acetylated histone H3 (H3-K56Ac, a mark that promotes replication-independent nucleosome turnover. Here, we find that H3-K56Ac promotes RNA polymerase II occupancy at many protein-coding and noncoding loci, yet neither H3-K56Ac nor H2A.Z has a significant impact on steady-state mRNA levels in yeast. Instead, broad effects of H3-K56Ac or H2A.Z on RNA levels are revealed only in the absence of the nuclear RNA exosome. H2A.Z is also necessary for the expression of divergent, promoter-proximal noncoding RNAs (ncRNAs in mouse embryonic stem cells. Finally, we show that H2A.Z functions with H3-K56Ac to facilitate formation of chromosome interaction domains (CIDs. Our study suggests that H2A.Z and H3-K56Ac work in concert with the RNA exosome to control mRNA and ncRNA expression, perhaps in part by regulating higher-order chromatin structures.

  1. Systematic Study of Drosophila MicroRNA Functions Using a Collection of Targeted Knockout Mutations

    DEFF Research Database (Denmark)

    Chen, Ya-Wen; Song, Shilin; Weng, Ruifen

    2014-01-01

    MicroRNAs are abundant in animal genomes, yet little is known about their functions in vivo. Here, we report the production of 80 new Drosophila miRNA mutants by targeted homologous recombination. These mutants remove 104 miRNAs. Together with 15 previously reported mutants, this collection inclu...... analysis of the biological roles of Drosophila miRNAs....

  2. Sheep oocyte expresses leptin and functional leptin receptor mRNA

    Directory of Open Access Journals (Sweden)

    Seyyed Jalil Taheri

    2016-09-01

    Conclusions: The result of present study reveals that leptin and its functional receptor (Ob-Rb mRNA are expressed in sheep oocyte and further studies should investigate the role(s of leptin on sheep oocyte physiology and embryo development.

  3. LNA-modified oligonucleotides mediate specific inhibition of microRNA function

    DEFF Research Database (Denmark)

    Ørom, Ulf Andersson; Kauppinen, Sakari; Lund, Anders H

    2006-01-01

    microRNAs are short, endogenous non-coding RNAs that act as post-transcriptional modulators of gene expression. Important functions for microRNAs have been found in the regulation of development, cellular proliferation and differentiation, while perturbed miRNA expression patterns have been...

  4. A Total Variation regulator for the extraction of step function depth profiles from ARXPS data

    International Nuclear Information System (INIS)

    Paynter, R.W.

    2015-01-01

    Highlights: • A Total Variation (TV) regulator for the inversion of ARXPS data was evaluated. • Simulations were performed to compare the TV regulator with a 1st-order Tikhonov regulator. • The TV regulator performed well on single step-function depth profiles. - Abstract: Simulations were conducted to generate and invert noisy ARXPS data, starting from known pseudo-step-function depth profiles. In the recovery of the depth profiles, a 1st-order Tikhonov regulator was compared with a Total Variation regulator. It was found that the profiles extracted using the Total Variation regulator tended to better resemble the true profile in the case of a single step function.

  5. Characterization of the TRBP domain required for Dicer interaction and function in RNA interference

    Directory of Open Access Journals (Sweden)

    El Far Mohamed

    2009-05-01

    Full Text Available Abstract Background Dicer, Ago2 and TRBP are the minimum components of the human RNA-induced silencing complex (RISC. While Dicer and Ago2 are RNases, TRBP is the double-stranded RNA binding protein (dsRBP that loads small interfering RNA into the RISC. TRBP binds directly to Dicer through its C-terminal domain. Results We show that the TRBP binding site in Dicer is a 165 amino acid (aa region located between the ATPase and the helicase domains. The binding site in TRBP is a 69 aa domain, called C4, located at the C-terminal end of TRBP. The TRBP1 and TRBP2 isoforms, but not TRBPs lacking the C4 site (TRBPsΔC4, co-immunoprecipitated with Dicer. The C4 domain is therefore necessary to bind Dicer, irrespective of the presence of RNA. Immunofluorescence shows that while full-length TRBPs colocalize with Dicer, TRBPsΔC4 do not. tarbp2-/- cells, which do not express TRBP, do not support RNA interference (RNAi mediated by short hairpin or micro RNAs against EGFP. Both TRBPs, but not TRBPsΔC4, were able to rescue RNAi function. In human cells with low RNAi activity, addition of TRBP1 or 2, but not TRBPsΔC4, rescued RNAi function. Conclusion The mapping of the interaction sites between TRBP and Dicer show unique domains that are required for their binding. Since TRBPsΔC4 do not interact or colocalize with Dicer, we suggest that TRBP and Dicer, both dsRBPs, do not interact through bound dsRNA. TRBPs, but not TRBPsΔC4, rescue RNAi activity in RNAi-compromised cells, indicating that the binding of Dicer to TRBP is critical for RNAi function.

  6. Alteration of protein function by a silent polymorphism linked to tRNA abundance.

    Directory of Open Access Journals (Sweden)

    Sebastian Kirchner

    2017-05-01

    Full Text Available Synonymous single nucleotide polymorphisms (sSNPs are considered neutral for protein function, as by definition they exchange only codons, not amino acids. We identified an sSNP that modifies the local translation speed of the cystic fibrosis transmembrane conductance regulator (CFTR, leading to detrimental changes to protein stability and function. This sSNP introduces a codon pairing to a low-abundance tRNA that is particularly rare in human bronchial epithelia, but not in other human tissues, suggesting tissue-specific effects of this sSNP. Up-regulation of the tRNA cognate to the mutated codon counteracts the effects of the sSNP and rescues protein conformation and function. Our results highlight the wide-ranging impact of sSNPs, which invert the programmed local speed of mRNA translation and provide direct evidence for the central role of cellular tRNA levels in mediating the actions of sSNPs in a tissue-specific manner.

  7. Alteration of protein function by a silent polymorphism linked to tRNA abundance.

    Science.gov (United States)

    Kirchner, Sebastian; Cai, Zhiwei; Rauscher, Robert; Kastelic, Nicolai; Anding, Melanie; Czech, Andreas; Kleizen, Bertrand; Ostedgaard, Lynda S; Braakman, Ineke; Sheppard, David N; Ignatova, Zoya

    2017-05-01

    Synonymous single nucleotide polymorphisms (sSNPs) are considered neutral for protein function, as by definition they exchange only codons, not amino acids. We identified an sSNP that modifies the local translation speed of the cystic fibrosis transmembrane conductance regulator (CFTR), leading to detrimental changes to protein stability and function. This sSNP introduces a codon pairing to a low-abundance tRNA that is particularly rare in human bronchial epithelia, but not in other human tissues, suggesting tissue-specific effects of this sSNP. Up-regulation of the tRNA cognate to the mutated codon counteracts the effects of the sSNP and rescues protein conformation and function. Our results highlight the wide-ranging impact of sSNPs, which invert the programmed local speed of mRNA translation and provide direct evidence for the central role of cellular tRNA levels in mediating the actions of sSNPs in a tissue-specific manner.

  8. Glycine receptors caught between genome and proteome - functional implications of RNA editing and splicing

    Directory of Open Access Journals (Sweden)

    Pascal Legendre

    2009-11-01

    Full Text Available Information processing in the brain requires a delicate balance between excitation and inhibition. Glycine receptors (GlyR are involved in inhibitory mechanisms mainly at a synaptic level, but potential novel roles for these receptors recently emerged due to the discovery of posttranscriptional processing. GLR transcripts are edited through enzymatic modification of a single nucleotide leading to amino acid substitution within the neurotransmitter binding domain. RNA editing produces gain-of-function receptors well suited for generation and maintenance of tonic inhibition of neuronal excitability. As neuronal activity deprivation in early stages of development or in epileptic tissue is detrimental to neurons and because RNA editing of GlyR is up-regulated in temporal lobe epilepsy patients with a severe course of disease a pathophysiological role of these receptors emerges. This review contains a state-of-the-art discussion of (pathophysiological implications of GlyR RNA editing.

  9. Early patient-reported outcomes versus objective function after total hip and knee arthroplasty

    DEFF Research Database (Denmark)

    Luna, I E; Kehlet, H; Peterson, B

    2017-01-01

    AIMS: The purpose of this study was to assess early physical function after total hip or knee arthroplasty (THA/TKA), and the correlation between patient-reported outcome measures, physical performance and actual physical activity (measured by actigraphy). PATIENTS AND METHODS: A total of 80 pati...

  10. Function and anatomy of plant siRNA pools derived from hairpin transgenes

    Directory of Open Access Journals (Sweden)

    Lee Kevin AW

    2007-11-01

    Full Text Available Abstract Background RNA interference results in specific gene silencing by small-interfering RNAs (siRNAs. Synthetic siRNAs provide a powerful tool for manipulating gene expression but high cost suggests that novel siRNA production methods are desirable. Strong evolutionary conservation of siRNA structure suggested that siRNAs will retain cross-species function and that transgenic plants expressing heterologous siRNAs might serve as useful siRNA bioreactors. Here we report a detailed evaluation of the above proposition and present evidence regarding structural features of siRNAs extracted from plants. Results Testing the gene silencing capacity of plant-derived siRNAs in mammalian cells proved to be very challenging and required partial siRNA purification and design of a highly sensitive assay. Using the above assay we found that plant-derived siRNAs are ineffective for gene silencing in mammalian cells. Plant-derived siRNAs are almost exclusively double-stranded and most likely comprise a mixture of bona fide siRNAs and aberrant partially complementary duplexes. We also provide indirect evidence that plant-derived siRNAs may contain a hitherto undetected physiological modification, distinct from 3' terminal 2-O-methylation. Conclusion siRNAs produced from plant hairpin transgenes and extracted from plants are ineffective for gene silencing in mammalian cells. Thus our findings establish that a previous claim that transgenic plants offer a cost-effective, scalable and sustainable source of siRNAs is unwarranted. Our results also indicate that the presence of aberrant siRNA duplexes and possibly a plant-specific siRNA modification, compromises the gene silencing capacity of plant-derived siRNAs in mammalian cells.

  11. Walking the interactome to identify human miRNA-disease associations through the functional link between miRNA targets and disease genes

    Science.gov (United States)

    2013-01-01

    Background MicroRNAs (miRNAs) are important post-transcriptional regulators that have been demonstrated to play an important role in human diseases. Elucidating the associations between miRNAs and diseases at the systematic level will deepen our understanding of the molecular mechanisms of diseases. However, miRNA-disease associations identified by previous computational methods are far from completeness and more effort is needed. Results We developed a computational framework to identify miRNA-disease associations by performing random walk analysis, and focused on the functional link between miRNA targets and disease genes in protein-protein interaction (PPI) networks. Furthermore, a bipartite miRNA-disease network was constructed, from which several miRNA-disease co-regulated modules were identified by hierarchical clustering analysis. Our approach achieved satisfactory performance in identifying known cancer-related miRNAs for nine human cancers with an area under the ROC curve (AUC) ranging from 71.3% to 91.3%. By systematically analyzing the global properties of the miRNA-disease network, we found that only a small number of miRNAs regulated genes involved in various diseases, genes associated with neurological diseases were preferentially regulated by miRNAs and some immunological diseases were associated with several specific miRNAs. We also observed that most diseases in the same co-regulated module tended to belong to the same disease category, indicating that these diseases might share similar miRNA regulatory mechanisms. Conclusions In this study, we present a computational framework to identify miRNA-disease associations, and further construct a bipartite miRNA-disease network for systematically analyzing the global properties of miRNA regulation of disease genes. Our findings provide a broad perspective on the relationships between miRNAs and diseases and could potentially aid future research efforts concerning miRNA involvement in disease pathogenesis

  12. A Role for the F-Box Protein HAWAIIAN SKIRT in Plant microRNA Function.

    Science.gov (United States)

    Lang, Patricia L M; Christie, Michael D; Dogan, Ezgi S; Schwab, Rebecca; Hagmann, Jörg; van de Weyer, Anna-Lena; Scacchi, Emanuele; Weigel, Detlef

    2018-01-01

    As regulators of gene expression in multicellular organisms, microRNAs (miRNAs) are crucial for growth and development. Although a plethora of factors involved in their biogenesis and action in Arabidopsis ( Arabidopsis thaliana ) has been described, these processes and their fine-tuning are not fully understood. Here, we used plants expressing an artificial miRNA target mimic (MIM) to screen for negative regulators of miR156. We identified a new mutant allele of the F-box gene HAWAIIAN SKIRT ( HWS ; At3G61590), hws - 5 , as a suppressor of the MIM156 -induced developmental and molecular phenotypes. In hws plants, levels of some endogenous miRNAs are increased and their mRNA targets decreased. Plants constitutively expressing full-length HWS-but not a truncated version lacking the F-box domain-display morphological and molecular phenotypes resembling those of mutants defective in miRNA biogenesis and activity. In combination with such mutants, hws loses its delayed floral organ abscission ("skirt") phenotype, suggesting epistasis. Also, the hws transcriptome profile partially resembles those of well-known miRNA mutants hyl1 - 2 , se - 3 , and ago1 - 27 , pointing to a role in a common pathway. We thus propose HWS as a novel, F-box dependent factor involved in miRNA function. © 2018 American Society of Plant Biologists. All Rights Reserved.

  13. MicroRNA function and dysregulation in bone tumors: the evidence to date.

    LENUS (Irish Health Repository)

    Nugent, Mary

    2014-01-01

    Micro ribonucleic acids (miRNAs) are small non-coding RNA segments that have a role in the regulation of normal cellular development and proliferation including normal osteogenesis. They exert their effects through inhibition of specific target genes at the post-transcriptional level. Many miRNAs have altered expression levels in cancer (either increased or decreased depending on the specific miRNA). Altered miRNA expression profiles have been identified in several malignancies including primary bone tumors such as osteosarcoma and Ewing\\'s sarcoma. It is thought that they may function as tumor suppressor genes or oncogenes and hence when dysregulated contribute to the initiation and progression of malignancy. miRNAs are also thought to have a role in the development of bone metastases in other malignancies. In addition, evidence increasingly suggests that miRNAs may play a part in determining the response to chemotherapy in the treatment of osteosarcoma. These molecules are readily detectable in tissues, both fresh and formalin fixed paraffin embedded and, more recently, in blood. Although there are fewer published studies regarding circulating miRNA profiles, they appear to reflect changes in tissue expression. Thus miRNAs may serve as potential indicators of disease presence but more importantly, may have a role in disease characterization or as potential therapeutic targets. This review gives a brief overview of miRNA biochemistry and explores the evidence to date implicating these small molecules in the pathogenesis of bone tumors.

  14. Structure-function relationship of substituted bromomethylcoumarins in nucleoside specificity of RNA alkylation.

    Science.gov (United States)

    Kellner, Stefanie; Kollar, Laura Bettina; Ochel, Antonia; Ghate, Manjunath; Helm, Mark

    2013-01-01

    Selective alkylation of RNA nucleotides is an important field of RNA biochemistry, e.g. in applications of fluorescent labeling or in structural probing experiments, yet detailed structure-function studies of labeling agents are rare. Here, bromomethylcoumarins as reactive compounds for fluorescent labeling of RNA are developed as an attractive scaffold on which electronic properties can be modulated by varying the substituents. Six different 4-bromomethyl-coumarins of various substitution patterns were tested for nucleotide specificity of RNA alkylation using tRNA from Escherichia coli as substrate. Using semi-quantitative LC-MS/MS analysis, reactions at mildly acidic and slightly alkaline pH were compared. For all tested compounds, coumarin conjugates with 4-thiouridine, pseudouridine, guanosine, and uridine were identified, with the latter largely dominating. This data set shows that selectivity of ribonucleotide alkylation depends on the substitution pattern of the reactive dye, and even more strongly on the modulation of the reaction conditions. The latter should be therefore carefully optimized when striving to achieve selectivity. Interestingly, the highest selectivity for labeling of a modified nucleoside, namely of 4-thiouridine, was achieved with a compound whose selectivity was somewhat less dependent on reaction conditions than the other compounds. In summary, bromomethylcoumarin derivatives are a highly interesting class of compounds, since their selectivity for 4-thiouridine can be efficiently tuned by variation of substitution pattern and reaction conditions.

  15. Kinetics of lipid-nanoparticle-mediated intracellular mRNA delivery and function

    Science.gov (United States)

    Zhdanov, Vladimir P.

    2017-10-01

    mRNA delivery into cells forms the basis for one of the new and promising ways to treat various diseases. Among suitable carriers, lipid nanoparticles (LNPs) with a size of about 100 nm are now often employed. Despite high current interest in this area, the understanding of the basic details of LNP-mediated mRNA delivery and function is limited. To clarify the kinetics of mRNA release from LNPs, the author uses three generic models implying (i) exponential, (ii) diffusion-controlled, and (iii) detachment-controlled kinetic regimes, respectively. Despite the distinct differences in these kinetics, the associated transient kinetics of mRNA translation to the corresponding protein and its degradation are shown to be not too sensitive to the details of the mRNA delivery by LNPs (or other nanocarriers). In addition, the author illustrates how this protein may temporarily influence the expression of one gene or a few equivalent genes. The analysis includes positive or negative regulation of the gene transcription via the attachment of the protein without or with positive or negative feedback in the gene expression. Stable, bistable, and oscillatory schemes have been scrutinized in this context.

  16. Structure and Biological Function of the RNA Pyrophosphohydrolase BdRppH from Bdellovibrio bacteriovorus

    Energy Technology Data Exchange (ETDEWEB)

    Messing, S.; Gabelli, S; Liu, Q; Celesnik, H; Belasco, J; Pineiro, S; Amzel, L

    2009-01-01

    Until recently, the mechanism of mRNA decay in bacteria was thought to be different from that of eukaryotes. This paradigm changed with the discovery that RppH (ORF176/NudH/YgdP), an Escherichia coli enzyme that belongs to the Nudix superfamily, is an RNA pyrophosphohydrolase that initiates mRNA decay by cleaving pyrophosphate from the 5?-triphosphate. Here we report the 1.9 A resolution structure of the Nudix hydrolase BdRppH from Bdellovibrio bacteriovorus, a bacterium that feeds on other Gram-negative bacteria. Based on the structure of the enzyme alone and in complex with GTP-Mg2+, we propose a mode of RNA binding similar to that of the nuclear decapping enzyme from Xenopus laevis, X29. In additional experiments, we show that BdRppH can indeed function in vitro and in vivo as an RNA pyrophosphohydrolase. These findings set the basis for the identification of possible decapping enzymes in other bacteria.

  17. Structural Analysis of ‘key’ Interactions in Functional RNA Molecules

    KAUST Repository

    Chawla, Mohit

    2018-04-01

    The main objective of the thesis is to carry out structural bioinformatics study along with usage of advanced quantum chemical methods to look at the structural stability and energetics of RNA building blocks. The main focus of the work described here lies on understanding the reasons behind the intrinsic stability of key interactions in nucleic acids. Crystal structures of RNA molecules exhibit fascinating variety of non-covalent interactions, which play an important role in maintaining the three dimensional structures. An accurate atomic level description of these interactions in the structural building blocks of RNA is a key to understand the structure-function relationship in these molecules. An effort has been made to link the conclusions drawn from quantum chemical computations on RNA base pairs in wide biochemical context of their occurrence in RNA structures. The initial attention was on the impact of natural and non-natural modifications of the nucleic acid bases on the structure and stability of base pairs that they are involved in. In the remaining sections we cover other molecular interactions shaping nucleic acids, as the interaction between ribose and the bases, and the fluoride sensing riboswitch system in order to investigate structure and dynamics of nucleic acids at the atomic level and to gain insight into the physical chemistry behind.

  18. Structure-function relationship of substituted bromomethylcoumarins in nucleoside specificity of RNA alkylation.

    Directory of Open Access Journals (Sweden)

    Stefanie Kellner

    Full Text Available Selective alkylation of RNA nucleotides is an important field of RNA biochemistry, e.g. in applications of fluorescent labeling or in structural probing experiments, yet detailed structure-function studies of labeling agents are rare. Here, bromomethylcoumarins as reactive compounds for fluorescent labeling of RNA are developed as an attractive scaffold on which electronic properties can be modulated by varying the substituents. Six different 4-bromomethyl-coumarins of various substitution patterns were tested for nucleotide specificity of RNA alkylation using tRNA from Escherichia coli as substrate. Using semi-quantitative LC-MS/MS analysis, reactions at mildly acidic and slightly alkaline pH were compared. For all tested compounds, coumarin conjugates with 4-thiouridine, pseudouridine, guanosine, and uridine were identified, with the latter largely dominating. This data set shows that selectivity of ribonucleotide alkylation depends on the substitution pattern of the reactive dye, and even more strongly on the modulation of the reaction conditions. The latter should be therefore carefully optimized when striving to achieve selectivity. Interestingly, the highest selectivity for labeling of a modified nucleoside, namely of 4-thiouridine, was achieved with a compound whose selectivity was somewhat less dependent on reaction conditions than the other compounds. In summary, bromomethylcoumarin derivatives are a highly interesting class of compounds, since their selectivity for 4-thiouridine can be efficiently tuned by variation of substitution pattern and reaction conditions.

  19. A Note on Using Unbounded Functions on Totally Bounded Sets in ...

    African Journals Online (AJOL)

    From a real-valued function f, unbounded on a totally bounded subset of a metric space, we construct a Cauchy sequence in S on which f is unbounded. Taking f to be a reciprocal Lebesgue number function, for an open cover of S, gives a rapid proof that S is compact when it is complete, without recourse to ...

  20. Comparative functional characterization of the CSR-1 22G-RNA pathway in Caenorhabditis nematodes.

    Science.gov (United States)

    Tu, Shikui; Wu, Monica Z; Wang, Jie; Cutter, Asher D; Weng, Zhiping; Claycomb, Julie M

    2015-01-01

    As a champion of small RNA research for two decades, Caenorhabditis elegans has revealed the essential Argonaute CSR-1 to play key nuclear roles in modulating chromatin, chromosome segregation and germline gene expression via 22G-small RNAs. Despite CSR-1 being preserved among diverse nematodes, the conservation and divergence in function of the targets of small RNA pathways remains poorly resolved. Here we apply comparative functional genomic analysis between C. elegans and Caenorhabditis briggsae to characterize the CSR-1 pathway, its targets and their evolution. C. briggsae CSR-1-associated small RNAs that we identified by immunoprecipitation-small RNA sequencing overlap with 22G-RNAs depleted in cbr-csr-1 RNAi-treated worms. By comparing 22G-RNAs and target genes between species, we defined a set of CSR-1 target genes with conserved germline expression, enrichment in operons and more slowly evolving coding sequences than other genes, along with a small group of evolutionarily labile targets. We demonstrate that the association of CSR-1 with chromatin is preserved, and show that depletion of cbr-csr-1 leads to chromosome segregation defects and embryonic lethality. This first comparative characterization of a small RNA pathway in Caenorhabditis establishes a conserved nuclear role for CSR-1 and highlights its key role in germline gene regulation across multiple animal species. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  1. Pervasive Regulatory Functions of mRNA Structure Revealed by High-Resolution SHAPE Probing.

    Science.gov (United States)

    Mustoe, Anthony M; Busan, Steven; Rice, Greggory M; Hajdin, Christine E; Peterson, Brant K; Ruda, Vera M; Kubica, Neil; Nutiu, Razvan; Baryza, Jeremy L; Weeks, Kevin M

    2018-03-22

    mRNAs can fold into complex structures that regulate gene expression. Resolving such structures de novo has remained challenging and has limited our understanding of the prevalence and functions of mRNA structure. We use SHAPE-MaP experiments in living E. coli cells to derive quantitative, nucleotide-resolution structure models for 194 endogenous transcripts encompassing approximately 400 genes. Individual mRNAs have exceptionally diverse architectures, and most contain well-defined structures. Active translation destabilizes mRNA structure in cells. Nevertheless, mRNA structure remains similar between in-cell and cell-free environments, indicating broad potential for structure-mediated gene regulation. We find that the translation efficiency of endogenous genes is regulated by unfolding kinetics of structures overlapping the ribosome binding site. We discover conserved structured elements in 35% of UTRs, several of which we validate as novel protein binding motifs. RNA structure regulates every gene studied here in a meaningful way, implying that most functional structures remain to be discovered. Copyright © 2018 Elsevier Inc. All rights reserved.

  2. Demonstration of two novel methods for predicting functional siRNA efficiency

    Directory of Open Access Journals (Sweden)

    Shi Tieliu

    2006-05-01

    Full Text Available Abstract Background siRNAs are small RNAs that serve as sequence determinants during the gene silencing process called RNA interference (RNAi. It is well know that siRNA efficiency is crucial in the RNAi pathway, and the siRNA efficiency for targeting different sites of a specific gene varies greatly. Therefore, there is high demand for reliable siRNAs prediction tools and for the design methods able to pick up high silencing potential siRNAs. Results In this paper, two systems have been established for the prediction of functional siRNAs: (1 a statistical model based on sequence information and (2 a machine learning model based on three features of siRNA sequences, namely binary description, thermodynamic profile and nucleotide composition. Both of the two methods show high performance on the two datasets we have constructed for training the model. Conclusion Both of the two methods studied in this paper emphasize the importance of sequence information for the prediction of functional siRNAs. The way of denoting a bio-sequence by binary system in mathematical language might be helpful in other analysis work associated with fixed-length bio-sequence.

  3. RNA sequencing analysis to demonstrate Erk dependent and independent functions of Mek.

    Science.gov (United States)

    Liu, Chang; Chen, Haixia; Liu, Lin; Chen, Lingyi

    2016-03-01

    Mek inhibition and Erk knockout (KO) have quite distinct effects on pluripotency maintenance in mouse embryonic stem cells (ESCs). To test whether there is an Erk-independent function of Mek, RNA-sequencing (RNA-seq) is carried out on six samples, WT KH2 ESCs treated with or without PD0325901 (PD) for 48 h (KH2_PD and KH2, respectively), iErk1; Erk KO ESCs cultured in the presence of Dox (P0), 48 and 96 h after Dox withdrawal (P1 and P2, respectively), and iErk1; Erk KO ESCs cultured without Dox for 96 h, and treated with PD in the last 48 h (P2_PD). These RNA-seq data demonstrate that Mek inhibition has quite different effect on the transcriptional profile of mouse ESCs, compared to Erk KO. Moreover, a significant fraction of genes is regulated by Mek inhibition, regardless of the presence or absence of Erk, indicating an Erk-independent function of Mek. RNA-seq data are deposited in Gene Expression Omnibus (GEO) datasets under accession number GSE70304.

  4. saRNA guided iNOS up-regulation improves erectile function of diabetic rats.

    Science.gov (United States)

    Wang, Tao; Li, Mingchao; Yuan, Huixin; Zhan, Yin; Xu, Hua; Wang, Shaogang; Yang, Weiming; Liu, Jihong; Ye, Zhangqun; Li, Long-Cheng

    2013-08-01

    Promoter targeted saRNAs mediate sequence specific up-regulation of gene expression. We explored the therapeutic effect of RNA activation mediated iNOS gene activation on improving erectile function in a rat model of diabetes mellitus. An optimal saRNA sequence specific for iNOS promoter was cloned into an adenoviral vector, resulting in AdU6/shiNOS and AdU6/shControl. The corresponding viruses were used to transduce cultured rat cavernous smooth muscle cells. Streptozotocin induced diabetes models were established in rats and used to test the effects of intracavernous delivery of iNOS saRNA viruses on erectile function. iNOS expression in the cavernous smooth muscle cells or penile tissue of treated rats was assessed by reverse transcriptase-polymerase chain reaction and Western blot. Cyclic guanosine monophosphate was analyzed by enzyme-linked immunosorbent assay. Intracavernous pressure in response to cavernous nerve stimulation was measured using a data acquisition system on post-injection days 1, 3, 5, 7, 10 and 14. Adenovirus mediated expression of iNOS saRNA caused sustained up-regulation of iNOS in cavernous smooth muscle cells. Intracavernous injection of AdU6/shiNOS activated iNOS expression in vivo and significantly increased peak intracavernous pressure in streptozotocin induced diabetic rats via nitric oxide/intracellular cyclic guanosine monophosphate activation. Results show that saRNA mediated iNOS over expression in the penis can restore erectile function in streptozocin diabetic rats via the nitric oxide-cyclic guanosine monophosphate pathway. Copyright © 2013 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

  5. Effect of pre-operative neuromuscular training on functional outcome after total knee replacement

    DEFF Research Database (Denmark)

    Huber, Erika O; de Bie, Rob A; Roos, Ewa M.

    2013-01-01

    Total Knee Replacement (TKR) is the standard treatment for patients with severe knee osteoarthritis (OA). Significant improvement in pain and function are seen after TKR and approximately 80% of patients are very satisfied with the outcome. Functional status prior to TKR is a major predictor...... of outcome after the intervention. Thus, improving functional status prior to surgery through exercise may improve after surgery outcome. However, results from several previous trials testing the concept have been inconclusive after surgery....

  6. Flexible and Versatile as a Chameleon—Sophisticated Functions of microRNA-199a

    Directory of Open Access Journals (Sweden)

    Shen Gu

    2012-07-01

    Full Text Available Although widely studied in the past decade, our knowledge of the functional role of microRNAs (miRNAs remains limited. Among the many miRNAs identified in humans, we focus on miR-199a due to its varied and important functions in diverse models and systems. Its expression is finely regulated by promoter methylation and direct binding of transcription factors such as TWIST1. During tumorigenesis, depending on the nature of the cancer, miR-199a, especially its -3p mature form, may act as either a potential tumor suppressor or an oncogene. Its 5p mature form has been shown to protect cardiomyocytes from hypoxic damage via its action on HIF1α. It also has a functional role in stem cell differentiation, embryo development, hepatitis, liver fibrosis, etc. Though it has varied biological activities, its regulation has not been reviewed. The varied and protean functions of miR-199a suggest that efforts to generalize the action of a miRNA are problematic. This review provides a comprehensive survey of the literature on miR-199a as an example of the complexity of miRNA biology and suggests future directions for miRNA research.

  7. Site-Specific Incorporation of Functional Components into RNA by an Unnatural Base Pair Transcription System

    Directory of Open Access Journals (Sweden)

    Rie Kawai

    2012-03-01

    Full Text Available Toward the expansion of the genetic alphabet, an unnatural base pair between 7-(2-thienylimidazo[4,5-b]pyridine (Ds and pyrrole-2-carbaldehyde (Pa functions as a third base pair in replication and transcription, and provides a useful tool for the site-specific, enzymatic incorporation of functional components into nucleic acids. We have synthesized several modified-Pa substrates, such as alkylamino-, biotin-, TAMRA-, FAM-, and digoxigenin-linked PaTPs, and examined their transcription by T7 RNA polymerase using Ds-containing DNA templates with various sequences. The Pa substrates modified with relatively small functional groups, such as alkylamino and biotin, were efficiently incorporated into RNA transcripts at the internal positions, except for those less than 10 bases from the 3′-terminus. We found that the efficient incorporation into a position close to the 3′-terminus of a transcript depended on the natural base contexts neighboring the unnatural base, and that pyrimidine-Ds-pyrimidine sequences in templates were generally favorable, relative to purine-Ds-purine sequences. The unnatural base pair transcription system provides a method for the site-specific functionalization of large RNA molecules.

  8. LncRNA-HIT Functions as an Epigenetic Regulator of Chondrogenesis through Its Recruitment of p100/CBP Complexes.

    Directory of Open Access Journals (Sweden)

    Hanqian L Carlson

    2015-12-01

    Full Text Available Gene expression profiling in E 11 mouse embryos identified high expression of the long noncoding RNA (lncRNA, LNCRNA-HIT in the undifferentiated limb mesenchyme, gut, and developing genital tubercle. In the limb mesenchyme, LncRNA-HIT was found to be retained in the nucleus, forming a complex with p100 and CBP. Analysis of the genome-wide distribution of LncRNA-HIT-p100/CBP complexes by ChIRP-seq revealed LncRNA-HIT associated peaks at multiple loci in the murine genome. Ontological analysis of the genes contacted by LncRNA-HIT-p100/CBP complexes indicate a primary role for these loci in chondrogenic differentiation. Functional analysis using siRNA-mediated reductions in LncRNA-HIT or p100 transcripts revealed a significant decrease in expression of many of the LncRNA-HIT-associated loci. LncRNA-HIT siRNA treatments also impacted the ability of the limb mesenchyme to form cartilage, reducing mesenchymal cell condensation and the formation of cartilage nodules. Mechanistically the LncRNA-HIT siRNA treatments impacted pro-chondrogenic gene expression by reducing H3K27ac or p100 activity, confirming that LncRNA-HIT is essential for chondrogenic differentiation in the limb mesenchyme. Taken together, these findings reveal a fundamental epigenetic mechanism functioning during early limb development, using LncRNA-HIT and its associated proteins to promote the expression of multiple genes whose products are necessary for the formation of cartilage.

  9. Long-term, efficient inhibition of microRNA function in mice using rAAV vectors.

    Science.gov (United States)

    Xie, Jun; Ameres, Stefan L; Friedline, Randall; Hung, Jui-Hung; Zhang, Yu; Xie, Qing; Zhong, Li; Su, Qin; He, Ran; Li, Mengxin; Li, Huapeng; Mu, Xin; Zhang, Hongwei; Broderick, Jennifer A; Kim, Jason K; Weng, Zhiping; Flotte, Terence R; Zamore, Phillip D; Gao, Guangping

    2012-03-04

    Understanding the function of individual microRNA (miRNA) species in mice would require the production of hundreds of loss-of-function strains. To accelerate analysis of miRNA biology in mammals, we combined recombinant adeno-associated virus (rAAV) vectors with miRNA 'tough decoys' (TuDs) to inhibit specific miRNAs. Intravenous injection of rAAV9 expressing anti-miR-122 or anti-let-7 TuDs depleted the corresponding miRNA and increased its mRNA targets. rAAV producing anti-miR-122 TuD but not anti-let-7 TuD reduced serum cholesterol by >30% for 25 weeks in wild-type mice. High-throughput sequencing of liver miRNAs from the treated mice confirmed that the targeted miRNAs were depleted and revealed that TuDs induced miRNA tailing and trimming in vivo. rAAV-mediated miRNA inhibition thus provides a simple way to study miRNA function in adult mammals and a potential therapy for dyslipidemia and other diseases caused by miRNA deregulation.

  10. A comparison of six total rna isolation methods for diagnosis of gysvd-1 (grapevine yellow speckle viroid-1) on vitis vinifera L. leaves

    International Nuclear Information System (INIS)

    Paylan, I.C.; Candar, A.; Gumus, M.; Pazarlar, S.

    2014-01-01

    Isolation of high quality RNA from plant tissues is one of the most critical steps for the successful application of diagnostic tests such as reverse transcription polymerase chain reaction (RT-PCR), northern blotting, microarray hybridization. The presence of inhibitors such as secondary metabolites, phenolic compounds and RNAses can cause inaccurate and undesirable results. Grapevine is rich in a wide range of metabolites which interfere with RNA isolation. From this point of view, we researched six different total RNA extraction methods on leaves of Vitisvinifera L. to find the best one that contribute the purity and high quality. The methods tested are silica-capture, modified silica-capture, commercial kit, the new combined, lithium chloride and citric buffer. RNA quality was analyzed spectrophotometrically by nanodrop, agarose gel electrophoresis and RT-PCR. As a result of all, it is clear that the most suitable TNA isolation protocol is the new combined method which experienced and named firstly by us, in terms of RNA purity, concentration, less time consuming of isolation step and achievement on detection of GYSVd-1. (author)

  11. The influence of optimism on functionality after total hip replacement surgery.

    Science.gov (United States)

    Balck, Friedrich; Lippmann, Maike; Jeszenszky, Csilla; Günther, Klaus-Peter; Kirschner, Stephan

    2016-08-01

    Among other factors, optimism has been shown to significantly influence the course of some diseases (cancer, HIV, coronary heart disease). This study investigated whether optimism of a patient before a total hip replacement can predict the functionality of the lower limbs 3 and 6 months after surgery. A total of 325 patients took part in the study (age: 58.7 years; w: 55%). The functionality was measured with the Western Ontario and McMaster Universities arthrosis index, and optimism with the Life Orientation Test. To analyse the influences of age, gender and optimism, general linear models were calculated. In optimistic patients, functionality improved significantly over time. The study showed a clear influence of dispositional optimism on the recovery after total hip replacement in the first 3 months after surgery. © The Author(s) 2015.

  12. Multiple functions of DDX3 RNA helicase in gene regulation, tumorigenesis and viral infection

    Directory of Open Access Journals (Sweden)

    YASUO eARIUMI

    2014-12-01

    Full Text Available The DEAD-box RNA helicase DDX3 is a multifunctional protein involved in all aspects of RNA metabolism, including transcription, splicing, mRNA nuclear export, translation, RNA decay and ribosome biogenesis. In addition, DDX3 is also implicated in cell cycle regulation, apoptosis, Wnt-ß-catenin signaling, tumorigenesis, and viral infection. Notably, recent studies suggest that DDX3 is a component of anti-viral innate immune signaling pathways. Indeed, DDX3 contributes to enhance the induction of anti-viral mediators, interferon regulatory factor (IRF 3 and type I interferon (IFN. However, DDX3 seems to be an important target for several viruses, such as human immunodeficiency virus (HIV-1, hepatitis C virus (HCV, hepatitis B virus (HBV, and poxvirus. DDX3 interacts with HIV-1 Rev or HCV Core protein and modulates its function. At least, DDX3 is required for both HIV-1 and HCV replication. Therefore, DDX3 could be a novel therapeutic target for the development of drug against HIV-1 and HCV.

  13. Total hip arthroplasty in the elderly: impact on functional performance Artroplastia total de quadril em idosos: impacto na funcionalidade

    Directory of Open Access Journals (Sweden)

    Rita C. Guedes

    2011-04-01

    Full Text Available OBJECTIVES: To compare gait and functional performance parameters in elderly subjects who had or had not total hip arthroplasty (THA. METHODS: Our sample comprised 23 elderly subjects (72±6.5 years of age with a mean of 2.6±1.3 years following THA, named the arthroplasty group (AG and 23 asymptomatic elderly subjects (70.1±5.9 years of age, named the control group (CG. Case and control subjects were paired by gender, age, body mass index and level of physical activity. The GAITRite® system was used to analyze various gait parameters in four situations: usual speed (US, fast speed (FS, slow speed (SS and dual task (DT; the Dynamic Gait Index (DGI and Timed Up and Go test (TUG methods were used to evaluate functional capacity. The statistical analyses were carried out using the Shapiro-Wilk test, Student's-t-tests for independent samples, chi-square tests, ANOVA for repeated measurements and paired Student's-t-tests. RESULTS: The AG had the worst results for gait speed (AG=1.18±0.13 and CG=1.39±0.09; p=0.012, symmetry index (SI of step length (AG=3.60±1.01 and CG=1.12±0.59; p=0.000, SI of step time (AG=-2.65±0.92 and CG=0.99±0.74; p=0.000, SI of stance phase (AG=-2.55 and CG=-1.04±0.50; p=0.005, SI of single support phase (AG=-2.17±0.78 and CG=1.21±0.51; p=0.003, DGI (AG=20.04±1.91 and CG=21.69±1.45; p=0.001 and TUG (AG=14.67±1.94 and CG=10.08±1.49; p=0.001. CONCLUSION: Elderly subjects with a history of THA had changes in gait parameters and lower performance in TUG test even 2.6±1,3 years after surgery, which suggests functional impairment.OBJETIVOS: Comparar os parâmetros da marcha e o desempenho funcional de idosos com e sem artroplastia total de quadril (ATQ. MÉTODOS: Foram selecionados 23 idosos (72±6,5 anos após média de 2,6±1,3 anos de ATQ e 23 idosos assintomáticos (70,1±5,9 anos, pareados por gênero, idade, índice de massa corpórea (IMC e nível de atividade física. Utilizou-se o sistema GAITRite® em quatro

  14. A simple functional form for proton-208Pb total reaction cross sections

    International Nuclear Information System (INIS)

    Majumdar, S.; Deb, P.K.; Amos, K.

    2001-01-01

    A simple functional form has been found that gives a good representation of the total reaction cross sections for the scattering from 208 Pb of protons with energies in the range 30 to 300 MeV. The ratios of the total reaction cross sections calculated under this approximations compared well (to within a few percent) to those determined from the microscopic optical model potentials

  15. Long non-coding RNA profile in mantle cell lymphoma identifies a functional lncRNA ROR1-AS1 associated with EZH2/PRC2 complex

    Science.gov (United States)

    Hu, Guangzhen; Gupta, Shiv K.; Troska, Tammy P.; Nair, Asha; Gupta, Mamta

    2017-01-01

    Mantle cell lymphoma (MCL) is an aggressive B-cell lymphoma characterized by rapid disease progression. The needs for new therapeutic strategies for MCL patients call for further understanding on the molecular mechanisms of pathogenesis of MCL. Recently, long noncoding RNAs (lncRNAs) have been recognized as key regulators of gene expression and disease development, however, the role of lncRNAs in non-Hodgkin lymphoma and specifically in MCL is still unknown. Next generation RNA-sequencing was carried out on MCL patient samples along with normal controls and data was analyzed. As a result, several novel lncRNAs were found significantly overexpressed in the MCL samples with lncRNA ROR1-AS1 the most significant one. We cloned the ROR1-AS1 lncRNA in expression vector and ectopically transfected in MCL cell lines. Results showed that overexpression of ROR1-AS1 lncRNA promoted growth of MCL cells while decreased sensitivity to the treatment with drugs ibrutinib and dexamethasone. ROR-AS1 overexpression also decreased the mRNA expression of P16 (P = 0.21), and SOX11 (p = 0.017), without much effect on P53, ATM and P14 mRNA. RNA-immunoprecipitation assays demonstrated high affinity binding of lncRNA ROR1-AS1 with EZH2 and SUZ12 proteins of the polycomb repressive complex-2 (PRC2). Suppressing EZH2 activity with pharmacological inhibitor GSK343 abolished binding of ROR1-AS1 with EZH2. Taken together, this study identified a functional lncRNA ROR-AS1 involved with regulation of gene transcription via associating with PRC2 complex, and may serve as a novel biomarker in MCL patients. PMID:29113297

  16. Alteration of protein function by a silent polymorphism linked to tRNA abundance

    OpenAIRE

    Kirchner, Sebastian; Cai, Zhiwei; Rauscher, Robert; Kastelic, Nicolai; Anding, Melanie; Czech, Andreas; Kleizen, Bertrand; Ostedgaard, Lynda S.; Braakman, Ineke; Sheppard, David N.; Ignatova, Zoya

    2017-01-01

    Synonymous single nucleotide polymorphisms (sSNPs) are considered neutral for protein function, as by definition they exchange only codons, not amino acids. We identified an sSNP that modifies the local translation speed of the cystic fibrosis transmembrane conduc-tance regulator (CFTR), leading to detrimental changes to protein stability and function. This sSNP introduces a codon pairing to a low-abundance tRNA that is particularly rare in human bronchial epithelia, but not in other human ti...

  17. Developmental and functional expression of miRNA-stability related genes in the nervous system.

    Science.gov (United States)

    de Sousa, Érica; Walter, Lais Takata; Higa, Guilherme Shigueto Vilar; Casado, Otávio Augusto Nocera; Kihara, Alexandre Hiroaki

    2013-01-01

    In the nervous system, control of gene expression by microRNAs (miRNAs) has been investigated in fundamental processes, such as development and adaptation to ambient demands. The action of these short nucleotide sequences on specific genes depends on intracellular concentration, which in turn reflects the balance of biosynthesis and degradation. Whereas mechanisms underlying miRNA biogenesis has been investigated in recent studies, little is known about miRNA-stability related proteins. We first detected two genes in the retina that have been associated to miRNA stability, XRN2 and PAPD4. These genes are highly expressed during retinal development, however with distinct subcellular localization. We investigated whether these proteins are regulated during specific phases of the cell cycle. Combined analyses of nuclei position in neuroblastic layer and labeling using anti-cyclin D1 revealed that both proteins do not accumulate in S or M phases of the cell cycle, being poorly expressed in progenitor cells. Indeed, XRN2 and PAPD4 were observed mainly after neuronal differentiation, since low expression was also observed in astrocytes, endothelial and microglial cells. XRN2 and PAPD4 are expressed in a wide variety of neurons, including horizontal, amacrine and ganglion cells. To evaluate the functional role of both genes, we carried out experiments addressed to the retinal adaptation in response to different ambient light conditions. PAPD4 is upregulated after 3 and 24 hours of dark- adaptation, revealing that accumulation of this protein is governed by ambient light levels. Indeed, the fast and functional regulation of PAPD4 was not related to changes in gene expression, disclosing that control of protein levels occurs by post-transcriptional mechanisms. Furthermore, we were able to quantify changes in PAPD4 in specific amacrine cells after dark -adaptation, suggesting for circuitry-related roles in visual perception. In summary, in this study we first described the

  18. The total cross section as a function of energy for elastic scattering of noble gas atoms

    International Nuclear Information System (INIS)

    Linse, C.A.

    1978-01-01

    Precise relative measurements of the total cross-sections as a function of velocity is presented for the systems Ar-Ar, Ar-Kr, Kr-Ar, Ar-Xe, Ne-Ar, Ne-Kr, and Ne-Xe, the primary beam particle being mentioned first. A discription of the apparatus is given. Then the method for extracting total cross-sections from the measured beam attenuation is analyzed. A comparison is made with total cross-sections calculated from various potentials that have been proposed in the literature

  19. Orthogonal use of a human tRNA synthetase active site to achieve multi-functionality

    Science.gov (United States)

    Zhou, Quansheng; Kapoor, Mili; Guo, Min; Belani, Rajesh; Xu, Xiaoling; Kiosses, William B.; Hanan, Melanie; Park, Chulho; Armour, Eva; Do, Minh-Ha; Nangle, Leslie A.; Schimmel, Paul; Yang, Xiang-Lei

    2011-01-01

    Protein multi-functionality is an emerging explanation for the complexity of higher organisms. In this regard, while aminoacyl tRNA synthetases catalyze amino acid activation for protein synthesis, some also act in pathways for inflammation, angiogenesis, and apoptosis. How multiple functions evolved and their relationship to the active site is not clear. Here structural modeling analysis, mutagenesis, and cell-based functional studies show that the potent angiostatic, natural fragment of human TrpRS associates via Trp side chains that protrude from the cognate cellular receptor VE-cadherin. Modeling indicates that (I prefer the way it was because the conclusion was reached not only by modeling, but more so by experimental studies.)VE-cadherin Trp side chains fit into the Trp-specific active site of the synthetase. Thus, specific side chains of the receptor mimic (?) amino acid substrates and expand the functionality of the active site of the synthetase. We propose that orthogonal use of the same active site may be a general way to develop multi-functionality of human tRNA synthetases and other proteins. PMID:20010843

  20. Totally Optimal Decision Trees for Monotone Boolean Functions with at Most Five Variables

    KAUST Repository

    Chikalov, Igor

    2013-01-01

    In this paper, we present the empirical results for relationships between time (depth) and space (number of nodes) complexity of decision trees computing monotone Boolean functions, with at most five variables. We use Dagger (a tool for optimization of decision trees and decision rules) to conduct experiments. We show that, for each monotone Boolean function with at most five variables, there exists a totally optimal decision tree which is optimal with respect to both depth and number of nodes.

  1. Decreased thalamocortical functional connectivity after 36 hours of total sleep deprivation: evidence from resting state FMRI.

    Directory of Open Access Journals (Sweden)

    Yongcong Shao

    Full Text Available OBJECTIVES: The thalamus and cerebral cortex are connected via topographically organized, reciprocal connections, which hold a key function in segregating internally and externally directed awareness information. Previous task-related studies have revealed altered activities of the thalamus after total sleep deprivation (TSD. However, it is still unclear how TSD impacts on the communication between the thalamus and cerebral cortex. In this study, we examined changes of thalamocortical functional connectivity after 36 hours of total sleep deprivation by using resting state function MRI (fMRI. MATERIALS AND METHODS: Fourteen healthy volunteers were recruited and performed fMRI scans before and after 36 hours of TSD. Seed-based functional connectivity analysis was employed and differences of thalamocortical functional connectivity were tested between the rested wakefulness (RW and TSD conditions. RESULTS: We found that the right thalamus showed decreased functional connectivity with the right parahippocampal gyrus, right middle temporal gyrus and right superior frontal gyrus in the resting brain after TSD when compared with that after normal sleep. As to the left thalamus, decreased connectivity was found with the right medial frontal gyrus, bilateral middle temporal gyri and left superior frontal gyrus. CONCLUSION: These findings suggest disruptive changes of the thalamocortical functional connectivity after TSD, which may lead to the decline of the arousal level and information integration, and subsequently, influence the human cognitive functions.

  2. Relationship between circulating microRNA-30c with total- and LDL-cholesterol, their circulatory transportation and effect of statins.

    Science.gov (United States)

    Sodi, Ravinder; Eastwood, Jarlath; Caslake, Muriel; Packard, Chris J; Denby, Laura

    2017-03-01

    Small non-coding microRNAs (miR) have important regulatory roles and are used as biomarkers of disease. We investigated the relationship between lipoproteins and circulating miR-30c, evaluated how they are transported in circulation and determined whether statins altered the circulating concentration of miR-30c. To determine the relationship between lipoproteins and circulating miR-30c, serum samples from 79 subjects recruited from a lipid clinic were evaluated. Ultracentrifugation and nanoparticle tracking analysis was used to evaluate the transportation of miR-30c in the circulation by lipoproteins and extracellular vesicles in three healthy volunteers. Using archived samples from previous studies, the effects of 40mg rosuvastatin (n=22) and 40mg pravastatin (n=24) on miR-30c expression was also examined. RNA extraction, reverse transcription-quantitative real-time polymerase chain reaction was carried out using standard procedures. When stratified according to total cholesterol concentration, there was increased miR-30c expression in the highest compared to the lowest tertile (p=0.035). There was significant positive correlation between miR-30c and total- (r=0.367; p=0.002) and LDL-cholesterol (r=0.391; p=0.001). We found that miR-30c was transported in both exosomes and on HDL3. There was a 3.8-fold increased expression of circulating miR-30c after pravastatin treatment for 1year (p=0.005) but no significant change with atorvastatin after 8weeks (p=0.145). This study shows for the first-time in humans that circulating miR-30c is significantly, positively correlated with total- and LDL-cholesterol implicating regulatory functions in lipid homeostasis. We show miR-30c is transported in both exosomes and on HDL3 and pravastatin therapy significantly increased circulating miR-30c expression adding to the pleiotropic dimensions of statins. Copyright © 2017 Elsevier B.V. All rights reserved.

  3. RNA interference for functional genomics and improvement of cotton (Gossypium spp.

    Directory of Open Access Journals (Sweden)

    Ibrokhim Y. Abdurakhmonov

    2016-02-01

    Full Text Available RNA interference (RNAi, is a powerful new technology in the discovery of genetic sequence functions, and has become a valuable tool for functional genomics of cotton (Gossypium ssp.. The rapid adoption of RNAi has replaced previous antisense technology. RNAi has aided in the discovery of function and biological roles of many key cotton genes involved in fiber development, fertility and somatic embryogenesis, resistance to important biotic and abiotic stresses, and oil and seed quality improvements as well as the key agronomic traits including yield and maturity. Here, we have comparatively reviewed seminal research efforts in previously used antisense approaches and currently applied breakthrough RNAi studies in cotton, analyzing developed RNAi methodologies, achievements, limitations, and future needs in functional characterizations of cotton genes. We also highlighted needed efforts in the development of RNAi-based cotton cultivars, and their safety and risk assessment, small and large-scale field trials, and commercialisation.

  4. The functional half-life of an mRNA depends on the ribosome spacing in an early coding region

    DEFF Research Database (Denmark)

    Pedersen, Margit; Nissen, Søren; Mitarai, Namiko

    2011-01-01

    Bacterial mRNAs are translated by closely spaced ribosomes and degraded from the 5'-end, with half-lives of around 2 min at 37 °C in most cases. Ribosome-free or "naked" mRNA is known to be readily degraded, but the initial event that inactivates the mRNA functionally has not been fully described...

  5. Acetabulum-Only Revision Total Hip Arthroplasty Is Associated With Good Functional Outcomes and Survivorship.

    Science.gov (United States)

    Lim, Jun Wei; Ridley, David; Johnston, Linda R; Clift, Benedict A

    2017-07-01

    The coexistence of a stable femoral and a loose acetabular component may pose a clinical dilemma for the surgeon. Our study aims at comparing the intermediate functional outcomes and survivorship of acetabulum-only revision total hip arthroplasty (ArTHA) with an age-matched and gender-matched total revision THA (TrTHA) group. We retrospectively reviewed prospectively collected data on the pain, function, and total Harris Hip Scores (HHS) and complication profile for ArTHA and TrTHA cohorts from our regional arthroplasty database. Kaplan-Meier survivorship, with the need for repeat revision surgery as the end point, was used for survival analysis. Among 538 cases, there were fewer acute medical complications in ArTHA and a similar dislocation rate for both cohorts. Preoperative HHS for pain, function, and total were better in the ArTHA cohort, but only the function score reached statistical significance. No significant differences in subsequent years for all aspects of HHS, except the function score was significantly better in the ArTHA cohort at year 1. And 10.0% of ArTHAs and 7.8% of TrTHAs had required rerevision. The 5-year survivorship was 90.3% (95% confidence interval ± 2.1%) for the ArTHA cohort and 92.7% (95% confidence interval ± 1.8%) for the TrTHA cohort (P = .394). The ArTHA with posterior approach (n = 118) group had the lowest dislocation rate and the best trend of functional outcomes. ArTHA can provide similar functional outcomes and dislocation rate to TrTHA, with an acceptable rerevision rate. The posterior approach in this study was not associated with a significant dislocation rate. Crown Copyright © 2017. Published by Elsevier Inc. All rights reserved.

  6. Functional exercise after total hip replacement (FEATHER): a randomised control trial.

    LENUS (Irish Health Repository)

    Monaghan, Brenda

    2012-11-01

    Prolonged physical impairments in range of movement, postural stability and walking speed are commonly reported following total hip replacement (THR). It is unclear from the current body of evidence what kind of exercises should be performed to maximize patient function and quality of life.

  7. Evaluation of the Function and Quality of Life after Total Hip Arthroplasty by Different Approaches

    Directory of Open Access Journals (Sweden)

    Paulo Araújo

    2017-09-01

    Conclusion: After 24 months post total hip arthroplasty there are no differences between the two approaches with regard to function or quality of life. However, the patients operated by the anterolateral approach had greater muscle strength deficits and higher percentage of positive Trendelenburg test.

  8. Knee awareness and functionality after simultaneous bilateral vs unilateral total knee arthroplasty

    DEFF Research Database (Denmark)

    Latifi, Roshan; Thomsen, Morten Grove; Kallemose, Thomas

    2016-01-01

    AIM: To investigate knee awareness and functional outcomes in patients treated with simultaneous bilateral vs unilateral total knee arthroplasty (TKA). METHODS: Through a database search, we identified 210 patients who had undergone unilateral TKA (UTKA) and 65 patients who had undergone simultan...

  9. Scapular allograft reconstruction after total scapulectomy: surgical technique and functional results

    NARCIS (Netherlands)

    Capanna, R.; Totti, F.; Geest, I.C.M. van der; Muller, D.A.

    2015-01-01

    HYPOTHESIS: Scapular allograft reconstruction after total scapulectomy preserving the rotator cuff muscles is an oncologically safe procedure and results in good functional outcome with a low complication rate. METHODS: The data of 6 patients who underwent scapular allograft reconstruction after a

  10. Experimental study of cardial function in delayed period after total radiotherapy

    International Nuclear Information System (INIS)

    Efimov, V.I.

    1983-01-01

    Data on functional cardial changes and its reactivity in experimental animals in delayed period after total exposure to different types of ionizing radiation in different dose range are analysed and generalized. Consequences of acute, subacute and chronic radiation action was revealed by electrocardiographic polycardiographic and roentgenological investigations. It is shown that many common features are observed in pictures of functional cardial changes of animals and a man in delayed period of radiation exposures. It was assumed that changes of heart reactivity can be conditioned both by changes of its receptor system and functional deviations in ve-- getative nervous system, vascular centers, endocrine, tropic and electrolyte metabolic disorders of general and local character

  11. Total and Direct Correlation Function Integrals from Molecular Simulation of Binary Systems

    DEFF Research Database (Denmark)

    Wedberg, Nils Hejle Rasmus Ingemar; O’Connell, John P.; Peters, Günther H.J.

    2011-01-01

    The possibility for obtaining derivative properties for mixtures from integrals of spatial total and direct correlation functions obtained from molecular dynamics simulations is explored. Theoretically well-supported methods are examined to extend simulation radial distribution functions to long...... are consistent with an excess Helmholtz energy model fitted to available simulations. In addition, simulations of water/methanol and water/t-butanol mixtures have been carried out. The method yields results for partial molar volumes, activity coefficient derivatives, and individual correlation function integrals...

  12. Function and Application Areas in Medicine of Non-Coding RNA

    Directory of Open Access Journals (Sweden)

    Figen Guzelgul

    2009-06-01

    Full Text Available RNA is the genetic material converting the genetic code that it gets from DNA into protein. While less than 2 % of RNA is converted into protein , more than 98 % of it can not be converted into protein and named as non-coding RNAs. 70 % of noncoding RNAs consists of introns , however, the rest part of them consists of exons. Non-coding RNAs are examined in two classes according to their size and functions. Whereas they are classified as long non-coding and small non-coding RNAs according to their size , they are grouped as housekeeping non-coding RNAs and regulating non-coding RNAs according to their function. For long years ,these non-coding RNAs have been considered as non-functional. However, today, it has been proved that these non-coding RNAs play role in regulating genes and in structural, functional and catalitic roles of RNAs converted into protein. Due to its taking a role in gene silencing mechanism, particularly in medical world , non-coding RNAs have led to significant developments. RNAi technolgy , which is used in designing drugs to be used in treatment of various diseases , is a ray of hope for medical world. [Archives Medical Review Journal 2009; 18(3.000: 141-155

  13. A novel method for accurate and reproducible functional cup positioning in total hip arthroplasty.

    Science.gov (United States)

    Meftah, Morteza; Yadav, Akhilesh; Wong, Anthony C; Ranawat, Amar S; Ranawat, Chitranjan S

    2013-08-01

    Cup positioning is an important variable for short and long term function, stability, and durability of total hip arthroplasty (THA). This novel method utilizes internal and external bony landmarks, and the transverse acetabular ligament for positioning the acetabular component. The cup is placed parallel and superior to the transverse ligament and inside the anterior wall notch of the true acetabulum, then adjusted for femoral version and pelvic tilt and obliquity based on weight bearing radiographs. In 78 consecutive THAs, the mean functional anteversion and abduction angles were 17.9° ± 4.7° and 41.7° ± 3.8°, respectively. 96% of the functional anteversion measurements and 100% of the functional abduction angles were within the safe zone. This technique is an easy, reproducible, and accurate method for functional cup placement. Copyright © 2013 Elsevier Inc. All rights reserved.

  14. Short and long-term impact of parathyroid autotransplantation on parathyroid function after total thyroidectomy

    Science.gov (United States)

    George, Robert; Sywak, Mark

    2017-01-01

    The most common complication of total thyroidectomy is parathyroid insufficiency. Acute, transient, post-operative hypoparathyroidism increases length of hospitalization, morbidity and cost associated with total thyroidectomy. While permanent hypoparathyroidism poses a significant medical burden with lifetime medication, regular follow up and considerable disease burden related to chronic renal failure and other sequelae. Parathyroid autotransplantation has been demonstrated to result in biochemically functional grafts, leading to the procedures’ common use during total thyroidectomy. The clearest indications for parathyroid auto transplantation are inadvertently removed or devascularized parathyroid glands. Some centers utilize routine autotransplantation to reduce the risk of permanent hypoparathyroidism. Novel fluorescence techniques to aid in parathyroid detection during thyroid surgery are under evaluation. This review aims to define the role and impact of parathyroid autotransplantation undertaken during total thyroidectomy. PMID:29322025

  15. Respective Functions of Two Distinct Siwi Complexes Assembled during PIWI-Interacting RNA Biogenesis in Bombyx Germ Cells

    Directory of Open Access Journals (Sweden)

    Kazumichi M. Nishida

    2015-01-01

    Full Text Available PIWI-interacting RNA (piRNA biogenesis consists of two sequential steps: primary piRNA processing and the ping-pong cycle that depends on reciprocal Slicer-mediated RNA cleavage by PIWI proteins. However, the molecular functions of the factors involved remain elusive. Here, we show that RNAs cleaved by a Bombyx mori PIWI, Siwi, remain bound to the protein upon cleavage but are released by a DEAD box protein BmVasa. BmVasa copurifies with Siwi but not another PIWI BmAgo3. A lack of BmVasa does not affect primary piRNA processing but abolishes the ping-pong cycle. Siwi also forms a complex with BmSpn-E and BmQin. This complex is physically separable from the Siwi/BmVasa complex. BmSpn-E, unlike BmVasa, is necessary for primary piRNA production. We propose a model for piRNA biogenesis, where the BmSpn-E/BmQin dimer binds Siwi to function in primary piRNA processing, whereas BmVasa, by associating with Siwi, ensures target RNA release upon cleavage to facilitate the ping-pong cycle.

  16. Structural similarities and functional differences clarify evolutionary relationships between tRNA healing enzymes and the myelin enzyme CNPase.

    Science.gov (United States)

    Muruganandam, Gopinath; Raasakka, Arne; Myllykoski, Matti; Kursula, Inari; Kursula, Petri

    2017-05-16

    Eukaryotic tRNA splicing is an essential process in the transformation of a primary tRNA transcript into a mature functional tRNA molecule. 5'-phosphate ligation involves two steps: a healing reaction catalyzed by polynucleotide kinase (PNK) in association with cyclic phosphodiesterase (CPDase), and a sealing reaction catalyzed by an RNA ligase. The enzymes that catalyze tRNA healing in yeast and higher eukaryotes are homologous to the members of the 2H phosphoesterase superfamily, in particular to the vertebrate myelin enzyme 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase). We employed different biophysical and biochemical methods to elucidate the overall structural and functional features of the tRNA healing enzymes yeast Trl1 PNK/CPDase and lancelet PNK/CPDase and compared them with vertebrate CNPase. The yeast and the lancelet enzymes have cyclic phosphodiesterase and polynucleotide kinase activity, while vertebrate CNPase lacks PNK activity. In addition, we also show that the healing enzymes are structurally similar to the vertebrate CNPase by applying synchrotron radiation circular dichroism spectroscopy and small-angle X-ray scattering. We provide a structural analysis of the tRNA healing enzyme PNK and CPDase domains together. Our results support evolution of vertebrate CNPase from tRNA healing enzymes with a loss of function at its N-terminal PNK-like domain.

  17. A novel Drosophila antisense scaRNA with a predicted guide function.

    Science.gov (United States)

    Tortoriello, Giuseppe; Accardo, Maria Carmela; Scialò, Filippo; Angrisani, Alberto; Turano, Mimmo; Furia, Maria

    2009-05-01

    A significant portion of eukaryotic small ncRNA transcriptome is composed by small nucleolar RNAs. From archaeal to mammalian cells, these molecules act as guides in the site-specific pseudouridylation or methylation of target RNAs. We used a bioinformatics search program to detect Drosophila putative orthologues of U79, one out of ten snoRNAs produced by GAS5, a human ncRNA involved in apoptosis, susceptibility to cancer and autoimmune diseases. This search led to the definition of a list of U79-related fly snoRNAs whose genomic organization, evolution and expression strategy are discussed here. We report that an intriguing novel specimen, named Dm46E3, is transcribed as a longer, unspliced precursor from the reverse strand of eiger, a fly regulatory gene that plays a key role in cell differentiation, apoptosis and immune response. Expression of Dm46E3 was found significantly up-regulated in a mutant strain in which eiger transcription is greatly reduced, suggesting that these two sense-antisense genes may be mutually regulated. Relevant to its function, Dm46E3 concentrated specifically in the Cajal bodies, followed a dynamic spatial expression profile during embryogenesis and displayed a degenerate antisense element that enables it to target U1b, a developmentally regulated isoform of the U1 spliceosomal snRNA that is particularly abundant in embryos.

  18. Ionization and fragmentation of DNA-RNA bases: a density functional theory study

    International Nuclear Information System (INIS)

    Sadr-Arani, Leila

    2014-01-01

    Ionizing radiation (IR) cross human tissue, deposit energy and dissipate fragmenting molecules. The resulting fragments may be highlighted by mass spectrometry. Despite the amount of information obtained experimentally by the interpretation of the mass spectrum, experience alone cannot answer all the questions of the mechanism of fragmentation of DNA/RNA bases and a theoretical study is a complement to this information. A theoretical study allows us to know the weakest bonds in the molecule during ionization and thus may help to provide mechanisms of dissociation and produced fragments. The purpose of this work, using the DFT with the PBE functional, is to study the ionization and fragmentation mechanisms of DNA/RNA bases (Uracil, Cytosine, Adenine and Guanine) and to identify the cations corresponding to each peak in mass spectra. For all RNA bases, the retro Diels-Alder reaction (elimination of HNCO or NCO*) is a major route for dissociating, with the exception of adenine for which there is no atom oxygen in its structure. Loss of NH 3 (NH 2 *) molecule is another common way to all bases that contain amine group. The possibility of the loss of hydrogen from the cations is also investigated, as well as the dissociation of dehydrogenated cations and protonated uracil. This work shows the interest of providing DFT calculation in the interpretation of mass spectra of DNA bases. (author)

  19. LINE retrotransposon RNA is an essential structural and functional epigenetic component of a core neocentromeric chromatin.

    Directory of Open Access Journals (Sweden)

    Anderly C Chueh

    2009-01-01

    Full Text Available We have previously identified and characterized the phenomenon of ectopic human centromeres, known as neocentromeres. Human neocentromeres form epigenetically at euchromatic chromosomal sites and are structurally and functionally similar to normal human centromeres. Recent studies have indicated that neocentromere formation provides a major mechanism for centromere repositioning, karyotype evolution, and speciation. Using a marker chromosome mardel(10 containing a neocentromere formed at the normal chromosomal 10q25 region, we have previously mapped a 330-kb CENP-A-binding domain and described an increased prevalence of L1 retrotransposons in the underlying DNA sequences of the CENP-A-binding clusters. Here, we investigated the potential role of the L1 retrotransposons in the regulation of neocentromere activity. Determination of the transcriptional activity of a panel of full-length L1s (FL-L1s across a 6-Mb region spanning the 10q25 neocentromere chromatin identified one of the FL-L1 retrotransposons, designated FL-L1b and residing centrally within the CENP-A-binding clusters, to be transcriptionally active. We demonstrated the direct incorporation of the FL-L1b RNA transcripts into the CENP-A-associated chromatin. RNAi-mediated knockdown of the FL-L1b RNA transcripts led to a reduction in CENP-A binding and an impaired mitotic function of the 10q25 neocentromere. These results indicate that LINE retrotransposon RNA is a previously undescribed essential structural and functional component of the neocentromeric chromatin and that retrotransposable elements may serve as a critical epigenetic determinant in the chromatin remodelling events leading to neocentromere formation.

  20. Identification and functional analysis of novel phosphorylation sites in the RNA surveillance protein Upf1.

    Science.gov (United States)

    Lasalde, Clarivel; Rivera, Andrea V; León, Alfredo J; González-Feliciano, José A; Estrella, Luis A; Rodríguez-Cruz, Eva N; Correa, María E; Cajigas, Iván J; Bracho, Dina P; Vega, Irving E; Wilkinson, Miles F; González, Carlos I

    2014-02-01

    One third of inherited genetic diseases are caused by mRNAs harboring premature termination codons as a result of nonsense mutations. These aberrant mRNAs are degraded by the Nonsense-Mediated mRNA Decay (NMD) pathway. A central component of the NMD pathway is Upf1, an RNA-dependent ATPase and helicase. Upf1 is a known phosphorylated protein, but only portions of this large protein have been examined for phosphorylation sites and the functional relevance of its phosphorylation has not been elucidated in Saccharomyces cerevisiae. Using tandem mass spectrometry analyses, we report the identification of 11 putative phosphorylated sites in S. cerevisiae Upf1. Five of these phosphorylated residues are located within the ATPase and helicase domains and are conserved in higher eukaryotes, suggesting a biological significance for their phosphorylation. Indeed, functional analysis demonstrated that a small carboxy-terminal motif harboring at least three phosphorylated amino acids is important for three Upf1 functions: ATPase activity, NMD activity and the ability to promote translation termination efficiency. We provide evidence that two tyrosines within this phospho-motif (Y-738 and Y-742) act redundantly to promote ATP hydrolysis, NMD efficiency and translation termination fidelity.

  1. Regulation of functional KCNQ1OT1 lncRNA by β-catenin

    OpenAIRE

    Naohiro Sunamura; Takahito Ohira; Miki Kataoka; Daigo Inaoka; Hideyuki Tanabe; Yuji Nakayama; Mitsuo Oshimura; Hiroyuki Kugoh

    2016-01-01

    Long noncoding RNAs (lncRNAs) have been implicated in many biological processes through epigenetic mechanisms. We previously reported that KCNQ1OT1, an imprinted antisense lncRNA in the human KCNQ1 locus on chromosome 11p15.5, is involved in cis-limited silencing within an imprinted KCNQ1 cluster. Furthermore, aberration of KCNQ1OT1 transcription was observed with a high frequency in colorectal cancers. However, the molecular mechanism of the transcriptional regulation and the functional role...

  2. Regulation of functional KCNQ1OT1 lncRNA by β-catenin.

    OpenAIRE

    SUNAMURA, Naohiro; OHIRA, Takahito; KATAOKA, Miki; INAOKA, Daigo; TANABE, Hideyuki; NAKAYAMA, Yuji; OSHIMURA, Mitsuo; KUGOH, Hiroyuki

    2016-01-01

    Long noncoding RNAs (lncRNAs) have been implicated in many biological processes through epigenetic mechanisms. We previously reported that KCNQ1OT1, an imprinted antisense lncRNA in the human KCNQ1 locus on chromosome 11p15.5, is involved in cis-limited silencing within an imprinted KCNQ1 cluster. Furthermore, aberration of KCNQ1OT1 transcription was observed with a high frequency in colorectal cancers. However, the molecular mechanism of the transcriptional regulation and the functional role...

  3. UPF201 archaeal specific family members reveal structural similarity to RNA-binding proteins but low likelihood for RNA-binding function.

    Directory of Open Access Journals (Sweden)

    Krishnamurthy N Rao

    Full Text Available We have determined X-ray crystal structures of four members of an archaeal specific family of proteins of unknown function (UPF0201; Pfam classification: DUF54 to advance our understanding of the genetic repertoire of archaea. Despite low pairwise amino acid sequence identities (10-40% and the absence of conserved sequence motifs, the three-dimensional structures of these proteins are remarkably similar to one another. Their common polypeptide chain fold, encompassing a five-stranded antiparallel beta-sheet and five alpha-helices, proved to be quite unexpectedly similar to that of the RRM-type RNA-binding domain of the ribosomal L5 protein, which is responsible for binding the 5S- rRNA. Structure-based sequence alignments enabled construction of a phylogenetic tree relating UPF0201 family members to L5 ribosomal proteins and other structurally similar RNA binding proteins, thereby expanding our understanding of the evolutionary purview of the RRM superfamily. Analyses of the surfaces of these newly determined UPF0201 structures suggest that they probably do not function as RNA binding proteins, and that this domain specific family of proteins has acquired a novel function in archaebacteria, which awaits experimental elucidation.

  4. Hemorrhagic iliopsoas bursitis complicating well-functioning ceramic-on-ceramic total hip arthroplasty.

    Science.gov (United States)

    Park, Kyung Soon; Diwanji, Sanket R; Kim, Hyung Keun; Song, Eun Kyoo; Yoon, Taek Rim

    2009-08-01

    Iliopsoas bursitis has been increasingly recognized as a complication of total hip arthroplasty and is usually associated with polyethylene wear. Here, the authors report a case of hemorrhagic iliopsoas bursitis complicating an otherwise well-functioning ceramic-on-ceramic arthroplasty performed by minimal invasive modified 2-incision technique. The bursitis in turn resulted in femoral nerve palsy and femoral vein compression. In this report, there was no evidence to support that the bursitis was due to an inflammatory response to ceramic wear particles or any other wear particles originating from the total hip arthroplasty.

  5. Deep small RNA sequencing from the nematode Ascaris reveals conservation, functional diversification, and novel developmental profiles.

    Science.gov (United States)

    Wang, Jianbin; Czech, Benjamin; Crunk, Amanda; Wallace, Adam; Mitreva, Makedonka; Hannon, Gregory J; Davis, Richard E

    2011-09-01

    Eukaryotic cells express several classes of small RNAs that regulate gene expression and ensure genome maintenance. Endogenous siRNAs (endo-siRNAs) and Piwi-interacting RNAs (piRNAs) mainly control gene and transposon expression in the germline, while microRNAs (miRNAs) generally function in post-transcriptional gene silencing in both somatic and germline cells. To provide an evolutionary and developmental perspective on small RNA pathways in nematodes, we identified and characterized known and novel small RNA classes through gametogenesis and embryo development in the parasitic nematode Ascaris suum and compared them with known small RNAs of Caenorhabditis elegans. piRNAs, Piwi-clade Argonautes, and other proteins associated with the piRNA pathway have been lost in Ascaris. miRNAs are synthesized immediately after fertilization in utero, before pronuclear fusion, and before the first cleavage of the zygote. This is the earliest expression of small RNAs ever described at a developmental stage long thought to be transcriptionally quiescent. A comparison of the two classes of Ascaris endo-siRNAs, 22G-RNAs and 26G-RNAs, to those in C. elegans, suggests great diversification and plasticity in the use of small RNA pathways during spermatogenesis in different nematodes. Our data reveal conserved characteristics of nematode small RNAs as well as features unique to Ascaris that illustrate significant flexibility in the use of small RNAs pathways, some of which are likely an adaptation to Ascaris' life cycle and parasitism. The transcriptome assembly has been submitted to NCBI Transcriptome Shotgun Assembly Sequence Database(http://www.ncbi.nlm.nih.gov/genbank/TSA.html) under accession numbers JI163767–JI182837 and JI210738–JI257410.

  6. Application of two direct C(sp3)-H functionalizations for total synthesis of (+)-lactacystin.

    Science.gov (United States)

    Yoshioka, Shun; Nagatomo, Masanori; Inoue, Masayuki

    2015-01-02

    Herein, we report a new synthetic route from (S)-pyroglutaminol to (+)-lactacystin, a potent inhibitor of the 20S proteasome. The photoinduced intermolecular C(sp(3))-H alkynylation and intramolecular C(sp(3))-H acylation chemo- and stereoselectively constructed the tetra- and trisubstituted carbon centers, respectively. The obtained bicycle was transformed into the target compound in a concise manner. The present total synthesis demonstrates the power of the direct C(sp(3))-H functionalizations for the assembly of multiple functionalized structures of natural products.

  7. Developmental and functional expression of miRNA-stability related genes in the nervous system.

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    Érica de Sousa

    Full Text Available In the nervous system, control of gene expression by microRNAs (miRNAs has been investigated in fundamental processes, such as development and adaptation to ambient demands. The action of these short nucleotide sequences on specific genes depends on intracellular concentration, which in turn reflects the balance of biosynthesis and degradation. Whereas mechanisms underlying miRNA biogenesis has been investigated in recent studies, little is known about miRNA-stability related proteins. We first detected two genes in the retina that have been associated to miRNA stability, XRN2 and PAPD4. These genes are highly expressed during retinal development, however with distinct subcellular localization. We investigated whether these proteins are regulated during specific phases of the cell cycle. Combined analyses of nuclei position in neuroblastic layer and labeling using anti-cyclin D1 revealed that both proteins do not accumulate in S or M phases of the cell cycle, being poorly expressed in progenitor cells. Indeed, XRN2 and PAPD4 were observed mainly after neuronal differentiation, since low expression was also observed in astrocytes, endothelial and microglial cells. XRN2 and PAPD4 are expressed in a wide variety of neurons, including horizontal, amacrine and ganglion cells. To evaluate the functional role of both genes, we carried out experiments addressed to the retinal adaptation in response to different ambient light conditions. PAPD4 is upregulated after 3 and 24 hours of dark- adaptation, revealing that accumulation of this protein is governed by ambient light levels. Indeed, the fast and functional regulation of PAPD4 was not related to changes in gene expression, disclosing that control of protein levels occurs by post-transcriptional mechanisms. Furthermore, we were able to quantify changes in PAPD4 in specific amacrine cells after dark -adaptation, suggesting for circuitry-related roles in visual perception. In summary, in this study we

  8. IntNetLncSim: an integrative network analysis method to infer human lncRNA functional similarity.

    Science.gov (United States)

    Cheng, Liang; Shi, Hongbo; Wang, Zhenzhen; Hu, Yang; Yang, Haixiu; Zhou, Chen; Sun, Jie; Zhou, Meng

    2016-07-26

    Increasing evidence indicated that long non-coding RNAs (lncRNAs) were involved in various biological processes and complex diseases by communicating with mRNAs/miRNAs each other. Exploiting interactions between lncRNAs and mRNA/miRNAs to lncRNA functional similarity (LFS) is an effective method to explore function of lncRNAs and predict novel lncRNA-disease associations. In this article, we proposed an integrative framework, IntNetLncSim, to infer LFS by modeling the information flow in an integrated network that comprises both lncRNA-related transcriptional and post-transcriptional information. The performance of IntNetLncSim was evaluated by investigating the relationship of LFS with the similarity of lncRNA-related mRNA sets (LmRSets) and miRNA sets (LmiRSets). As a result, LFS by IntNetLncSim was significant positively correlated with the LmRSet (Pearson correlation γ2=0.8424) and LmiRSet (Pearson correlation γ2=0.2601). Particularly, the performance of IntNetLncSim is superior to several previous methods. In the case of applying the LFS to identify novel lncRNA-disease relationships, we achieved an area under the ROC curve (0.7300) in experimentally verified lncRNA-disease associations based on leave-one-out cross-validation. Furthermore, highly-ranked lncRNA-disease associations confirmed by literature mining demonstrated the excellent performance of IntNetLncSim. Finally, a web-accessible system was provided for querying LFS and potential lncRNA-disease relationships: http://www.bio-bigdata.com/IntNetLncSim.

  9. Comparative Analyses of MicroRNA Microarrays during Cardiogenesis: Functional Perspectives

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    Diego Franco

    2013-04-01

    Full Text Available Cardiovascular development is a complex process in which several transcriptional pathways are operative, providing instructions to the developing cardiomyocytes, while coping with contraction and morphogenetic movements to shape the mature heart. The discovery of microRNAs has added a new layer of complexity to the molecular mechanisms governing the formation of the heart. Discrete genetic ablation of the microRNAs processing enzymes, such as Dicer and Drosha, has highlighted the functional roles of microRNAs during heart development. Importantly, selective deletion of a single microRNA, miR-1-2, results in an embryonic lethal phenotype in which both morphogenetic, as well as impaired conduction, phenotypes can be observed. In an effort to grasp the variability of microRNA expression during cardiac morphogenesis, we recently reported the dynamic expression profile during ventricular development, highlighting the importance of miR-27 on the regulation of a key cardiac transcription factor, Mef2c. In this review, we compare the microRNA expression profile in distinct models of cardiogenesis, such as ventricular chamber development, induced pluripotent stem cell (iPS-derived cardiomyocytes and the aging heart. Importantly, out of 486 microRNAs assessed in the developing heart, 11% (55 displayed increased expression, many of which are also differentially expressed in distinct cardiogenetic experimental models, including iPS-derived cardiomyocytes. A review on the functional analyses of these differentially expressed microRNAs will be provided in the context of cardiac development, highlighting the resolution and power of microarrays analyses on the quest to decipher the most relevant microRNAs in the developing, aging and diseased heart.

  10. Comparative Analyses of MicroRNA Microarrays during Cardiogenesis: Functional Perspectives.

    Science.gov (United States)

    Bonet, Fernando; Hernandez-Torres, Francisco; Esteban, Franciso J; Aranega, Amelia; Franco, Diego

    2013-04-03

    Cardiovascular development is a complex process in which several transcriptional pathways are operative, providing instructions to the developing cardiomyocytes, while coping with contraction and morphogenetic movements to shape the mature heart. The discovery of microRNAs has added a new layer of complexity to the molecular mechanisms governing the formation of the heart. Discrete genetic ablation of the microRNAs processing enzymes, such as Dicer and Drosha, has highlighted the functional roles of microRNAs during heart development. Importantly, selective deletion of a single microRNA, miR-1-2, results in an embryonic lethal phenotype in which both morphogenetic, as well as impaired conduction, phenotypes can be observed. In an effort to grasp the variability of microRNA expression during cardiac morphogenesis, we recently reported the dynamic expression profile during ventricular development, highlighting the importance of miR-27 on the regulation of a key cardiac transcription factor, Mef2c. In this review, we compare the microRNA expression profile in distinct models of cardiogenesis, such as ventricular chamber development, induced pluripotent stem cell (iPS)-derived cardiomyocytes and the aging heart. Importantly, out of 486 microRNAs assessed in the developing heart, 11% (55) displayed increased expression, many of which are also differentially expressed in distinct cardiogenetic experimental models, including iPS-derived cardiomyocytes. A review on the functional analyses of these differentially expressed microRNAs will be provided in the context of cardiac development, highlighting the resolution and power of microarrays analyses on the quest to decipher the most relevant microRNAs in the developing, aging and diseased heart.

  11. A loss of function analysis of host factors influencing Vaccinia virus replication by RNA interference.

    Directory of Open Access Journals (Sweden)

    Philippa M Beard

    Full Text Available Vaccinia virus (VACV is a large, cytoplasmic, double-stranded DNA virus that requires complex interactions with host proteins in order to replicate. To explore these interactions a functional high throughput small interfering RNA (siRNA screen targeting 6719 druggable cellular genes was undertaken to identify host factors (HF influencing the replication and spread of an eGFP-tagged VACV. The experimental design incorporated a low multiplicity of infection, thereby enhancing detection of cellular proteins involved in cell-to-cell spread of VACV. The screen revealed 153 pro- and 149 anti-viral HFs that strongly influenced VACV replication. These HFs were investigated further by comparisons with transcriptional profiling data sets and HFs identified in RNAi screens of other viruses. In addition, functional and pathway analysis of the entire screen was carried out to highlight cellular mechanisms involved in VACV replication. This revealed, as anticipated, that many pro-viral HFs are involved in translation of mRNA and, unexpectedly, suggested that a range of proteins involved in cellular transcriptional processes and several DNA repair pathways possess anti-viral activity. Multiple components of the AMPK complex were found to act as pro-viral HFs, while several septins, a group of highly conserved GTP binding proteins with a role in sequestering intracellular bacteria, were identified as strong anti-viral VACV HFs. This screen has identified novel and previously unexplored roles for cellular factors in poxvirus replication. This advancement in our understanding of the VACV life cycle provides a reliable knowledge base for the improvement of poxvirus-based vaccine vectors and development of anti-viral theraputics.

  12. Measurements of long-range interactions between protein-functionalized surfaces by total internal reflection microscopy.

    Science.gov (United States)

    Wang, Zhaohui; Gong, Xiangjun; Ngai, To

    2015-03-17

    Understanding the interaction between protein-functionalized surfaces is an important subject in a variety of protein-related processes, ranging from coatings for biomedical implants to targeted drug carriers and biosensors. In this work, utilizing a total internal reflection microscope (TIRM), we have directly measured the interactions between micron-sized particles decorated with three types of common proteins concanavalin A (ConA), bovine serum albumin (BSA), lysozyme (LYZ), and glass surface coated with soy proteins (SP). Our results show that the protein adsorption greatly affects the charge property of the surfaces, and the interactions between those protein-functionalized surfaces depend on solution pH values. At pH 7.5-10.0, all these three protein-functionalized particles are highly negatively charged, and they move freely above the negatively charged SP-functionalized surface. The net interaction between protein-functionalized surfaces captured by TIRM was found as a long-range, nonspecific double-layer repulsion. When pH was decreased to 5.0, both protein-functionalized surfaces became neutral and double-layer repulsion was greatly reduced, resulting in adhesion of all three protein-functionalized particles to the SP-functionalized surface due to the hydrophobic attraction. The situation is very different at pH = 4.0: BSA-decorated particles, which are highly charged, can move freely above the SP-functionalized surfaces, while ConA- and LYZ-decorated particles can only move restrictively in a limited range. Our results quantify these nonspecific kT-scale interactions between protein-functionalized surfaces, which will enable the design of surfaces for use in biomedical applications and study of biomolecular interactions.

  13. Late effects on gonadal function of cyclophosphamide, total-body irradiation, and marrow transplantation

    Energy Technology Data Exchange (ETDEWEB)

    Sanders, J.E.; Buckner, C.D.; Leonard, J.M.; Sullivan, K.M.; Witherspoon, R.P.; Deeg, H.J.; Storb, R.; Thomas, E.D.

    1983-09-01

    One hundred thirty-seven patients had gonadal function evaluated 1-11 years after marrow transplantation. All 15 women less than age 26 and three of nine older than age 26 who were treated with 200 mg/kg cyclophosphamide recovered normal gonadotropin levels and menstruation. Five have had five pregnancies resulting in three live births, one spontaneous abortion, and one elective abortion. Three of 38 women who were prepared with 120 mg/kg cyclophosphamide and 920-1200 rad total-body irradiation had normal gonadotropin levels and menstruation. Two had pregnancies resulting in one spontaneous and one elective abortion. Of 31 men prepared with 200 mg/kg cyclophosphamide, 30 had normal luteinizing hormone levels, 20 had normal follicle-stimulating hormone levels, and 10 of 15 had spermatogenesis. Four have fathered five normal children. Thirty-six of 41 men prepared with 120 mg/kg cyclophosphamide and 920-1750 rad total-body irradiation had normal luteinizing hormone levels, ten had normal follicle-stimulating hormone levels, and 2 of 32 studied had spermatogenesis. One has fathered two normal children. It was concluded that cyclophosphamide does not prevent return of normal gonadal function in younger women and in most men. Total-body irradiation prevents return of normal gonadal function in the majority of patients.

  14. Isolation, expression and functional analysis of a putative RNA-dependent RNA polymerase gene from maize (Zea mays L.).

    Science.gov (United States)

    He, Junguang; Dong, Zhigang; Jia, Zhiwei; Wang, Jianhua; Wang, Guoying

    2010-02-01

    RNA-dependent RNA polymerases (RdRPs) in plants have been reported to be involved in post-transcriptional gene silencing (PTGS) and antiviral defense. In this report, an RdRP gene from maize (ZmRdRP1) was obtained by rapid amplification of cDNA ends (RACE) and RT-PCR. The mRNA of ZmRdRP1 was composed of 3785 nucleotides, including a 167 nt 5' untranslated region (UTR), a 291 nt 3'UTR and a 3327 nt open reading frame (ORF), which encodes a putative protein of 1108 amino acids with an estimated molecular mass of 126.9 kDa and a predicated isoelectric point (pI) of 8.37. Real-time quantitative RT-PCR analysis showed that ZmRdRP1 was elicited by salicylic acid (SA) treatment, methyl jasmonate (MeJA) treatment and sugarcane mosaic virus (SCMV) infection. We silenced ZmRdRP1 by constitutively expressing an inverted-repeat fragment of ZmRdRP1 (ir-RdRP1) in transgenic maize plants. Further studies revealed that the ir-RdRP1 transgenic plants were more susceptible to SCMV infection than wild type plants. Virus-infected transgenic maize plants developed more serious disease symptoms and accumulated more virus than wild type plants. These findings suggested that ZmRdRP1 was involved in antiviral defense in maize.

  15. Reprogramming transcription by distinct classes of enhancers functionally defined by eRNA.

    Science.gov (United States)

    Wang, Dong; Garcia-Bassets, Ivan; Benner, Chris; Li, Wenbo; Su, Xue; Zhou, Yiming; Qiu, Jinsong; Liu, Wen; Kaikkonen, Minna U; Ohgi, Kenneth A; Glass, Christopher K; Rosenfeld, Michael G; Fu, Xiang-Dong

    2011-05-15

    Mammalian genomes are populated with thousands of transcriptional enhancers that orchestrate cell-type-specific gene expression programs, but how those enhancers are exploited to institute alternative, signal-dependent transcriptional responses remains poorly understood. Here we present evidence that cell-lineage-specific factors, such as FoxA1, can simultaneously facilitate and restrict key regulated transcription factors, exemplified by the androgen receptor (AR), to act on structurally and functionally distinct classes of enhancer. Consequently, FoxA1 downregulation, an unfavourable prognostic sign in certain advanced prostate tumours, triggers dramatic reprogramming of the hormonal response by causing a massive switch in AR binding to a distinct cohort of pre-established enhancers. These enhancers are functional, as evidenced by the production of enhancer-templated non-coding RNA (eRNA) based on global nuclear run-on sequencing (GRO-seq) analysis, with a unique class apparently requiring no nucleosome remodelling to induce specific enhancer-promoter looping and gene activation. GRO-seq data also suggest that liganded AR induces both transcription initiation and elongation. Together, these findings reveal a large repository of active enhancers that can be dynamically tuned to elicit alternative gene expression programs, which may underlie many sequential gene expression events in development, cell differentiation and disease progression.

  16. microRNA-184 functions as tumor suppressor in renal cell carcinoma.

    Science.gov (United States)

    Su, Zhengming; Chen, Duqun; Li, Yifan; Zhang, Enpu; Yu, Zuhu; Chen, Ting; Jiang, Zhimao; Ni, Liangchao; Yang, Shangqi; Gui, Yaoting; Ye, Jiongxian; Lai, Yongqing

    2015-03-01

    microRNAs (miRNAs) are evolutionarily conserved, endogenous, small, noncoding RNA molecules of approximately 22 nucleotides in length that function as post-transcriptional gene regulators. Their aberrant expression may be involved in human diseases, including cancer. Although miRNA-184 (miR-184) has been reported in other tumors, its function in renal cell carcinoma (RCC) is still unknown. The aim of the present study was to investigate the role of miR-184 in RCC. The impacts of miR-184 on cell migration, proliferation and apoptosis were evaluated using migration scratch, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and flow cytometry assay. Our studies revealed that miR-184 mimic significantly inhibits cell migration, suppresses cell proliferation and induces renal cancer cell apoptosis in vitro when compared with the negative control (P184 played a significant role as a tumor suppressor in RCC. Therefore, miR-184 may be a promising therapeutic target for renal cancer treatment in the future.

  17. Noncoding RNA Shows Context-Dependent Function | Center for Cancer Research

    Science.gov (United States)

    In addition to well-studied protein coding sequences, it is known that the genomes of higher organisms produce numerous noncoding RNAs (ncRNAs). Important roles for some ncRNAs in cell function have been demonstrated, though usually on a case-by-case basis, leading some scientists to argue that the majority of ncRNA production is just “noise” that results from the imperfect transcription machinery. The fact that many ncRNAs overlap with coding genes has hampered studies of their activities. Thus, a general understanding of whether ncRNA production is functional or not is lacking. To address this issue, Daniel Larson, Ph.D., of CCR’s Laboratory of Receptor Biology and Gene Expression, and his colleagues developed a new approach using single-molecule imaging in living cells. The researchers specifically labeled coding and ncRNAs from the GAL locus in yeast, which regulates the galactose response. Glucose is the preferred source of carbon for yeast, but when it is scarce, genes within the GAL locus, including GAL10 and GAL1, are activated to allow the metabolism of galactose.

  18. Transcriptional Repressor NIR Functions in the Ribosome RNA Processing of Both 40S and 60S Subunits

    Science.gov (United States)

    Wang, Yingshuang; Kong, Ruirui; Hu, Lelin; Schuele, Roland; Du, Xiaojuan; Ke, Yang

    2012-01-01

    Background NIR was identified as an inhibitor of histone acetyltransferase and it represses transcriptional activation of p53. NIR is predominantly localized in the nucleolus and known as Noc2p, which is involved in the maturation of the 60S ribosomal subunit. However, how NIR functions in the nucleolus remains undetermined. In the nucleolus, a 47S ribosomal RNA precursor (pre-rRNA) is transcribed and processed to produce 18S, 5.8S and 28S rRNAs. The 18S rRNA is incorporated into the 40S ribosomal subunit, whereas the 28S and 5.8S rRNAs are incorporated into the 60S subunit. U3 small nucleolar RNA (snoRNA) directs 18S rRNA processing and U8 snoRNA mediates processing of 28S and 5.8 S rRNAs. Functional disruption of nucleolus often causes p53 activation to inhibit cell proliferation. Methodology/Principal Findings Western blotting showed that NIR is ubiquitously expressed in different human cell lines. Knock-down of NIR by siRNA led to inhibition of the 18S, 28S and 5.8S rRNAs evaluated by pulse-chase experiment. Pre-rRNA particles (pre-rRNPs) were fractionated from the nucleus by sucrose gradient centrifugation and analysis of the pre-RNPs components showed that NIR existed in the pre-RNPs of both the 60S and 40S subunits and co-fractionated with 32S and 12S pre-rRNAs in the 60S pre-rRNP. Protein-RNA binding experiments demonstrated that NIR is associated with the 32S pre-rRNA and U8 snoRNA. In addition, NIR bound U3 snoRNA. It is a novel finding that depletion of NIR did not affect p53 protein level but de-repressed acetylation of p53 and activated p21. Conclusions We provide the first evidence for a transcriptional repressor to function in the rRNA biogenesis of both the 40S and 60S subunits. Our findings also suggested that a nucleolar protein may alternatively signal to p53 by affecting the p53 modification rather than affecting p53 protein level. PMID:22363708

  19. Personality, function and satisfaction in patients undergoing total hip or knee replacement.

    Science.gov (United States)

    Ramaesh, Rishikesan; Jenkins, Paul; Lane, Judith V; Knight, Sara; Macdonald, Deborah; Howie, Colin

    2014-03-01

    The aim of this study was to investigate the relationships between personality and joint-specific function, general physical and general mental health in patients undergoing total hip (THA) and knee arthroplasty (TKA). One hundred and eighty-four patients undergoing THA and 205 undergoing TKA were assessed using the Eysneck Personality Questionnaire, brief version (EPQ-BV). General physical and mental health was measured using the Short-Form 12 (SF-12) questionnaire and the EuroQol (EQ-5D). Joint-specific function was measured using the Oxford hip or knee score. The "unstable introvert" personality type was associated with poorer pre-operative function and health in patients with hip arthrosis. In patients with knee arthrosis, there was poorer general health in those with "stable extrovert" and "unstable introvert" types. Personality was not an independent predictor of outcome following TKA or THA. The main predictor was pre-operative function and health. Comorbidity was an important covariate of both pre-operative and postoperative function. Personality may play a role in the interaction of these disease processes with function and health perception. It may also affect the response and interpretation of psychometric and patient-reported outcome measures. It may be important to characterise and identify these traits in potential arthroplasty patients as it may help deliver targeted education and management to improve outcomes in certain groups.

  20. Small RNA and Transcriptome Sequencing Reveal a Potential miRNA-Mediated Interaction Network That Functions during Somatic Embryogenesis in Lilium pumilum DC. Fisch.

    Science.gov (United States)

    Zhang, Jing; Xue, Bingyang; Gai, Meizhu; Song, Shengli; Jia, Nana; Sun, Hongmei

    2017-01-01

    Plant somatic embryos are widely used in the fields of germplasm conservation, breeding for genetic engineering and artificial seed production. MicroRNAs (miRNAs) play pivotal roles in somatic embryogenesis (SE) regulation. However, their regulatory roles during various stages of SE remain unclear. In this study, six types of embryogenic samples of Lilium pumilum DC. Fisch., including organogenic callus, embryogenic callus induced for 4 weeks, embryogenic callus induced for 6 weeks, globular embryos, torpedo embryos and cotyledon embryos, were prepared for small RNA sequencing. The results revealed a total of 2,378,760 small RNA reads, among which the most common size was 24 nt. Four hundred and fifty-two known miRNAs, belonging to more than 86 families, 57 novel miRNAs and 40 miRNA*s were identified. The 86 known miRNA families were sorted according to an alignment with their homologs across 24 land plants into the following four categories: 23 highly conserved, 4 moderately conserved, 15 less conserved and 44 species-specific miRNAs. Differentially expressed known miRNAs were identified during various stages of SE. Subsequently, the expression levels of 12 differentially expressed miRNAs and 4 targets were validated using qRT-PCR. In addition, six samples were mixed in equal amounts for transcript sequencing, and the sequencing data were used as transcripts for miRNA target prediction. A total of 66,422 unigenes with an average length of 800 bp were assembled from 56,258,974 raw reads. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment indicated that 38,004 and 15,497 unigenes were successfully assigned to GO terms and KEGG pathways, respectively. Among the unigenes, 2,182 transcripts were predicted to be targets for 396 known miRNAs. The potential targets of the identified miRNAs were mostly classified into the following GO terms: cell, binding and metabolic process. Enriched KEGG analysis demonstrated that carbohydrate metabolism

  1. Functional exercise after total hip replacement (FEATHER a randomised control trial

    Directory of Open Access Journals (Sweden)

    Monaghan Brenda

    2012-11-01

    Full Text Available Abstract Background Prolonged physical impairments in range of movement, postural stability and walking speed are commonly reported following total hip replacement (THR. It is unclear from the current body of evidence what kind of exercises should be performed to maximize patient function and quality of life. Methods/design This will be a single blind multi centre randomized control trial with two arms. Seventy subjects post primary total hip arthroplasty will be randomized into either an experimental group (n=35, or to a control group (n=35. The experimental group will attend a functional exercise class twice weekly for a six week period from week 12 to week 18 post surgery. The functional exercise group will follow a circuit based functional exercise class supervised by a chartered Physiotherapist. The control group will receive usual care. The principal investigator (BM will perform blinded outcome assessments on all patients using validated measures for pain, stiffness, and function using the Western Ontario and Mc Master Universities Osteoarthritis index (WOMAC. This is the primary outcome measurement tool. Secondary outcome measurements include Quality of life (SF-36, 6 min walk test, Visual Analogue Scale, and the Berg Balance score. The WOMAC score will be collated on day five post surgery and repeated at week twelve and week eighteen. All other measurements will be taken at week 12 and repeated at week eighteen. In addition a blinded radiologist will measure gluteus medius cross sectional area using real time ultrasound for all subjects at week 12 and at week 18 to determine if the functional exercise programme has any effect on muscle size. Discussion This randomised controlled trial will add to the body of evidence on the relationship between muscle size, functional ability, balance, quality of life and time post surgery in patients following total hip arthroplasty. The CONSORT guidelines will be followed to throughout. Ethical

  2. An ecological function and services approach to total maximum daily load (TMDL) prioritization.

    Science.gov (United States)

    Hall, Robert K; Guiliano, David; Swanson, Sherman; Philbin, Michael J; Lin, John; Aron, Joan L; Schafer, Robin J; Heggem, Daniel T

    2014-04-01

    Prioritizing total maximum daily load (TMDL) development starts by considering the scope and severity of water pollution and risks to public health and aquatic life. Methodology using quantitative assessments of in-stream water quality is appropriate and effective for point source (PS) dominated discharge, but less so in watersheds with mostly nonpoint source (NPS) related impairments. For NPSs, prioritization in TMDL development and implementation of associated best management practices should focus on restoration of ecosystem physical functions, including how restoration effectiveness depends on design, maintenance and placement within the watershed. To refine the approach to TMDL development, regulators and stakeholders must first ask if the watershed, or ecosystem, is at risk of losing riparian or other ecologically based physical attributes and processes. If so, the next step is an assessment of the spatial arrangement of functionality with a focus on the at-risk areas that could be lost, or could, with some help, regain functions. Evaluating stream and wetland riparian function has advantages over the traditional means of water quality and biological assessments for NPS TMDL development. Understanding how an ecosystem functions enables stakeholders and regulators to determine the severity of problem(s), identify source(s) of impairment, and predict and avoid a decline in water quality. The Upper Reese River, Nevada, provides an example of water quality impairment caused by NPS pollution. In this river basin, stream and wetland riparian proper functioning condition (PFC) protocol, water quality data, and remote sensing imagery were used to identify sediment sources, transport, distribution, and its impact on water quality and aquatic resources. This study found that assessments of ecological function could be used to generate leading (early) indicators of water quality degradation for targeting pollution control measures, while traditional in-stream water

  3. Total ozone retrieval from GOME UV spectral data using the weighting function DOAS approach

    Directory of Open Access Journals (Sweden)

    M. Coldewey-Egbers

    2005-01-01

    Full Text Available A new algorithm approach called Weighting Function Differential Optical Absorption Spectroscopy (WFDOAS is presented which has been developed to retrieve total ozone columns from nadir observations of the Global Ozone Monitoring Experiment. By fitting the vertically integrated ozone weighting function rather than ozone cross-section to the sun-normalized radiances, a direct retrieval of vertical column amounts is possible. The new WFDOAS approach takes into account the slant path wavelength modulation that is usually neglected in the standard DOAS approach using single airmass factors. This paper focuses on the algorithm description and error analysis, while in a companion paper by Weber et al. (2004 a detailed validation with groundbased measurements is presented. For the first time several auxiliary quantities directly derived from the GOME spectral range such as cloud-top-height and cloud fraction (O2-A band and effective albedo using the Lambertian Equivalent Reflectivity (LER near 377nm are used in combination as input to the ozone retrieval. In addition the varying ozone dependent contribution to the Raman correction in scattered light known as Ring effect has been included. The molecular ozone filling-in that is accounted for in the new algorithm has the largest contribution to the improved total ozone results from WFDOAS compared to the operational product. The precision of the total ozone retrieval is estimated to be better than 3% for solar zenith angles below 80°.

  4. Extracellular vesicle-mediated transfer of processed and functional RNY5 RNA

    Science.gov (United States)

    Chakrabortty, Sudipto K.; Prakash, Ashwin; Nechooshtan, Gal; Hearn, Stephen; Gingeras, Thomas R.

    2015-01-01

    Extracellular vesicles (EVs) have been proposed as a means to promote intercellular communication. We show that when human primary cells are exposed to cancer cell EVs, rapid cell death of the primary cells is observed, while cancer cells treated with primary or cancer cell EVs do not display this response. The active agents that trigger cell death are 29- to 31-nucleotide (nt) or 22- to 23-nt processed fragments of an 83-nt primary transcript of the human RNY5 gene that are highly likely to be formed within the EVs. Primary cells treated with either cancer cell EVs, deproteinized total RNA from either primary or cancer cell EVs, or synthetic versions of 31- and 23-nt fragments trigger rapid cell death in a dose-dependent manner. The transfer of processed RNY5 fragments through EVs may reflect a novel strategy used by cancer cells toward the establishment of a favorable microenvironment for their proliferation and invasion. PMID:26392588

  5. Psychological factors as risk factors for poor hip function after total hip arthroplasty

    Directory of Open Access Journals (Sweden)

    Benditz A

    2017-02-01

    Full Text Available Achim Benditz,1 Petra Jansen,2 Jan Schaible,1 Christina Roll,1 Joachim Grifka,1 Jürgen Götz1 1Department of Orthopedics, University Medical Center Regensburg, Asklepios Klinikum Bad Abbach, Bad Abbach, 2Department of Sport Science, University of Regensburg, Regensburg, Germany Abstract: Recovery after total hip arthroplasty (THA is influenced by several psychological aspects, such as depression, anxiety, resilience, and personality traits. We hypothesized that preoperative depression impedes early functional outcome after THA (primary outcome measure. Additional objectives were perioperative changes in the psychological status and their influence on perioperative outcome. This observational study analyzed depression, anxiety, resilience, and personality traits in 50 patients after primary unilateral THA. Hip functionality was measured by means of the Harris Hip Score. Depression, state anxiety, and resilience were evaluated preoperatively as well as 1 and 5 weeks postoperatively. Trait anxiety and personality traits were measured once preoperatively. Patients with low depression and anxiety levels had significantly better outcomes with respect to early hip functionality. Resilience and personality traits did not relate to hip functionality. Depression and state anxiety levels significantly decreased within the 5-week stay in the acute and rehabilitation clinic, whereas resilience remained at the same level. Our study suggests that low depression and anxiety levels are positively related to early functionality after THA. Therefore, perioperative measurements of these factors seem to be useful to provide the best support for patients with risk factors. Keywords: total hip arthroplasty, psychological factors, depression, state anxiety, trait anxiety, resilience, personality traits

  6. The power of cross-functional teams in driving total quality

    Science.gov (United States)

    Mcclung, Tim M.; Mcmaster, Tom J.

    1992-01-01

    Garrett Canada, a Division of Allied-Signal Aerospace Canada, has been a member of the Canadian aerospace industry for 40 years. Although Garrett Canada has always been a profitable division with a solid market share, the changing and turbulent business environment and globalization of the aerospace industry has created new demands and challenges. The marketplace is demanding faster introduction of new products, as well as shorter leadtimes for repairs and spares. It was recognized that reducing cycle times for new products and for ongoing production would not only satisfy our customers, it would also enhance our business performance through reduced inventories, lower past due, and more responsiveness to change. It was evident that drastic function changes were required if we were to maintain our position as a premier aerospace supplier. The challenge was to convert a stable, somewhat slow-paced work environment with strong functional boundaries into a boundaryless world class team functioning in a total quality environment and focused on customer satisfaction. Complete and uncompromised customer satisfaction has become our driving force, with Total Quality being our engine to continuously improve our processes and increase our speed. The way in which this transition has been brought about is the subject of this presentation.

  7. The power of cross-functional teams in driving total quality

    Science.gov (United States)

    McClung, Tim M.; McMaster, Tom J.

    Garrett Canada, a Division of Allied-Signal Aerospace Canada, has been a member of the Canadian aerospace industry for 40 years. Although Garrett Canada has always been a profitable division with a solid market share, the changing and turbulent business environment and globalization of the aerospace industry has created new demands and challenges. The marketplace is demanding faster introduction of new products, as well as shorter leadtimes for repairs and spares. It was recognized that reducing cycle times for new products and for ongoing production would not only satisfy our customers, it would also enhance our business performance through reduced inventories, lower past due, and more responsiveness to change. It was evident that drastic function changes were required if we were to maintain our position as a premier aerospace supplier. The challenge was to convert a stable, somewhat slow-paced work environment with strong functional boundaries into a boundaryless world class team functioning in a total quality environment and focused on customer satisfaction. Complete and uncompromised customer satisfaction has become our driving force, with Total Quality being our engine to continuously improve our processes and increase our speed. The way in which this transition has been brought about is the subject of this presentation.

  8. Multiple RNA processing defects and impaired chloroplast function in plants deficient in the organellar protein-only RNase P enzyme.

    Directory of Open Access Journals (Sweden)

    Wenbin Zhou

    Full Text Available Transfer RNA (tRNA precursors undergo endoribonucleolytic processing of their 5' and 3' ends. 5' cleavage of the precursor transcript is performed by ribonuclease P (RNase P. While in most organisms RNase P is a ribonucleoprotein that harbors a catalytically active RNA component, human mitochondria and the chloroplasts (plastids and mitochondria of seed plants possess protein-only RNase P enzymes (PRORPs. The plant organellar PRORP (PRORP1 has been characterized to some extent in vitro and by transient gene silencing, but the molecular, phenotypic and physiological consequences of its down-regulation in stable transgenic plants have not been assessed. Here we have addressed the function of the dually targeted organellar PRORP enzyme in vivo by generating stably transformed Arabidopsis plants in which expression of the PRORP1 gene was suppressed by RNA interference (RNAi. PRORP1 knock-down lines show defects in photosynthesis, while mitochondrial respiration is not appreciably affected. In both plastids and mitochondria, the effects of PRORP1 knock-down on the processing of individual tRNA species are highly variable. The drastic reduction in the levels of mature plastid tRNA-Phe(GAA and tRNA-Arg(ACG suggests that these two tRNA species limit plastid gene expression in the PRORP1 mutants and, hence, are causally responsible for the mutant phenotype.

  9. Time to parathyroid function recovery in patients with protracted hypoparathyroidism after total thyroidectomy.

    Science.gov (United States)

    Villarroya-Marquina, Inés; Sancho, Juan; Lorente-Poch, Leyre; Gallego-Otaegui, Lander; Sitges-Serra, Antonio

    2018-01-01

    Hypocalcaemia is the most common adverse effect after total thyroidectomy. It recovers in about two-thirds of the patients within the first postoperative month. Little is known, however, about recovery of the parathyroid function (RPF) after this time period. The aim of the present study was to investigate the time to RPF in patients with protracted (>1 month) hypoparathyroidism after total thyroidectomy. Cohort prospective observational study. Adult patients undergoing total thyroidectomy for goitre or thyroid cancer. Cases with protracted hypoparathyroidism were studied for RPF during the following months. Time to RPF and variables associated with RPF or permanent hypoparathyroidism were recorded. Out of 854 patients undergoing total thyroidectomy, 142 developed protracted hypoparathyroidism. Of these, 36 (4.2% of the entire cohort) developed permanent hypoparathyroidism and 106 recovered: 73 before 6 months, 21 within 6-12 months and 12 after 1 year follow-up. Variables significantly associated with RPF were the number of parathyroid glands remaining in situ (not autografted nor inadvertently resected) and a serum calcium concentration >2.25 mmol/L at one postoperative month. Late RPF (>6 months) was associated with surgery for thyroid cancer. RPF was still possible after one year in patients with four parathyroid glands preserved in situ and serum calcium concentration at one month >2.25 mmol/L. Permanent hypoparathyroidism should not be diagnosed in patients requiring replacement therapy for more than six months, especially if the four parathyroid glands were preserved. © 2018 European Society of Endocrinology.

  10. The function of the inner nuclear envelope protein SUN1 in mRNA export is regulated by phosphorylation.

    Science.gov (United States)

    Li, Ping; Stumpf, Maria; Müller, Rolf; Eichinger, Ludwig; Glöckner, Gernot; Noegel, Angelika A

    2017-08-22

    SUN1, a component of the LINC (Linker of Nucleoskeleton and Cytoskeleton) complex, functions in mammalian mRNA export through the NXF1-dependent pathway. It associates with mRNP complexes by direct interaction with NXF1. It also binds to the NPC through association with the nuclear pore component Nup153, which is involved in mRNA export. The SUN1-NXF1 association is at least partly regulated by a protein kinase C (PKC) which phosphorylates serine 113 (S113) in the N-terminal domain leading to reduced interaction. The phosphorylation appears to be important for the SUN1 function in nuclear mRNA export since GFP-SUN1 carrying a S113A mutation was less efficient in restoring mRNA export after SUN1 knockdown as compared to the wild type protein. By contrast, GFP-SUN1-S113D resembling the phosphorylated state allowed very efficient export of poly(A)+RNA. Furthermore, probing a possible role of the LINC complex component Nesprin-2 in this process we observed impaired mRNA export in Nesprin-2 knockdown cells. This effect might be independent of SUN1 as expression of a GFP tagged SUN-domain deficient SUN1, which no longer can interact with Nesprin-2, did not affect mRNA export.

  11. Empirical models of Total Electron Content based on functional fitting over Taiwan during geomagnetic quiet condition

    Directory of Open Access Journals (Sweden)

    Y. Kakinami

    2009-08-01

    Full Text Available Empirical models of Total Electron Content (TEC based on functional fitting over Taiwan (120° E, 24° N have been constructed using data of the Global Positioning System (GPS from 1998 to 2007 during geomagnetically quiet condition (Dst>−30 nT. The models provide TEC as functions of local time (LT, day of year (DOY and the solar activity (F, which are represented by 1–162 days mean of F10.7 and EUV. Other models based on median values have been also constructed and compared with the models based on the functional fitting. Under same values of F parameter, the models based on the functional fitting show better accuracy than those based on the median values in all cases. The functional fitting model using daily EUV is the most accurate with 9.2 TECu of root mean square error (RMS than the 15-days running median with 10.4 TECu RMS and the model of International Reference Ionosphere 2007 (IRI2007 with 14.7 TECu RMS. IRI2007 overestimates TEC when the solar activity is low, and underestimates TEC when the solar activity is high. Though average of 81 days centered running mean of F10.7 and daily F10.7 is often used as indicator of EUV, our result suggests that average of F10.7 mean from 1 to 54 day prior and current day is better than the average of 81 days centered running mean for reproduction of TEC. This paper is for the first time comparing the median based model with the functional fitting model. Results indicate the functional fitting model yielding a better performance than the median based one. Meanwhile we find that the EUV radiation is essential to derive an optimal TEC.

  12. Altered resting-state amygdala functional connectivity after 36 hours of total sleep deprivation.

    Directory of Open Access Journals (Sweden)

    Yongcong Shao

    Full Text Available Recent neuroimaging studies have identified a potentially critical role of the amygdala in disrupted emotion neurocircuitry in individuals after total sleep deprivation (TSD. However, connectivity between the amygdala and cerebral cortex due to TSD remains to be elucidated. In this study, we used resting-state functional MRI (fMRI to investigate the functional connectivity changes of the basolateral amygdala (BLA and centromedial amygdala (CMA in the brain after 36 h of TSD.Fourteen healthy adult men aged 25.9 ± 2.3 years (range, 18-28 years were enrolled in a within-subject crossover study. Using the BLA and CMA as separate seed regions, we examined resting-state functional connectivity with fMRI during rested wakefulness (RW and after 36 h of TSD.TSD resulted in a significant decrease in the functional connectivity between the BLA and several executive control regions (left dorsolateral prefrontal cortex [DLPFC], right dorsal anterior cingulate cortex [ACC], right inferior frontal gyrus [IFG]. Increased functional connectivity was found between the BLA and areas including the left posterior cingulate cortex/precuneus (PCC/PrCu and right parahippocampal gyrus. With regard to CMA, increased functional connectivity was observed with the rostral anterior cingulate cortex (rACC and right precentral gyrus.These findings demonstrate that disturbance in amygdala related circuits may contribute to TSD psychophysiology and suggest that functional connectivity studies of the amygdala during the resting state may be used to discern aberrant patterns of coupling within these circuits after TSD.

  13. Development of DNA affinity techniques for the functional characterization of purified RNA polymerase II transcription factors

    International Nuclear Information System (INIS)

    Garfinkel, S.; Thompson, J.A.; Cohen, R.B.; Brendler, T.; Safer, B.

    1987-01-01

    Affinity adsorption, precipitation, and partitioning techniques have been developed to purify and characterize RNA Pol II transcription components from whole cell extracts (WCE) (HeLa) and nuclear extracts (K562). The titration of these extracts with multicopy constructs of the Ad2 MLP but not pUC8, inhibits transcriptional activity. DNA-binding factors precipitated by this technique are greatly enriched by centrifugation. Using this approach, factors binding to the upstream promoter sequence (UPS) of the Ad2 MLP have been rapidly isolated by Mono Q, Mono S, and DNA affinity chromatography. By U.V. crosslinking to nucleotides containing specific 32 P-phosphodiester bonds within the recognition sequence, this factor is identified as a M/sub r/ = 45,000 polypeptide. To generate an assay system for the functional evaluation of single transcription components, a similar approach using synthetic oligonucleotide sequences spanning single promoter binding sites has been developed. The addition of a synthetic 63-mer containing the UPS element of the Ad2 MLP to HeLa WCE inhibited transcription by 60%. The addition of partially purified UPS binding protein, but not RNA Pol II, restored transcriptional activity. The addition of synthetic oligonucleotides containing other regulatory sequences not present in the Ad2 MLP was without effect

  14. Leg position influences early blood loss and functional recovery following total knee arthroplasty: A randomized study.

    Science.gov (United States)

    Yang, Yang; Yong-Ming, Lv; Pei-jian, Ding; Jia, Li; Ying-ze, Zhang

    2015-11-01

    Hidden blood loss is a major factor influencing functional recovery and quality of life in patients undergoing total knee arthroplasty. Special hip and knee flexion positions after have been reported to have promising results with respect to reducing perioperative blood loss. The purpose of this study was to determine the effect of postoperative leg position on blood loss and functional recovery after total knee arthroplasty. We enrolled 46 consecutive patients with degenerative osteoarthritis of the knee in this prospective, randomized study. The patients were randomly allocated to a flexion or an extension group. In the flexion group, the affected leg was elevated by 60° at the hip, and the knee was flexed by 60°, while in the extension group, the affected knee was fully extended postoperatively. Blood loss, hemoglobin level, knee circumference and range of motion (ROM) were recorded to determine the influence of postoperative leg position on clinical outcomes. Although the transfusion rate was similar between the two groups (P > 0.05), other parameters related to blood loss (including calculated blood loss, hidden blood loss and postoperative knee circumference) were significantly lower in the flexion group than in the extension group (P group had gained a better ROM in the affected knee than had patients from the extension group (P = 0.04). At 6 months, however, the ROM of the affected knee was similar in both groups. The hospital stay was 1.9 days shorter in the flexion group than in the extension group. Wound infection rates were similar in both groups, and no proven case of deep vein thrombosis was observed in either group. Elevation of the hip by 60° with 60° knee flexion is an effective and simple method to reduce blood loss after primary unilateral total knee arthroplasty, and contributes to better recovery of the functional ROM in the early postoperative period. Copyright © 2015 IJS Publishing Group Limited. Published by Elsevier Ltd. All rights

  15. The hot pepper (Capsicum annuum microRNA transcriptome reveals novel and conserved targets: a foundation for understanding MicroRNA functional roles in hot pepper.

    Directory of Open Access Journals (Sweden)

    Dong-Gyu Hwang

    Full Text Available MicroRNAs (miRNAs are a class of non-coding RNAs approximately 21 nt in length which play important roles in regulating gene expression in plants. Although many miRNA studies have focused on a few model plants, miRNAs and their target genes remain largely unknown in hot pepper (Capsicum annuum, one of the most important crops cultivated worldwide. Here, we employed high-throughput sequencing technology to identify miRNAs in pepper extensively from 10 different libraries, including leaf, stem, root, flower, and six developmental stage fruits. Based on a bioinformatics pipeline, we successfully identified 29 and 35 families of conserved and novel miRNAs, respectively. Northern blot analysis was used to validate further the expression of representative miRNAs and to analyze their tissue-specific or developmental stage-specific expression patterns. Moreover, we computationally predicted miRNA targets, many of which were experimentally confirmed using 5' rapid amplification of cDNA ends analysis. One of the validated novel targets of miR-396 was a domain rearranged methyltransferase, the major de novo methylation enzyme, involved in RNA-directed DNA methylation in plants. This work provides the first reliable draft of the pepper miRNA transcriptome. It offers an expanded picture of pepper miRNAs in relation to other plants, providing a basis for understanding the functional roles of miRNAs in pepper.

  16. Right ventricular function late after total repair of tetralogy of Fallot

    Energy Technology Data Exchange (ETDEWEB)

    Straten, Alexander van; Roos, Albert de [Leiden University Medical Center, Department of Radiology, Leiden (Netherlands); Vliegen, Hubert W. [Leiden University Medical Center, Department of Cardiology, Leiden (Netherlands); Hazekamp, Mark G. [Leiden University Medical Center, Department of Cardiothoracic Surgery, Leiden (Netherlands)

    2005-04-01

    Over the past decades, life expectancy in patients with congenital heart disease has increased dramatically. However, serious complications may develop late after total repair in infancy. These complications are usually the result of longstanding pulmonary regurgitation, which leads to dilatation of the right ventricle and an increased risk for severe arrhythmias. Therefore lifelong follow-up in these patients is required. Cardiac magnetic resonance imaging is the current imaging tool of choice because it offers superior imaging quality and enables accurate quantification of functional parameters such as flow volumes and systolic and diastolic performance. (orig.)

  17. Right ventricular function late after total repair of tetralogy of Fallot

    International Nuclear Information System (INIS)

    Straten, Alexander van; Roos, Albert de; Vliegen, Hubert W.; Hazekamp, Mark G.

    2005-01-01

    Over the past decades, life expectancy in patients with congenital heart disease has increased dramatically. However, serious complications may develop late after total repair in infancy. These complications are usually the result of longstanding pulmonary regurgitation, which leads to dilatation of the right ventricle and an increased risk for severe arrhythmias. Therefore lifelong follow-up in these patients is required. Cardiac magnetic resonance imaging is the current imaging tool of choice because it offers superior imaging quality and enables accurate quantification of functional parameters such as flow volumes and systolic and diastolic performance. (orig.)

  18. Predictors of pain and physical function at 3 and 12 months after total hip arthroplasty

    DEFF Research Database (Denmark)

    Plews, Sarah; Løvlund Nielsen, Randi; Overgaard, Søren

    with primary hip osteoarthritis responded to Hip dysfunction and Osteoarthritis Outcome Score (HOOS) questionnaires prior to and 3 and 12 months after THA. Preoperative pain intensity; joint space width (JSW), age, gender, and body mass index (BMI) were used to predict changes in pain and physical function....... Conclusions: Preoperative pain predicted changes in pain and physical function up to one year after THA. Such knowledge should be taken into consideration, when assessing OA patients prior to surgery. This study provides useful insight for clinicians, regarding the overall improvement patients can expect......Background: Few studies have combined preoperative patient-reported and objective outcome measures to predict outcomes after total hip arthroplasty (THA). Purpose / Aim of Study: to identify predictors of outcome 3 and 12 months after THA Materials and Methods: A cohort of 107 consecutive patients...

  19. Control of a Biped Robot by Total Rate of Angular Momentum Using the Task Function Approach

    Directory of Open Access Journals (Sweden)

    J. A. Rojas-Estrada

    2005-01-01

    Full Text Available In this work we address the control problem of biped robots by using the task function approach. A problem arrives when one of the feet is in contact with the ground, which presents imperfections. There is then the possibility that the biped robot undergoes a fall. It is difficult to track any trajectory due to the presence of unevenness on the ground. What we propose is to use the task function approach combined with the application of the total rate of angular momentum to obtain a control law for the ankle. By this technique, the tracking becomes more smooth and the balance is assured. The control law proposed allows the upper part of the robot to be controlled independently since only the ankle actuators are concerned. We enounce the formal problem and present some simulations with real parameters of a 21 degrees of freedom biped robot.

  20. A Simple and Effective Method for High Quality Co-Extraction of Genomic DNA and Total RNA from Low Biomass Ectocarpus siliculosus, the Model Brown Alga

    Science.gov (United States)

    Greco, Maria; Sáez, Claudio A.; Brown, Murray T.; Bitonti, Maria Beatrice

    2014-01-01

    The brown seaweed Ectocarpus siliculosus is an emerging model species distributed worldwide in temperate coastal ecosystems. Over 1500 strains of E. siliculosus are available in culture from a broad range of geographic locations and ecological niches. To elucidate the molecular mechanisms underlying its capacity to cope with different environmental and biotic stressors, genomic and transcriptomic studies are necessary; this requires the co-isolation of genomic DNA and total RNA. In brown algae, extraction of nucleic acids is hindered by high concentrations of secondary metabolites that co-precipitate with nucleic acids. Here, we propose a reliable, rapid and cost-effective procedure for the co-isolation of high-quality nucleic acids using small quantities of biomass (25-, 50- and 100 mg) from strains of E. siliculosus (RHO12; LIA4A; EC524 and REP10–11) isolated from sites with different environmental conditions. The procedure employs a high pH extraction buffer (pH 9.5) which contains 100 mM Tris-HCl and 150 mM NaCl, with the addition of 5 mM DTT and 1% sarkosyl to ensure maximum solubility of nucleic acids, effective inhibition of nuclease activity and removal of interfering contaminants (e.g. polysaccharides, polyphenols). The use of sodium acetate together with isopropanol shortened precipitation time and enhanced the yields of DNA/RNA. A phenol:chlorophorm:isoamyl alcohol step was subsequently used to purify the nucleic acids. The present protocol produces high yields of nucleic acids from only 25 mg of fresh algal biomass (0.195 and 0.284 µg mg−1 fresh weigh of RNA and DNA, respectively) and the high quality of the extracted nucleic acids was confirmed through spectrophotometric and electrophoretic analyses. The isolated RNA can be used directly in downstream applications such as RT-PCR and the genomic DNA was suitable for PCR, producing reliable restriction enzyme digestion patterns. Co-isolation of DNA/RNA from different strains indicates that this

  1. Changes in gait symmetry, gait velocity and self-reported function following total hip replacement.

    Science.gov (United States)

    Hodt-Billington, Caroline; Helbostad, Jorunn L; Vervaat, Willemijn; Rognsvåg, Turid; Moe-Nilssen, Rolf

    2011-09-01

    To investigate the magnitude of change at different time points in measures of gait symmetry, gait velocity and self-reported function following total hip replacement. Longitudinal with test occasions pre-surgery and 3, 6 and 12 months post-surgery. Thirty-four patients with hip osteoarthritis (mean age 63 years, standard deviation 11 years). Subjects walked back and forth along a 7-m walkway at slow, preferred and fast speed. Anteroposterior, vertical and mediolateral trunk symmetry was assessed by accelerometry, while single support symmetry, step-length symmetry and gait velocity was simultaneously assessed by an electronic walkway. Self-reported function was assessed by Hip disability and Osteoarthritis Outcome Score. Gait symmetry data were normalized for gait velocity. Changes between test occasions were reported as effect size. All measures showed effect sizes > 0.30 from pre-operative to 12-months postoperative assessments, and improvements were significant (p symmetry. In general, gait symmetry and gait velocity improved most 6 and 12 months postoperatively, while self-reported function improved most 3 months postoperatively. Early improvements were seen in self-reported function, suggesting immediate relief from stiffness and pain, while gait symmetry and velocity improved later postoperatively, suggesting that gait quality and performance require prolonged rehabilitation with postoperative guidance, muscular strengthening and motor relearning.

  2. Component alignment and functional outcome following computer assisted and jig based total knee arthroplasty.

    Science.gov (United States)

    Lad, Dnyanesh G; Thilak, Jai; Thadi, Mohan

    2013-01-01

    Incorrect positioning of the implant and improper alignment of the limb following total knee arthroplasty (TKA) can lead to rapid implant wear, loosening, and suboptimal function. Studies suggest that alignment errors of > 3° are associated with rapid failure and less satisfactory function. Computer navigated systems have been developed to enhance precision in instrumentation during surgery. The aim of the study was to compare component alignment following computer assisted surgery (CAS) and jig based TKA as well as functional outcome. This is a prospective study of 100 knees to compare computer-assisted TKA and jig-based surgery in relation to femoral and tibial component alignment and functional outcome. The postoperative x-rays (anteroposterior and lateral) of the knee and CT scanogram from hip to foot were obtained. The coronal alignment of the femoral and tibial components and rotational alignment of femoral component was calculated. Knee society score at 24 months was used to assess the function. Results of our study show that mean placement of the tibial component in coronal plane (91.3037°) and sagittal planes (3.6058°) was significantly better with CAS. The difference was statistically insignificant in case of mean coronal alignment of the femoral components (90.34210° in navigation group and 90.5444° in jig group) and in case of the mean femoral condylar twist angle (external rotation 2.3406° in navigation group versus 2.3593° in jig group). A significantly improved placement of the component was found in the coronal and sagittal planes of the tibial component by CAS. The placement of the components in the other planes was comparable with the values recorded in the jig-based surgery group. Functional outcome was not significantly different.

  3. The Fanconi anemia proteins functionally interact with the protein kinase regulated by RNA (PKR).

    Science.gov (United States)

    Zhang, Xiaoling; Li, June; Sejas, Daniel P; Rathbun, Keaney R; Bagby, Grover C; Pang, Qishen

    2004-10-15

    Protein kinase regulated by RNA (PKR) plays critical roles in cell growth and apoptosis and is implicated as a potential pathogenic factor of Alzheimer's, Parkinson's, and Huntington's diseases. Here we report that this proapoptotic kinase is also involved in Fanconi anemia (FA), a disease characterized by bone marrow (BM) failure and leukemia. We have used a BM extract to show that three FA proteins, FANCA, FANCC, and FANCG, functionally interact with the PKR kinase, which in turn regulates translational control. By using a combined immunoprecipitation and reconstituted kinase assay, in which an active PKR kinase complex was captured from a normal cell extract, we demonstrated functional interactions between the FA proteins and the PKR kinase. In primary human BM cells, mutations in the FANCA, FANCC, and FANCG genes markedly increase the amount of PKR bound to FANCC, and this PKR accumulation is correlated with elevated PKR activation and hypersensitivity of BM progenitor cells to growth repression mediated by the inhibitory cytokines interferon-gamma and tumor necrosis factor-alpha. Specific inhibition of PKR by 2-aminopurine in these FA BM cells attenuates PKR activation and apoptosis induction. In lymphoblasts derived from an FA-C patient, overexpression of a dominant negative mutant PKR (PKRK296R) suppressed PKR activation and apoptosis induced by interferon-gamma and tumor necrosis factor-alpha. Furthermore, by using genetically matched wild-type and PKR-null cells, we demonstrated that forced expression of a patient-derived FA-C mutant (FANCCL554P) augmented double-stranded RNA-induced PKR activation and cell death. Thus, inappropriate activation of PKR as a consequence of certain FA mutations might play a role in bone marrow failure that frequently occurred in FA.

  4. Tranexamic acid increases early perioperative functional outcomes after total knee arthroplasty

    Directory of Open Access Journals (Sweden)

    Matthew J. Grosso, MD

    2018-03-01

    Full Text Available Background: The purpose of this study was to investigate the influence of tranexamic acid (TXA on functional outcomes in the immediate postoperative period after total knee arthroplasty (TKA. We hypothesized that the known benefits of TXA would confer measurable clinical improvements in physical therapy (PT performance, decrease pain, and decrease hospital length of stay (LOS. Methods: We retrospectively analyzed 560 TKA patients, including 280 consecutive patients whose surgery was performed before the initiation of a standardized TXA protocol and the first 280 patients who received TXA after protocol initiation. Outcome measurements included postoperative changes in hemoglobin and hematocrit, LOS, pain scores, destination of discharge, and steps ambulated with PT over 5 sessions. Results: TXA administration resulted in less overall drops in hemoglobin (P < .001 and hematocrit levels (P < .001. Moreover, patients administered TXA ambulated more than their counterparts during every PT session, which was statistically significant during the second (P = .010, third (P = .011, and fourth (P = .024 sessions. On average, the TXA cohort ambulated 20% more per PT session than patients who did not receive TXA (P < .001. TXA administration did not influence pain levels during PT, hospital LOS, or discharge destination in this investigation. Conclusions: It is well known that TXA reduces postoperative anemia, but this study also demonstrates that it confers early perioperative functional benefits for TKA patients. Potential mechanisms for this benefit include reduced rates of postoperative anemia and reduced rates of hemarthroses. Keywords: Tranexamic acid, Total knee arthroplasty, Blood loss, Physical therapy

  5. Circ-ZNF609 Is a Circular RNA that Can Be Translated and Functions in Myogenesis.

    Science.gov (United States)

    Legnini, Ivano; Di Timoteo, Gaia; Rossi, Francesca; Morlando, Mariangela; Briganti, Francesca; Sthandier, Olga; Fatica, Alessandro; Santini, Tiziana; Andronache, Adrian; Wade, Mark; Laneve, Pietro; Rajewsky, Nikolaus; Bozzoni, Irene

    2017-04-06

    Circular RNAs (circRNAs) constitute a family of transcripts with unique structures and still largely unknown functions. Their biogenesis, which proceeds via a back-splicing reaction, is fairly well characterized, whereas their role in the modulation of physiologically relevant processes is still unclear. Here we performed expression profiling of circRNAs during in vitro differentiation of murine and human myoblasts, and we identified conserved species regulated in myogenesis and altered in Duchenne muscular dystrophy. A high-content functional genomic screen allowed the study of their functional role in muscle differentiation. One of them, circ-ZNF609, resulted in specifically controlling myoblast proliferation. Circ-ZNF609 contains an open reading frame spanning from the start codon, in common with the linear transcript, and terminating at an in-frame STOP codon, created upon circularization. Circ-ZNF609 is associated with heavy polysomes, and it is translated into a protein in a splicing-dependent and cap-independent manner, providing an example of a protein-coding circRNA in eukaryotes. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  6. Dopamine transporter polymorphism modulates oculomotor function and DAT1 mRNA expression in schizophrenia.

    Science.gov (United States)

    Wonodi, Ikwunga; Hong, L Elliot; Stine, O Colin; Mitchell, Braxton D; Elliott, Amie; Roberts, Rosalinda C; Conley, Robert R; McMahon, Robert P; Thaker, Gunvant K

    2009-03-05

    Smooth pursuit eye movement (SPEM) deficit is an established schizophrenia endophenotype with a similar neurocognitive construct to working memory. Frontal eye field (FEF) neurons controlling SPEM maintain firing when visual sensory information is removed, and their firing rates directly correlate with SPEM velocity. We previously demonstrated a paradoxical association between a functional polymorphism of dopamine signaling (COMT gene) and SPEM. Recent evidence implicates the dopamine transporter gene (DAT1) in modulating cortical dopamine and associated neurocognitive functions. We hypothesized that DAT1 10/10 genotype, which reduces dopamine transporter expression and increases extracellular dopamine, would affect SPEM. We examined the effects of DAT1 genotype on: Clinical diagnosis in the study sample (n = 418; 190 with schizophrenia), SPEM measures in a subgroup with completed oculomotor measures (n = 200; 87 schizophrenia), and DAT1 gene expression in FEF tissue obtained from postmortem brain samples (n = 32; 16 schizophrenia). DAT1 genotype was not associated with schizophrenia. DAT1 10/10 genotype was associated with better SPEM in healthy controls, intermediate SPEM in unaffected first-degree relatives of schizophrenia subjects, and worse SPEM in schizophrenia subjects. In the gene expression study, DAT1 10/10 genotype was associated with significantly reduced DAT1 mRNA transcript in FEF tissue from healthy control donors (P < 0.05), but higher expression in schizophrenia donors. Findings suggest regulatory effects of another gene(s) or etiological factor in schizophrenia, which modulate DAT1 gene function. 2008 Wiley-Liss, Inc.

  7. Playing RNase P evolution: swapping the RNA catalyst for a protein reveals functional uniformity of highly divergent enzyme forms.

    Directory of Open Access Journals (Sweden)

    Christoph Weber

    2014-08-01

    Full Text Available The RNase P family is a diverse group of endonucleases responsible for the removal of 5' extensions from tRNA precursors. The diversity of enzyme forms finds its extremes in the eukaryal nucleus where RNA-based catalysis by complex ribonucleoproteins in some organisms contrasts with single-polypeptide enzymes in others. Such structural contrast suggests associated functional differences, and the complexity of the ribonucleoprotein was indeed proposed to broaden the enzyme's functionality beyond tRNA processing. To explore functional overlap and differences between most divergent forms of RNase P, we replaced the nuclear RNase P of Saccharomyces cerevisiae, a 10-subunit ribonucleoprotein, with Arabidopsis thaliana PRORP3, a single monomeric protein. Surprisingly, the RNase P-swapped yeast strains were viable, displayed essentially unimpaired growth under a wide variety of conditions, and, in a certain genetic background, their fitness even slightly exceeded that of the wild type. The molecular analysis of the RNase P-swapped strains showed a minor disturbance in tRNA metabolism, but did not point to any RNase P substrates or functions beyond that. Altogether, these results indicate the full functional exchangeability of the highly dissimilar enzymes. Our study thereby establishes the RNase P family, with its combination of structural diversity and functional uniformity, as an extreme case of convergent evolution. It moreover suggests that the apparently gratuitous complexity of some RNase P forms is the result of constructive neutral evolution rather than reflecting increased functional versatility.

  8. The modified total body rotation test: a rapid, reliable assessment of physical function in older adults.

    Science.gov (United States)

    Stanziano, Damian C; Signorile, Joseph F; Mow, Sara; Davidson, Erin E; Ouslander, Joseph G; Roos, Bernard A

    2010-10-01

    To describe a quick, modified measure of total body rotation with performance rankings for elderly persons and to highlight the importance of total body rotational ability for overall physical performance. Scores were collected during the same testing session and were compared with other measures using a Pearson correlation. Community-based senior centers. One hundred forty-five participants aged 79.6±7.2 were recruited using flyers distributed at health fairs, senior centers, and lifelong learning programs. The modified total body rotation test (MTBR), the back-scratch test, the modified sit-and-reach test, the 30-second chair-stand test, the Short Physical Performance Battery, and the short version of the Continuous-Scale Physical Functional Performance Test. The MBTR significantly correlated to all standard measures of physical performance. Norms are reported for all participants, and reliability analyses were conducted using data from 18 participants. Data were divided into quintiles to create cutoffs for physical performance rankings that can be used when reporting results to the general public. Many evaluations measure isolated components of physical performance, but few examine whole body movements that allow all components necessary for independence and function to work in harmony as needed considering the task and the individual's movement strategy. The current study offers the MTBR as a quick, easy, and cost-effective evaluation method to quantify the degree of impairment or injury and the rate of improvement with treatment or training. © 2010, Copyright the Authors. Journal compilation © 2010, The American Geriatrics Society.

  9. Effects of total glucosides of peony on AQP-5 and its mRNA expression in submandibular glands of NOD mice with Sjogren's syndrome.

    Science.gov (United States)

    Wu, G-L; Pu, X-H; Yu, G-Y; Li, T-Y

    2015-01-01

    The aim of this study was to observe the effects of total glucosides of peony (TGP) on pathological change, Aquaporin-5 (AQP-5) and its mRNA expression in submandibular glands of non-obese diabetic (NOD) mice with Sjogren's Syndrome, to investigate its regulation on secretion of salivary glands. 40 NOD mice were randomly divided into model group, TGP group, hydroxychloroquine group, combination group (n = 10). For TGP group, the mice were intragastrically administrated with 0.4 ml TGP dilution per day in accordance with 300 g/kg dose; for hydroxychloroquine group, the mice were intragastrically administrated with 0.4 ml hydroxychloroquine per day in accordance with 60 mg/kg dose; for the combination group, the mice were intragastrically administrated with 0.4 ml TGP dilution and 0.4 ml hydroxychloroquine. 8 weeks later, the mice were sacrificed, and submandibular glands were collected by anatomy. Pathological changes of submandibular gland were observed under a light microscope; AQP-5 protein in submandibular glands was detected by immunohistochemical staining; and AQP-5 mRNA expression in submandibular glands was detected by RT-PCR. The lymphocytic infiltration score of model mice was significantly higher than that of other groups. The pathological morphology and score of NOD mice were significantly improved after administration, and the combination group was superior to the hydroxychloroquine group and TGP group (p TGP group and the combination group were higher than the hydroxychloroquine group (p TGP may improve pathological damage of submandibular glands of NOD mouse with Sjogren's syndrome by upregulating AQP-5 and its mRNA expression in submandibular glands.

  10. The natural history of unexplained early poor function following total hip replacement.

    Science.gov (United States)

    Watson, Bruce S; Jenkins, Paul J; Ballantyne, James A

    2014-01-01

    The majority of patients experience a significant improvement in quality of life and function after total hip replacement (THR). It has recently been shown that age and good pre-operative function are the best predictors of postoperative function. When patients fail to achieve a satisfactory outcome, a cause is often identified. Where there is no identifiable cause, advice, follow-up and management is not clear. The aim of this study was to determine the long-term outcome of patients who had early poor function, but no identifiable cause. From a regional database, we identified 1,564 patients who underwent unilateral THR between 1998 and 2004 and who were without complication or subsequent bilateral procedure at six months. These patients were divided into two groups according to their Harris hip score (HHS) at this stage: group A consisted of 270 patients with a 'poor' result (HHS less than 70). Group B consisted of 1,294 patients with a 'good' or 'excellent' result (HHS 70 or above). The patients were reviewed at five years. One hundred and ten patients from group A and 980 from group B completed five-year follow-up without further identifiable complication. Those with poor or fair function at six months were at an increased risk of developing an identified complication by five years including dislocation (OR 5.7, 95% CI 1.8-18.2), deep infection (OR 9.8, 95%CI 2.9-37.7) and death (OR 1.6, 95% CI 1.1-2.3). There was a greater rate of revision in group A versus group B (OR 5.7, 95% CI 2.9-11). The overall function measured by the Harris hip score significantly improved in group A, but never reached that of those with good or excellent function at six months (HHS 76.2 versus 90.3, P < 0.001). Patients with poor function at six months, but no obvious cause, are at higher risk of developing complications by five years. This group may benefit from more regular arthroplasty review and intervention.

  11. Functional results of robotic total intersphincteric resection with hand-sewn coloanal anastomosis.

    Science.gov (United States)

    Luca, F; Valvo, M; Guerra-Cogorno, M; Simo, D; Blesa-Sierra, E; Biffi, R; Garberoglio, C

    2016-06-01

    In recent decades there has been an increasing trend toward sphincter-preserving procedures for the treatment of low rectal cancer. Robotic surgery is considered to be particularly beneficial when operating in the deep pelvis, where laparoscopy presents technical limitations. The aim of this study was to prospectively evaluate the functional outcomes in patients affected by rectal cancer after robotic total intersphincteric resection (ISR) with hand-sewn coloanal anastomosis. From March 2008 to October 2012, 23 consecutive patients affected by distal rectal adenocarcinoma underwent robotic ISR. Operative, clinical, pathological and functional data regarding continence or presence of a low anterior resection syndrome (LARS) were prospectively collected in a database. Twenty-three consecutive patients were included in the study: 8 men and 15 women. The mean age was 60.2 years (range 28-73). Eighteen (78.3%) had neoadjuvant radiochemotherapy. Conversion rate was nil. The mean operative time was 296.01 min and the mean postoperative hospital stay was 7.43 ± 1.73 days. According to Kirwan's incontinence score, good fecal continence was shown in 85.7% of patients (Grade 1 and 2) and none required a colostomy (Grade 4). Concerning LARS score, the results were as follows: 57.1% patients had no LARS; 19% minor LARS and 23.8% major LARS. Robotic total ISR for low rectal cancer is an acceptable alternative to traditional procedures. Extensive discussion with the patient about the risk of poor functional outcomes or LARS syndrome is mandatory when considering an ISR for treatment of low rectal cancer. Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. Total hip arthroplasty outcomes assessment using functional and radiographic scores to compare canine systems.

    Science.gov (United States)

    Iwata, D; Broun, H C; Black, A P; Preston, C A; Anderson, G I

    2008-01-01

    A retrospective multi-centre study was carried out in order to compare outcomes between cemented and uncemented total hip arthoplasties (THA). A quantitative orthopaedic outcome assessment scoring system was devised in order to relate functional outcome to a numerical score, to allow comparison between treatments and amongst centres. The system combined a radiographic score and a clinical score. Lower scores reflect better outcomes than higher scores. Consecutive cases of THA were included from two specialist practices between July 2002 and December 2005. The study included 46 THA patients (22 uncemented THA followed for 8.3 +/- 4.7M and 24 cemented THA for 26.0 +/- 15.7M) with a mean age of 4.4 +/- 3.3 years at surgery. Multi-variable linear and logistical regression analyses were performed with adjustments for age at surgery, surgeon, follow-up time, uni- versus bilateral disease, gender and body weight. The differences between treatment groups in terms of functional scores or total scores were not significant (p > 0.05). Radiographic scores were different between treatment groups. However, these scores were usually assessed within two months of surgery and proved unreliable predictors of functional outcome (p > 0.05). The findings reflect relatively short-term follow-up, especially for the uncemented group, and do not include clinician-derived measures, such as goniometry and thigh circumference. Longer-term follow-up for the radiographic assessments is essential. A prospective study including the clinician-derived outcomes needs to be performed in order to validate the outcome instrument in its modified form.

  13. Avian reovirus L2 genome segment sequences and predicted structure/function of the encoded RNA-dependent RNA polymerase protein

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    Xu Wanhong

    2008-12-01

    Full Text Available Abstract Background The orthoreoviruses are infectious agents that possess a genome comprised of 10 double-stranded RNA segments encased in two concentric protein capsids. Like virtually all RNA viruses, an RNA-dependent RNA polymerase (RdRp enzyme is required for viral propagation. RdRp sequences have been determined for the prototype mammalian orthoreoviruses and for several other closely-related reoviruses, including aquareoviruses, but have not yet been reported for any avian orthoreoviruses. Results We determined the L2 genome segment nucleotide sequences, which encode the RdRp proteins, of two different avian reoviruses, strains ARV138 and ARV176 in order to define conserved and variable regions within reovirus RdRp proteins and to better delineate structure/function of this important enzyme. The ARV138 L2 genome segment was 3829 base pairs long, whereas the ARV176 L2 segment was 3830 nucleotides long. Both segments were predicted to encode λB RdRp proteins 1259 amino acids in length. Alignments of these newly-determined ARV genome segments, and their corresponding proteins, were performed with all currently available homologous mammalian reovirus (MRV and aquareovirus (AqRV genome segment and protein sequences. There was ~55% amino acid identity between ARV λB and MRV λ3 proteins, making the RdRp protein the most highly conserved of currently known orthoreovirus proteins, and there was ~28% identity between ARV λB and homologous MRV and AqRV RdRp proteins. Predictive structure/function mapping of identical and conserved residues within the known MRV λ3 atomic structure indicated most identical amino acids and conservative substitutions were located near and within predicted catalytic domains and lining RdRp channels, whereas non-identical amino acids were generally located on the molecule's surfaces. Conclusion The ARV λB and MRV λ3 proteins showed the highest ARV:MRV identity values (~55% amongst all currently known ARV and MRV

  14. A simple and efficient protocol for isolation of high quality functional RNA from different tissues of turmeric (Curcuma longa L.).

    Science.gov (United States)

    Deepa, K; Sheeja, T E; Santhi, R; Sasikumar, B; Cyriac, Anu; Deepesh, P V; Prasath, D

    2014-04-01

    Many experiments in plant molecular biology require processing of a large number of RNA samples and in some cases large quantities are required for a single application. In turmeric, a major spice and medicinal plant, a protocol for RNA isolation is not available. The major difficulty encountered while using other popular protocols is the low yield and quality of RNA which hampers the downstream applications like qRT-PCR, cDNA synthesis and micro RNA isolation. Commercial kits though available are costly and were found to be unsuccessful in case of rhizomes and root tissues that are rich in polyphenols, polysaccharides and alkaloids. It was thus felt that a quick, handy and cheap protocol of total RNA isolation from different tissues of turmeric was required for day to day working in our lab. The new protocol utilizes SDS based extraction buffer including β-mercaptoethanol and PVP with sequential acid phenol:chloroform extraction to remove polyphenols and proteins, followed by the purification with sodium acetate to eliminate polysaccharides. The protocol is simple and can be completed in less than 3 h. The RNA yield from rhizome was higher by more than fivefold with both A260/280 and A260/230 ratio in the range of 1.8-2.0. The protocol worked well with leaf, rhizome, pseudostem and root tissues with RIN >7.0 and the isolated RNA could be successfully used for cDNA synthesis, RT-PCR, qRT-PCR and small RNA isolation including microRNA.

  15. Essential Structural and Functional Roles of the Cmr4 Subunit in RNA Cleavage by the Cmr CRISPR-Cas Complex

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    Nancy F. Ramia

    2014-12-01

    Full Text Available The Cmr complex is the multisubunit effector complex of the type III-B clustered regularly interspaced short palindromic repeats (CRISPR-Cas immune system. The Cmr complex recognizes a target RNA through base pairing with the integral CRISPR RNA (crRNA and cleaves the target at multiple regularly spaced locations within the complementary region. To understand the molecular basis of the function of this complex, we have assembled information from electron microscopic and X-ray crystallographic structural studies and mutagenesis of a complete Pyrococcus furiosus Cmr complex. Our findings reveal that four helically packed Cmr4 subunits, which make up the backbone of the Cmr complex, act as a platform to support crRNA binding and target RNA cleavage. Interestingly, we found a hook-like structural feature associated with Cmr4 that is likely the site of target RNA binding and cleavage. Our results also elucidate analogies in the mechanisms of crRNA and target molecule binding by the distinct Cmr type III-A and Cascade type I-E complexes.

  16. Origins of biological function in DNA and RNA hairpin loop motifs from replica exchange molecular dynamics simulation.

    Science.gov (United States)

    Swadling, Jacob B; Ishii, Kunihiko; Tahara, Tahei; Kitao, Akio

    2018-01-31

    Deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) have remarkably similar chemical structures, but despite this, they play significantly different roles in modern biology. In this article, we explore the possible conformations of DNA and RNA hairpins to better understand the fundamental differences in structure formation and stability. We use large parallel temperature replica exchange molecular dynamics ensembles to sample the full conformational landscape of these hairpin molecules so that we can identify the stable structures formed by the hairpin sequence. Our simulations show RNA adopts a narrower distribution of folded structures compared to DNA at room temperature, which forms both hairpins and many unfolded conformations. RNA is capable of forming twice as many hydrogen bonds than DNA which results in a higher melting temperature. We see that local chemical differences lead to emergent molecular properties such as increased persistence length in RNA that is weakly temperature dependant. These discoveries provide fundamental insight into how RNA forms complex folded tertiary structures which confer enzymatic-like function in ribozymes, whereas DNA retains structural motifs in order to facilitate function such as translation of sequence.

  17. An update on recent methods applied for deciphering the diversity of the noncoding RNA genome structure and function.

    Science.gov (United States)

    Spicuglia, Salvatore; Maqbool, Muhammad Ahmad; Puthier, Denis; Andrau, Jean-Christophe

    2013-09-01

    The explosion of high throughput sequencing technologies marked a turn in our way of understanding the complexity and diversity of the transcriptome, including noncoding transcription dependent on RNA polymerase II. Many new ncRNA populations were described in recent years, including for example TSS RNAs, lincRNAs, eRNAs, PROMPTS and several others. Besides the advances in the average depth coverage of RNA-seq experiments, various additional protocols are now available that can be used to address qualitative and quantitative aspects of the noncoding transcriptome complexity and function. In this review, we will focus on methods allowing isolation and characterization of complex RNA populations using sequencing based approaches, including conventional strategies already used for coding genome and more specific developments allowing, for example, the study of nascent strand transcription, protein-bound or structured RNAs. Crown Copyright © 2013. Published by Elsevier Inc. All rights reserved.

  18. STUDY OF FUNCTIONAL RESULTS OF CEMENTED TOTAL HIP REPLACEMENT BY MOORE’S APPROACH

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    Arunim

    2016-02-01

    Full Text Available INTRODUCTION The total hip replacement (THR has probably become the surgical procedure of choice for vide variety of hip joint disabling diseases. The prosthesis used for THR is often grouped into cemented, cement-less and hybrid ones. There has been increasing trends in use of cement-less components citing more number of complications namely loosening, increased infection rate etc. however with additional cost factors as well. We conducted this study to ascertain whether in a developing country like ours should we really switch over to un-cemented hip replacements dreading such complications or can we still use cemented prosthesis with equally good if not better results. METHODS A study of functional results of cemented total hip replacement was done in patients with varied age groups ranging from 40 years to 75 years with the average age being 54.8 years. 20 patients with 21 diseased hips were treated with cemented total hip replacement by Moore’s posterior approach at NSCB Subharti medical college, Meerut, UP from December 2010 to December 2013 and reviewed thereafter with an average follow-up period of 4.2 years. Average surgical time required was one and half hour. Patients were asked to come for follow up on 1st month, 3rd month and 6th month and then every 6 months and were assessed as per modified Harris Hip Score. RESULTS All the patients were evaluated according to the Modified Harris Hip Scoring system. The results showed 14(67% hips with excellent results, 4(19% with good results, and 3(14% hips with fair results. No poor outcome was noted in this study. 2 cases of dislocation (10% were noted one on the 5th post-operative day and the other occurred after the patient was discharged from the institution. CONCLUSIONS The management of diseased and destroyed hips with chronic pain with cemented total hip replacement is effective and gives stable, mobile and painless hip joint to the patient. Functional results are excellent and

  19. Impact of gastro-oesophageal reflux on microRNA expression, location and function

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    Smith Cameron M

    2013-01-01

    Full Text Available Abstract Background Ulceration of the oesophageal squamous mucosa (ulcerative oesophagitis is a pathological manifestation of gastro-oesophageal reflux disease, and is a major risk factor for the development of Barrett’s oesophagus. Barrett’s oesophagus is characterised by replacement of reflux-damaged oesophageal squamous epithelium with a columnar intestinal-like epithelium. We previously reported discovery of microRNAs that are differentially expressed between oesophageal squamous mucosa and Barrett’s oesophagus mucosa. Now, to better understand early steps in the initiation of Barrett’s oesophagus, we assessed the expression, location and function of these microRNAs in oesophageal squamous mucosa from individuals with ulcerative oesophagitis. Methods Quantitative real-time PCR was used to compare miR-21, 143, 145, 194, 203, 205 and 215 expression levels in oesophageal mucosa from individuals without pathological gastro-oesophageal reflux to individuals with ulcerative oesophagitis. Correlations between microRNA expression and messenger RNA differentiation markers BMP-4, CK8 and CK14 were analyzed. The cellular localisation of microRNAs within the oesophageal mucosa was determined using in-situ hybridisation. microRNA involvement in proliferation and apoptosis was assessed following transfection of a human squamous oesophageal mucosal cell line (Het-1A. Results miR-143, miR-145 and miR-205 levels were significantly higher in gastro-oesophageal reflux compared with controls. Elevated miR-143 expression correlated with BMP-4 and CK8 expression, and elevated miR-205 expression correlated negatively with CK14 expression. Endogenous miR-143, miR-145 and miR-205 expression was localised to the basal layer of the oesophageal epithelium. Transfection of miR-143, 145 and 205 mimics into Het-1A cells resulted in increased apoptosis and decreased proliferation. Conclusions Elevated miR-143, miR-145 and miR-205 expression was observed in

  20. Nuclear Factor 90, a cellular dsRNA binding protein inhibits the HIV Rev-export function

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    St-Laurent Georges

    2006-11-01

    Full Text Available Abstract Background The HIV Rev protein is known to facilitate export of incompletely spliced and unspliced viral transcripts to the cytoplasm, a necessary step in virus life cycle. The Rev-mediated nucleo-cytoplasmic transport of nascent viral transcripts, dependents on interaction of Rev with the RRE RNA structural element present in the target RNAs. The C-terminal variant of dsRNA-binding nuclear protein 90 (NF90ctv has been shown to markedly attenuate viral replication in stably transduced HIV-1 target cell line. Here we examined a mechanism of interference of viral life cycle involving Rev-NF90ctv interaction. Results Since Rev:RRE complex formations depend on protein:RNA and protein:protein interactions, we investigated whether the expression of NF90ctv might interfere with Rev-mediated export of RRE-containing transcripts. When HeLa cells expressed both NF90ctv and Rev protein, we observed that NF90ctv inhibited the Rev-mediated RNA transport. In particular, three regions of NF90ctv protein are involved in blocking Rev function. Moreover, interaction of NF90ctv with the RRE RNA resulted in the expression of a reporter protein coding sequences linked to the RRE structure. Moreover, Rev influenced the subcellular localization of NF90ctv, and this process is leptomycin B sensitive. Conclusion The dsRNA binding protein, NF90ctv competes with HIV Rev function at two levels, by competitive protein:protein interaction involving Rev binding to specific domains of NF90ctv, as well as by its binding to the RRE-RNA structure. Our results are consistent with a model of Rev-mediated HIV-1 RNA export that envisions Rev-multimerization, a process interrupted by NF90ctv.

  1. MicroRNA-21 exhibits antiangiogenic function by targeting RhoB expression in endothelial cells.

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    Céline Sabatel

    Full Text Available BACKGROUND: MicroRNAs (miRNAs are endogenously expressed small non-coding RNAs that regulate gene expression at post-transcriptional level. The recent discovery of the involvement of these RNAs in the control of angiogenesis renders them very attractive in the development of new approaches for restoring the angiogenic balance. Whereas miRNA-21 has been demonstrated to be highly expressed in endothelial cells, the potential function of this miRNA in angiogenesis has never been investigated. METHODOLOGY/PRINCIPAL FINDINGS: We first observed in endothelial cells a negative regulation of miR-21 expression by serum and bFGF, two pro-angiogenic factors. Then using in vitro angiogenic assays, we observed that miR-21 acts as a negative modulator of angiogenesis. miR-21 overexpression reduced endothelial cell proliferation, migration and the ability of these cells to form tubes whereas miR-21 inhibition using a LNA-anti-miR led to opposite effects. Expression of miR-21 in endothelial cells also led to a reduction in the organization of actin into stress fibers, which may explain the decrease in cell migration. Further mechanistic studies showed that miR-21 targets RhoB, as revealed by a decrease in RhoB expression and activity in miR-21 overexpressing cells. RhoB silencing impairs endothelial cell migration and tubulogenesis, thus providing a possible mechanism for miR-21 to inhibit angiogenesis. Finally, the therapeutic potential of miR-21 as an angiogenesis inhibitor was demonstrated in vivo in a mouse model of choroidal neovascularization. CONCLUSIONS/SIGNIFICANCE: Our results identify miR-21 as a new angiogenesis inhibitor and suggest that inhibition of cell migration and tubulogenesis is mediated through repression of RhoB.

  2. Waiting for total knee replacement surgery: factors associated with pain, stiffness, function and quality of life

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    Dionne Clermont E

    2009-05-01

    Full Text Available Abstract Background Recent evidences show that education and rehabilitation while waiting for knee replacement have positive effects on the patients' health status. Identification of factors associated with worse pain, function and health-related quality of life (HRQoL while waiting for surgery could help develop pre-surgery rehabilitation interventions that target specifically these factors and prioritize patients that may benefit the most from them. The objectives of this study were to measure pain, stiffness, function and HRQoL in patients at enrolment on waiting lists for knee replacement and to identify demographic, clinical, socioeconomic and psychosocial characteristics associated with these outcomes. Methods This study is part of a broader study measuring the effects of pre-surgery wait in patients scheduled for knee replacement. From 02/2006 to 09/2007, 197 patients newly scheduled for total knee replacement were recruited from the waiting lists of three university hospitals in Quebec City, Canada. Pain, stiffness and function were measured with the Western Ontario and McMaster Osteoarthritis Index (WOMAC and HRQoL was measured with the SF-36 Health Survey. Stepwise multiple regression analysis was used to assess the strength of the associations between the independent variables and the WOMAC and SF-36 scores. Results The scores of all eight HRQoL physical and mental domains of the SF-36 were significantly lower than aged matched Canadian normative data (p Conclusion Patients waiting for knee replacement have poor function and HRQoL. Characteristics that were found to be associated with these outcomes could help develop pre-surgery rehabilitation program and prioritize patients that may benefit the most from them. Such programs could include interventions to reduce psychological distress, therapeutic exercises targeting both knees and weight loss management.

  3. Small RNA Functions Are Required for Growth and Development of Magnaporthe oryzae.

    Science.gov (United States)

    Raman, Vidhyavathi; Simon, Stacey A; Demirci, Feray; Nakano, Mayumi; Meyers, Blake C; Donofrio, Nicole M

    2017-07-01

    RNA interference (RNAi) is conserved in eukaryotic organisms, and it has been well studied in many animal and plant species and some fungal species, yet it is not well studied in fungal plant pathogens. In the rice blast fungus Magnaporthe oryzae, we examined small RNA (sRNA) and their biogenesis in the context of growth and pathogenicity. Through genetic and genomic analyses, we demonstrate that loss of a single gene encoding Dicer, RNA-dependent RNA polymerase, or Argonaute reduces sRNA levels. These three proteins are required for the biogenesis of sRNA-matching genome-wide regions (coding regions, repeats, and intergenic regions). The loss of one Argonaute reduced both sRNA and fungal virulence on barley leaves. Transcriptome analysis of multiple mutants revealed that sRNA play an important role in transcriptional regulation of repeats and intergenic regions in M. oryzae. Together, these data support that M. oryzae sRNA regulate developmental processes including, fungal growth and virulence.

  4. Structure-Function Model for Kissing Loop Interactions That Initiate Dimerization of Ty1 RNA.

    Science.gov (United States)

    Gamache, Eric R; Doh, Jung H; Ritz, Justin; Laederach, Alain; Bellaousov, Stanislav; Mathews, David H; Curcio, M Joan

    2017-04-26

    The genomic RNA of the retrotransposon Ty1 is packaged as a dimer into virus-like particles. The 5' terminus of Ty1 RNA harbors cis -acting sequences required for translation initiation, packaging and initiation of reverse transcription (TIPIRT). To identify RNA motifs involved in dimerization and packaging, a structural model of the TIPIRT domain in vitro was developed from single-nucleotide resolution RNA structural data. In general agreement with previous models, the first 326 nucleotides of Ty1 RNA form a pseudoknot with a 7-bp stem (S1), a 1-nucleotide interhelical loop and an 8-bp stem (S2) that delineate two long, structured loops. Nucleotide substitutions that disrupt either pseudoknot stem greatly reduced helper-Ty1-mediated retrotransposition of a mini-Ty1, but only mutations in S2 destabilized mini-Ty1 RNA in cis and helper-Ty1 RNA in trans. Nested in different loops of the pseudoknot are two hairpins with complementary 7-nucleotide motifs at their apices. Nucleotide substitutions in either motif also reduced retrotransposition and destabilized mini- and helper-Ty1 RNA. Compensatory mutations that restore base-pairing in the S2 stem or between the hairpins rescued retrotransposition and RNA stability in cis and trans. These data inform a model whereby a Ty1 RNA kissing complex with two intermolecular kissing-loop interactions initiates dimerization and packaging.

  5. Structure-Function Model for Kissing Loop Interactions That Initiate Dimerization of Ty1 RNA

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    Eric R. Gamache

    2017-04-01

    Full Text Available The genomic RNA of the retrotransposon Ty1 is packaged as a dimer into virus-like particles. The 5′ terminus of Ty1 RNA harbors cis-acting sequences required for translation initiation, packaging and initiation of reverse transcription (TIPIRT. To identify RNA motifs involved in dimerization and packaging, a structural model of the TIPIRT domain in vitro was developed from single-nucleotide resolution RNA structural data. In general agreement with previous models, the first 326 nucleotides of Ty1 RNA form a pseudoknot with a 7-bp stem (S1, a 1-nucleotide interhelical loop and an 8-bp stem (S2 that delineate two long, structured loops. Nucleotide substitutions that disrupt either pseudoknot stem greatly reduced helper-Ty1-mediated retrotransposition of a mini-Ty1, but only mutations in S2 destabilized mini-Ty1 RNA in cis and helper-Ty1 RNA in trans. Nested in different loops of the pseudoknot are two hairpins with complementary 7-nucleotide motifs at their apices. Nucleotide substitutions in either motif also reduced retrotransposition and destabilized mini- and helper-Ty1 RNA. Compensatory mutations that restore base-pairing in the S2 stem or between the hairpins rescued retrotransposition and RNA stability in cis and trans. These data inform a model whereby a Ty1 RNA kissing complex with two intermolecular kissing-loop interactions initiates dimerization and packaging.

  6. Proof of the effect of electric current on function of tara calor using total derivative

    Science.gov (United States)

    Soenarto, Y.; Kusdiwelirawan, A.; Fitriana, A.; Ermawaty, I. R.; Sari, V. M. A.

    2018-01-01

    The research aims to prove the effect of electric currents on taracalor by performing observations, analyze the results using the taracalor formulation which is the literature and by the application of mathematics is the total differential. Steps performed in the research start from the study of theory, subsequent experiments, analysis and comparison of matches against the literature. The result of this research shows that the average values of taracalor for 1 Ampere, 2 Ampere, and 3 Ampere are 0,7725, 0,8894, and 0.9786, respectively. While the average functions for 1 Ampere, 2 Ampere, and 3 Ampere are (2,235 × 10-3)(dt ‑ 90dΔT), (2,57 × 10-3)(dt ‑ 25,71dΔT), and (2,833 × 10-3)(dt ‑ 12,86dΔT), respectively.

  7. Functional brain imaging of a complex navigation task following one night of total sleep deprivation

    Science.gov (United States)

    Strangman, Gary; Thompson, John H.; Strauss, Monica M.; Marshburn, Thomas H.; Sutton, Jeffrey P.

    2006-01-01

    Study Objectives: To assess the cerebral effects associated with sleep deprivation in a simulation of a complex, real-world, high-risk task. Design and Interventions: A two-week, repeated measures, cross-over experimental protocol, with counterbalanced orders of normal sleep (NS) and total sleep deprivation (TSD). Setting: Each subject underwent functional magnetic resonance imaging (fMRI) while performing a dual-joystick, 3D sensorimotor navigation task (simulated orbital docking). Scanning was performed twice per subject, once following a night of normal sleep (NS), and once following a single night of total sleep deprivation (TSD). Five runs (eight 24s docking trials each) were performed during each scanning session. Participants: Six healthy, young, right-handed volunteers (2 women; mean age 20) participated. Measurements and Results: Behavioral performance on multiple measures was comparable in the two sleep conditions. Neuroimaging results within sleep conditions revealed similar locations of peak activity for NS and TSD, including left sensorimotor cortex, left precuneus (BA 7), and right visual areas (BA 18/19). However, cerebral activation following TSD was substantially larger and exhibited higher amplitude modulations from baseline. When directly comparing NS and TSD, most regions exhibited TSD>NS activity, including multiple prefrontal cortical areas (BA 8/9,44/45,47), lateral parieto-occipital areas (BA 19/39, 40), superior temporal cortex (BA 22), and bilateral thalamus and amygdala. Only left parietal cortex (BA 7) demonstrated NS>TSD activity. Conclusions: The large network of cerebral differences between the two conditions, even with comparable behavioral performance, suggests the possibility of detecting TSD-induced stress via functional brain imaging techniques on complex tasks before stress-induced failures.

  8. A hairpin within YAP mRNA 3′UTR functions in regulation at post-transcription level

    International Nuclear Information System (INIS)

    Gao, Yuen; Wang, Yuan; Feng, Jinyan; Feng, Guoxing; Zheng, Minying; Yang, Zhe; Xiao, Zelin; Lu, Zhanping; Ye, Lihong; Zhang, Xiaodong

    2015-01-01

    The central dogma of gene expression is that DNA is transcribed into messenger RNAs, which in turn serve as the template for protein synthesis. Recently, it has been reported that mRNAs display regulatory roles that rely on their ability to compete for microRNA binding, independent of their protein-coding function. However, the regulatory mechanism of mRNAs remains poorly understood. Here, we report that a hairpin within YAP mRNA 3′untranslated region (3′UTR) functions in regulation at post-transcription level through generating endogenous siRNAs (esiRNAs). Bioinformatics analysis for secondary structure showed that YAP mRNA displayed a hairpin structure (termed standard hairpin, S-hairpin) within its 3′UTR. Surprisingly, we observed that the overexpression of S-hairpin derived from YAP 3′UTR (YAP-sh) increased the luciferase reporter activities of transcriptional factor NF-κB and AP-1 in 293T cells. Moreover, we identified that a fragment from YAP-sh, an esiRNA, was able to target mRNA 3′UTR of NF2 (a member of Hippo-signaling pathway) and YAP mRNA 3′UTR itself in hepatoma cells. Thus, we conclude that the YAP-sh within YAP mRNA 3′UTR may serve as a novel regulatory element, which functions in regulation at post-transcription level. Our finding provides new insights into the mechanism of mRNAs in regulatory function. - Highlights: • An S-hairpin within YAP mRNA 3′UTR possesses regulatory function. • YAP-sh acts as a regulatory element for YAP at post-transcription level. • YAP-sh-3p20, an esiRNA derived from YAP-sh, targets mRNAs of YAP and NF2. • YAP-sh-3p20 depresses the proliferation of HepG2 cells in vitro

  9. A hairpin within YAP mRNA 3′UTR functions in regulation at post-transcription level

    Energy Technology Data Exchange (ETDEWEB)

    Gao, Yuen; Wang, Yuan; Feng, Jinyan; Feng, Guoxing; Zheng, Minying; Yang, Zhe; Xiao, Zelin; Lu, Zhanping [State Key Laboratory of Medicinal Chemical Biology, Department of Cancer Research, College of Life Sciences, Nankai University, Tianjin 300071 (China); Ye, Lihong [State Key Laboratory of Medicinal Chemical Biology, Department of Biochemistry, College of Life Sciences, Nankai University, Tianjin 300071 (China); Zhang, Xiaodong, E-mail: zhangxd@nankai.edu.cn [State Key Laboratory of Medicinal Chemical Biology, Department of Cancer Research, College of Life Sciences, Nankai University, Tianjin 300071 (China)

    2015-04-03

    The central dogma of gene expression is that DNA is transcribed into messenger RNAs, which in turn serve as the template for protein synthesis. Recently, it has been reported that mRNAs display regulatory roles that rely on their ability to compete for microRNA binding, independent of their protein-coding function. However, the regulatory mechanism of mRNAs remains poorly understood. Here, we report that a hairpin within YAP mRNA 3′untranslated region (3′UTR) functions in regulation at post-transcription level through generating endogenous siRNAs (esiRNAs). Bioinformatics analysis for secondary structure showed that YAP mRNA displayed a hairpin structure (termed standard hairpin, S-hairpin) within its 3′UTR. Surprisingly, we observed that the overexpression of S-hairpin derived from YAP 3′UTR (YAP-sh) increased the luciferase reporter activities of transcriptional factor NF-κB and AP-1 in 293T cells. Moreover, we identified that a fragment from YAP-sh, an esiRNA, was able to target mRNA 3′UTR of NF2 (a member of Hippo-signaling pathway) and YAP mRNA 3′UTR itself in hepatoma cells. Thus, we conclude that the YAP-sh within YAP mRNA 3′UTR may serve as a novel regulatory element, which functions in regulation at post-transcription level. Our finding provides new insights into the mechanism of mRNAs in regulatory function. - Highlights: • An S-hairpin within YAP mRNA 3′UTR possesses regulatory function. • YAP-sh acts as a regulatory element for YAP at post-transcription level. • YAP-sh-3p20, an esiRNA derived from YAP-sh, targets mRNAs of YAP and NF2. • YAP-sh-3p20 depresses the proliferation of HepG2 cells in vitro.

  10. Accelerating Nanoscale Research with Neutron Total Scattering: Linking Structure and Function in Finite Materials

    Science.gov (United States)

    Page, Katharine

    2012-10-01

    h -abstract-pard In contrast to bulk materials, nanomaterials and nanoparticles, comprised of a few hundred to tens of thousands of atoms, require every atom's position to be located in order to understand their structure-property relationships. New behavior can arise with a constricted, expanded, or distorted lattice, variation in surface termination structure, ligand capping or stabilization, or with the increasingly diverse set of shapes and architectures appearing in nanoscience literature today: tubes, pyramids, stars, core-shell and matrix-confined particles, multilayer films, etc. Pair distribution function (PDF) analysis, based on spallation neutron or synchrotron x-ray total scattering data, has emerged as a very promising characterization method for nanomaterials in recent years. Total scattering methods provide information about every pair of atoms probed in a diffraction experiment and thus contain an unexploited wealth of information for finite systems. In this contribution we will present our work establishing the influence of particle size and shape on the nature and correlation of local atomic dipoles in finite ferroelectric systems. We also review current data-driven modeling capabilities and outline the need for evolution of robust computational tools to follow other complex nanoscale phenomena with scattering data. pard-/abstract-

  11. Bilateral Total Hip Arthroplasty in Femoral Head Avascular Necrosis: Functional Outcomes and Complications

    Directory of Open Access Journals (Sweden)

    Afshin Taheriazam

    2016-06-01

    Full Text Available Total hip arthroplasty (THA is one of the successful and cost-benefit surgical treatments. One-stage bilateral THA (BTHA offers many benefits. However, there are concerns about the safety of the procedure and higher complications. We aimed to evaluate the complications and outcomes of one-stage BTHA with Hardinge approach for femoral head avascular necrosis patients. A total of 60 patients from April 2009 and May 2013, were underwent one-stage bilateral total hip arthroplasty (BTHA in Milad and Erfan hospitals, Tehran, Iran. A prospective analysis of the functional outcomes and complications of one-stage BTHA through Hardinge approach in patients with femoral head avascular necrosis (AVN performed. We evaluated all patients clinically and radiologically with serial follow-ups. A clinical hip score based upon the modified Harris Hip Score (MHHS was performed preoperatively and again postoperatively. During period of study 44 men (73.3% and 16 women (26.6% with a mean age of 31.40±4.08 years (range 25 to 36 years at the time of presentation were entered. The mean surgical time was 2.6±0.38 hrs. The mean hospital stay was 3 .50±0.72 days. Hemoglobin level decreased significa ntly after operation (P= 0.046. There was no reported patient with perioperative death, deep venous thrombosis, pulmonary embolism, infection, dislocation, periprosthetic fracture or heterotrophic ossification. The mean preoperative MHHS score was 47.93±7.33 in patients. MHHS score i mproved to 95.06±3.47 in the last follow-up (P=0.0001.Our results recommend the use of one-stage BTHA through Hardinge approach in femoral head avascular necrosis patients.

  12. Integrative Analysis of MicroRNA and mRNA Data Reveals an Orchestrated Function of MicroRNAs in Skeletal Myocyte Differentiation in Response to TNF-α or IGF1

    Science.gov (United States)

    Meyer, Swanhild U.; Sass, Steffen; Mueller, Nikola S.; Krebs, Stefan; Bauersachs, Stefan; Kaiser, Sebastian; Blum, Helmut; Thirion, Christian; Krause, Sabine; Theis, Fabian J.; Pfaffl, Michael W.

    2015-01-01

    Introduction Skeletal muscle cell differentiation is impaired by elevated levels of the inflammatory cytokine tumor necrosis factor-α (TNF-α) with pathological significance in chronic diseases or inherited muscle disorders. Insulin like growth factor-1 (IGF1) positively regulates muscle cell differentiation. Both, TNF-α and IGF1 affect gene and microRNA (miRNA) expression in this process. However, computational prediction of miRNA-mRNA relations is challenged by false positives and targets which might be irrelevant in the respective cellular transcriptome context. Thus, this study is focused on functional information about miRNA affected target transcripts by integrating miRNA and mRNA expression profiling data. Methodology/Principal Findings Murine skeletal myocytes PMI28 were differentiated for 24 hours with concomitant TNF-α or IGF1 treatment. Both, mRNA and miRNA expression profiling was performed. The data-driven integration of target prediction and paired mRNA/miRNA expression profiling data revealed that i) the quantity of predicted miRNA-mRNA relations was reduced, ii) miRNA targets with a function in cell cycle and axon guidance were enriched, iii) differential regulation of anti-differentiation miR-155-5p and miR-29b-3p as well as pro-differentiation miR-335-3p, miR-335-5p, miR-322-3p, and miR-322-5p seemed to be of primary importance during skeletal myoblast differentiation compared to the other miRNAs, iv) the abundance of targets and affected biological processes was miRNA specific, and v) subsets of miRNAs may collectively regulate gene expression. Conclusions Joint analysis of mRNA and miRNA profiling data increased the process-specificity and quality of predicted relations by statistically selecting miRNA-target interactions. Moreover, this study revealed miRNA-specific predominant biological implications in skeletal muscle cell differentiation and in response to TNF-α or IGF1 treatment. Furthermore, myoblast differentiation-associated mi

  13. Functional and clinical outcomes of total ankle arthroplasty in elderly compared to younger patients.

    Science.gov (United States)

    Tenenbaum, Shay; Bariteau, Jason; Coleman, Scott; Brodsky, James

    2017-06-01

    Total ankle arthroplasty (TAA) is becoming an increasingly utilized procedure for the management of end-stage ankle arthritis. Elderly patients are the fastest growing segment of the population in the western world, creating a unique challenge to the health economics of our era. Determining if elderly patients with end-stage ankle arthritis demonstrate the same improvements in clinical outcomes and functional measures of gait following TAA would be valuable. This can aid to evaluate the utilization of TAA in this enlarging cohort of our population. Consecutive series of twenty-one patients over the age of 70, who underwent TAA for end-stage ankle arthritis, was prospectively compared to a series of twenty-one patients aged 50-60, who underwent the same procedure by single surgeon during same time period. Clinical outcomes were measured with outcome scores including VAS pain score, AOFAS Ankle and Hindfoot Score, and the SF-36. Three-dimensional gait analysis was performed preoperatively and at a minimum of one year postoperatively, to measure temporal-spatial, kinematic, and kinetic parameters of gait. Mixed model multivariate statistical analysis was used to evaluate and compare the independent contributions to outcomes of the surgical intervention over time; of patient age; and of time-plus-age interaction, as these influenced both the clinical outcomes and the functional gait outcomes. Statistically significant improvements in VAS pain scores, AOFAS ankle/hindfoot scores, and SF-36 scores were demonstrated in both age groups. Following surgery, there were improvements in all parameters of gait, including temporal-spatial parameters as step length and walking velocity; kinematic parameters, including, increase in total range of motion to a total of 17-19°; and kinetic parameters, including increase in ankle power and moment. The improvements both in clinical and gait outcomes were equivalent in the two age groups. In this comparative study, it is shown that both

  14. Convex composite wavelet frame and total variation-based image deblurring using nonconvex penalty functions

    Science.gov (United States)

    Shen, Zhengwei; Cheng, Lishuang

    2017-09-01

    Total variation (TV)-based image deblurring method can bring on staircase artifacts in the homogenous region of the latent images recovered from the degraded images while a wavelet/frame-based image deblurring method will lead to spurious noise spikes and pseudo-Gibbs artifacts in the vicinity of discontinuities of the latent images. To suppress these artifacts efficiently, we propose a nonconvex composite wavelet/frame and TV-based image deblurring model. In this model, the wavelet/frame and the TV-based methods may complement each other, which are verified by theoretical analysis and experimental results. To further improve the quality of the latent images, nonconvex penalty function is used to be the regularization terms of the model, which may induce a stronger sparse solution and will more accurately estimate the relative large gradient or wavelet/frame coefficients of the latent images. In addition, by choosing a suitable parameter to the nonconvex penalty function, the subproblem that splits by the alternative direction method of multipliers algorithm from the proposed model can be guaranteed to be a convex optimization problem; hence, each subproblem can converge to a global optimum. The mean doubly augmented Lagrangian and the isotropic split Bregman algorithms are used to solve these convex subproblems where the designed proximal operator is used to reduce the computational complexity of the algorithms. Extensive numerical experiments indicate that the proposed model and algorithms are comparable to other state-of-the-art model and methods.

  15. Quality of life and functionality after non-cemented total hiparthroplasty

    International Nuclear Information System (INIS)

    Shah, Z.A.; Arif, U.; Aslam, M.N.; Bilal, A.; Khan, M.N.B.

    2013-01-01

    Objectives: To evaluate the functional outcome of non-cemented total hip arthroplasty in terms of pain relief, functional capacity, range of motion and absence of deformity using Harris hip score. Study Design: Descriptive study. Place and Duration of Study: From January 2012 to December 2012, at Nawaz Sharif Social Security Teaching Hospital, Lahore (University College Medicine and Dentistry). Subject and Methods: Thirty patients meeting the inclusion criteria were admitted through orthopedics outpatient department of Nawaz Sharif social security hospital Lahore. Pre operative Harris scoring was done and was compared with the post operative score to find the improvement. Results: Mean age of patients was 52.53 +- 18.21 years, and 17 were males and 13 females. Average pre operative Harris Hip score was 23.77 +- 9.50 and post-operative score 87.90 +- 10.42. Conclusion: It was concluded that THR is a safe surgical procedure with promising results in relieving pain, improving movements and upgrading the quality of life. (author)

  16. Obstacles and opportunities in the functional analysis of extracellular vesicle RNA - an ISEV position paper

    OpenAIRE

    Mateescu, Bogdan; Kowal, Emma; Balkom, Bastiaan Wilhelmus Maria van; Bartel, Sabine; Bhattacharyya, Suvendra N.; Buzás, Edit I.; Buck, Amy H.; de-Candia, Paola; Chow, Franklin W. N.; Das, Saumya; Driedonks, Tom A. P.; Fernández-Messina, Lola; Haderk, Franziska; Hill, Andrew F.; Jones, Jennifer C.

    2017-01-01

    The release of RNA-containing extracellular vesicles (EV) into the extracellular milieu has been demonstrated in a multitude of different in vitro cell systems and in a variety of body fluids. RNA-containing EV are in the limelight for their capacity to communicate genetically encoded messages to other cells, their suitability as candidate biomarkers for diseases, and their use as therapeutic agents. Although EV-RNA has attracted enormous interest from basic researchers, clinicians, and indus...

  17. INFLUENCE OF RESIDUAL VARUS DEFORMITY ON CLINICAL, FUNCTIONAL, RADIOLOGICAL AND DYNAMOMETRIC OUTCOMES OF TOTAL KNEE ARTHROPLASTY

    Directory of Open Access Journals (Sweden)

    M. P. Zinoviev

    2017-01-01

    Full Text Available Purpose of the study – to evaluate the influence of residual varus deformity of the lower leg on clinical, functional and dynamometric outcomes of TKR.Material and methods. In the period from September 2014 till May 2015 951 total knee replacement surgeries were performed in Ural clinical and rehabilitation center in 933 patients with initial varus deformity of lower limb. However, in 52 cases (5.5% residual varus deformity of >3° persisted. The main group included 36 patients with mean residual varus deformity of lower leg of 3.9°±0.74° (from 3.1° to 5.6°, the control group included 34 patients with neutral mechanic axis of the lower leg. Evaluation of treatment outcomes was performed on average 14.2±1.8 months after the procedure using functional assessment of the patients according to KSS (Knee Society Score, subjective assessment of life quality (SF36, as well as evaluation of the static-dynamic function of the lower leg on diagnostic and treatment complex Biodex Systems 4 Quick Set and assessment of stability of components fixation by F.C. Ewald scale in modification of O.A. Kudinov et al.Results. Functional assessment of the patients according to KSS (Knee Society Score for the main group was 84.0±4.6, in control group – 82.2±4.1 points (p>0.05. Subjective assessment of life quality (SF36 in the main group was 162.6±6.4 points, in control group – 164.3±8.1 points (p>0.05. In terms of flexion and extension of lower leg there were no significant differences between two groups when assessing static and dynamic function by treatment and diagnostic complex «Biodex Systems 4 Quick Set» on average in 14.2±1.8 months after procedure: neither isometric nor isokinetic parameters in both groups demonstrated statistically significant differences (p>0.05. Based on X-rays evaluation there were no abnormalities in components fixation in both groups. There were no statistically significant differences identified in KSS and SF36 scales

  18. Functions of the nonsense-mediated mRNA decay pathway in Drosophila development.

    Directory of Open Access Journals (Sweden)

    Mark M Metzstein

    2006-12-01

    Full Text Available Nonsense-mediated mRNA decay (NMD is a cellular surveillance mechanism that degrades transcripts containing premature translation termination codons, and it also influences expression of certain wild-type transcripts. Although the biochemical mechanisms of NMD have been studied intensively, its developmental functions and importance are less clear. Here, we describe the isolation and characterization of Drosophila "photoshop" mutations, which increase expression of green fluorescent protein and other transgenes. Mapping and molecular analyses show that photoshop mutations are loss-of-function mutations in the Drosophila homologs of NMD genes Upf1, Upf2, and Smg1. We find that Upf1 and Upf2 are broadly active during development, and they are required for NMD as well as for proper expression of dozens of wild-type genes during development and for larval viability. Genetic mosaic analysis shows that Upf1 and Upf2 are required for growth and/or survival of imaginal cell clones, but this defect can be overcome if surrounding wild-type cells are eliminated. By contrast, we find that the PI3K-related kinase Smg1 potentiates but is not required for NMD or for viability, implying that the Upf1 phosphorylation cycle that is required for mammalian and Caenorhabditis elegans NMD has a more limited role during Drosophila development. Finally, we show that the SV40 3' UTR, present in many Drosophila transgenes, targets the transgenes for regulation by the NMD pathway. The results establish that the Drosophila NMD pathway is broadly active and essential for development, and one critical function of the pathway is to endow proliferating imaginal cells with a competitive growth advantage that prevents them from being overtaken by other proliferating cells.

  19. Structure-function studies of STAR family Quaking proteins bound to their in vivo RNA target sites

    Energy Technology Data Exchange (ETDEWEB)

    Teplova, Marianna; Hafner, Markus; Teplov, Dmitri; Essig, Katharina; Tuschl, Thomas; Patel, Dinshaw J. [MSKCC; (Rockefeller)

    2013-09-27

    Mammalian Quaking (QKI) and its Caenorhabditis elegans homolog, GLD-1 (defective in germ line development), are evolutionarily conserved RNA-binding proteins, which post-transcriptionally regulate target genes essential for developmental processes and myelination. We present X-ray structures of the STAR (signal transduction and activation of RNA) domain, composed of Qua1, K homology (KH), and Qua2 motifs of QKI and GLD-1 bound to high-affinity in vivo RNA targets containing YUAAY RNA recognition elements (RREs). The KH and Qua2 motifs of the STAR domain synergize to specifically interact with bases and sugar-phosphate backbones of the bound RRE. Qua1-mediated homodimerization generates a scaffold that enables concurrent recognition of two RREs, thereby plausibly targeting tandem RREs present in many QKI-targeted transcripts. Structure-guided mutations reduced QKI RNA-binding affinity in vitro and in vivo, and expression of QKI mutants in human embryonic kidney cells (HEK293) significantly decreased the abundance of QKI target mRNAs. Overall, our studies define principles underlying RNA target selection by STAR homodimers and provide insights into the post-transcriptional regulatory function of mammalian QKI proteins.

  20. Enhancer-associated long non-coding RNA LEENE regulates endothelial nitric oxide synthase and endothelial function.

    Science.gov (United States)

    Miao, Yifei; Ajami, Nassim E; Huang, Tse-Shun; Lin, Feng-Mao; Lou, Chih-Hong; Wang, Yun-Ting; Li, Shuai; Kang, Jian; Munkacsi, Hannah; Maurya, Mano R; Gupta, Shakti; Chien, Shu; Subramaniam, Shankar; Chen, Zhen

    2018-01-18

    The optimal expression of endothelial nitric oxide synthase (eNOS), the hallmark of endothelial homeostasis, is vital to vascular function. Dynamically regulated by various stimuli, eNOS expression is modulated at transcriptional, post-transcriptional, and post-translational levels. However, epigenetic modulations of eNOS, particularly through long non-coding RNAs (lncRNAs) and chromatin remodeling, remain to be explored. Here we identify an enhancer-associated lncRNA that enhances eNOS expression (LEENE). Combining RNA-sequencing and chromatin conformation capture methods, we demonstrate that LEENE is co-regulated with eNOS and that its enhancer resides in proximity to eNOS promoter in endothelial cells (ECs). Gain- and Loss-of-function of LEENE differentially regulate eNOS expression and EC function. Mechanistically, LEENE facilitates the recruitment of RNA Pol II to the eNOS promoter to enhance eNOS nascent RNA transcription. Our findings unravel a new layer in eNOS regulation and provide novel insights into cardiovascular regulation involving endothelial function.

  1. Functional genome-wide siRNA screen identifies KIAA0586 as mutated in Joubert syndrome

    NARCIS (Netherlands)

    Roosing, S.; Hofree, M.; Kim, S.; Scott, E.; Copeland, B.; Romani, M.; Silhavy, J.L.; Rosti, R.O.; Schroth, J.; Mazza, T.; Miccinilli, E.; Zaki, M.S.; Swoboda, K.J.; Milisa-Drautz, J.; Dobyns, W.B.; Mikati, M.A.; Incecik, F.; Azam, M.; Borgatti, R.; Romaniello, R.; Boustany, R.M.; Clericuzio, C.L.; D'Arrigo, S.; Stromme, P.; Boltshauser, E.; Stanzial, F.; Mirabelli-Badenier, M.; Moroni, I.; Bertini, E.; Emma, F.; Steinlin, M.; Hildebrandt, F.; Johnson, C.A.; Freilinger, M.; Vaux, K.K.; Gabriel, S.B.; Aza-Blanc, P.; Heynen-Genel, S.; Ideker, T.; Dynlacht, B.D.; Lee, J.E.; Valente, E.M.; Kim, J.; Gleeson, J.G.

    2015-01-01

    Defective primary ciliogenesis or cilium stability forms the basis of human ciliopathies, including Joubert syndrome (JS), with defective cerebellar vermis development. We performed a high-content genome-wide small interfering RNA (siRNA) screen to identify genes regulating ciliogenesis as

  2. Studies on the structure and function of 16S ribosomal RNA using ...

    Indian Academy of Sciences (India)

    Recent technological developments permit us to examine the accessibility of specific atoms on any nucleotide in any large RNA molecule to certain chemical probes. This can provide detailed information about the higher order structure of large RNA molecules, including secondary and tertiary structure, protein-RNA ...

  3. Obstacles and opportunities in the functional analysis of extracellular vesicle RNA - an ISEV position paper

    NARCIS (Netherlands)

    Mateescu, Bogdan; Kowal, Emma J K; van Balkom, Bas W M; Bartel, Sabine; Bhattacharyya, Suvendra N; Buzás, Edit I; Buck, Amy H; de Candia, Paola; Chow, Franklin W N; Das, Saumya; Driedonks, Tom A P; Fernández-Messina, Lola; Haderk, Franziska; Hill, Andrew F; Jones, Jennifer C; Van Keuren-Jensen, Kendall R; Lai, Charles P; Lässer, Cecilia; Liegro, Italia di; Lunavat, Taral R; Lorenowicz, Magdalena J; Maas, Sybren L N; Mäger, Imre; Mittelbrunn, Maria; Momma, Stefan; Mukherjee, Kamalika; Nawaz, Muhammed; Pegtel, D Michiel; Pfaffl, Michael W; Schiffelers, Raymond M|info:eu-repo/dai/nl/212909509; Tahara, Hidetoshi; Théry, Clotilde; Tosar, Juan Pablo; Wauben, Marca H M|info:eu-repo/dai/nl/112675735; Witwer, Kenneth W; Nolte-'t Hoen, Esther N M

    2017-01-01

    The release of RNA-containing extracellular vesicles (EV) into the extracellular milieu has been demonstrated in a multitude of different in vitro cell systems and in a variety of body fluids. RNA-containing EV are in the limelight for their capacity to communicate genetically encoded messages to

  4. Obstacles and opportunities in the functional analysis of extracellular vesicle RNA - An ISEV position paper

    NARCIS (Netherlands)

    Mateescu, Bogdan; Kowal, Emma J K; van Balkom, Bas W M|info:eu-repo/dai/nl/256594783; Bartel, Sabine; Bhattacharyya, Suvendra N.; Buzás, Edit I.; Buck, Amy H.; de Candia, Paola; Chow, Franklin W N; Das, Saumya; Driedonks, Tom A P; Fernández-Messina, Lola; Haderk, Franziska; Hill, Andrew F.; Jones, Jennifer C.; Van Keuren-Jensen, Kendall R.; Lai, Charles P.; Lässer, Cecilia; di Liegro, Italia; Lunavat, Taral R.; Lorenowicz, Magdalena J.; Maas, Sybren L N; Mäger, Imre; Mittelbrunn, Maria; Momma, Stefan; Mukherjee, Kamalika; Nawaz, Muhammed; Pegtel, D. Michiel; Pfaffl, Michael W.; Schiffelers, Raymond M.|info:eu-repo/dai/nl/212909509; Tahara, Hidetoshi; Théry, Clotilde; Tosar, Juan Pablo; Wauben, Marca H M; Witwer, Kenneth W.; Nolte-'t Hoen, Esther N M

    2017-01-01

    The release of RNA-containing extracellular vesicles (EV) into the extracellular milieu has been demonstrated in a multitude of different in vitro cell systems and in a variety of body fluids. RNA-containing EV are in the limelight for their capacity to communicate genetically encoded messages to

  5. Functional specialization of the small interfering RNA pathway in response to virus infection.

    Directory of Open Access Journals (Sweden)

    Joao Trindade Marques

    Full Text Available In Drosophila, post-transcriptional gene silencing occurs when exogenous or endogenous double stranded RNA (dsRNA is processed into small interfering RNAs (siRNAs by Dicer-2 (Dcr-2 in association with a dsRNA-binding protein (dsRBP cofactor called Loquacious (Loqs-PD. siRNAs are then loaded onto Argonaute-2 (Ago2 by the action of Dcr-2 with another dsRBP cofactor called R2D2. Loaded Ago2 executes the destruction of target RNAs that have sequence complementarity to siRNAs. Although Dcr-2, R2D2, and Ago2 are essential for innate antiviral defense, the mechanism of virus-derived siRNA (vsiRNA biogenesis and viral target inhibition remains unclear. Here, we characterize the response mechanism mediated by siRNAs against two different RNA viruses that infect Drosophila. In both cases, we show that vsiRNAs are generated by Dcr-2 processing of dsRNA formed during viral genome replication and, to a lesser extent, viral transcription. These vsiRNAs seem to preferentially target viral polyadenylated RNA to inhibit viral replication. Loqs-PD is completely dispensable for silencing of the viruses, in contrast to its role in silencing endogenous targets. Biogenesis of vsiRNAs is independent of both Loqs-PD and R2D2. R2D2, however, is required for sorting and loading of vsiRNAs onto Ago2 and inhibition of viral RNA expression. Direct injection of viral RNA into Drosophila results in replication that is also independent of Loqs-PD. This suggests that triggering of the antiviral pathway is not related to viral mode of entry but recognition of intrinsic features of virus RNA. Our results indicate the existence of a vsiRNA pathway that is separate from the endogenous siRNA pathway and is specifically triggered by virus RNA. We speculate that this unique framework might be necessary for a prompt and efficient antiviral response.

  6. Functional Outcomes 18 Months After Total and Midarm Transplantation: A Case Report.

    Science.gov (United States)

    Iglesias, M; Ramírez-Berumen, M; Butrón, P; Alberú-Gómez, J; Salazar-Hernández, F; Macias-Gallardo, J; Leal-Villalpando, R P; Zamudio-Bautista, J; Acosta, V; Jauregui, L; Hernández-Campos, A; Espinosa-Cruz, V; Vázquez-Lamadrid, J; González-Sánchez, J; Cuellar-Rodriguez, J; Sierra-Madero, J G; Gaytan-Cervantes, R; Contreras-Barbosa, S; Navarro-Lara, A; Guzman-Gonzalez, J; Domínguez-Cherit, J; Vilatoba, M; Toussaint-Caire, S; Vega-Boada, F; Gómez-Pérez, F J; Mayorquin-Ruiz, M

    2018-03-16

    The function reported after arm transplantation is deemed beneficial relative to the marked disability that upper arm amputation causes. We report a 51-year-old man with a Disabilities of the Arm, Shoulder and Hand (DASH) score of 75.83 who underwent bilateral arm transplantation in October 2015. The right arm was transplanted at the glenohumeral joint level, including transplantation of the humeral head, joint capsule, and rotator cuff ligaments and tendons. Additionally, neurorrhaphies were performed at the origin of the terminal branches of the brachial plexus, including the axillary and musculocutaneous nerves. Therefore, this was considered a total arm transplantation. The left arm was transplanted at the transhumeral level, with complete transplantation of the biceps and triceps brachii, and terminolateral neurorrhaphy of the donor musculocutaneous nerve to the receptor radial nerve. A maintenance triple immunosuppression scheme was administered, with tacrolimus levels kept at 10 ng/mL. At 18 months post-transplantation, the intrinsic musculature in the left hand showed electrical registry, DASH score was 67.5, Carroll test score was 28 in both extremities, Hand Transplant Score System was 67.5 in the right extremity and 77.5 in the left extremity, and Short Form-36 score was 96.1. The patient was healthy, with restored body integrity. He could lift medium-sized weightless objects, eat and go to the bathroom by himself, drink liquids with bimanual grasp, swim, dress almost independently, and drive. The functional evolution of the patient was similar to previously reported transplanted arms, even though the right arm transplant involved the glenohumeral joint and axillary and musculocutaneous nerve repair. Copyright © 2017 Elsevier Inc. All rights reserved.

  7. SL1 revisited: functional analysis of the structure and conformation of HIV-1 genome RNA.

    Science.gov (United States)

    Sakuragi, Sayuri; Yokoyama, Masaru; Shioda, Tatsuo; Sato, Hironori; Sakuragi, Jun-Ichi

    2016-11-11

    The dimer initiation site/dimer linkage sequence (DIS/DLS) region of HIV is located on the 5' end of the viral genome and suggested to form complex secondary/tertiary structures. Within this structure, stem-loop 1 (SL1) is believed to be most important and an essential key to dimerization, since the sequence and predicted secondary structure of SL1 are highly stable and conserved among various virus subtypes. In particular, a six-base palindromic sequence is always present at the hairpin loop of SL1 and the formation of kissing-loop structure at this position between the two strands of genomic RNA is suggested to trigger dimerization. Although the higher-order structure model of SL1 is well accepted and perhaps even undoubted lately, there could be stillroom for consideration to depict the functional SL1 structure while in vivo (in virion or cell). In this study, we performed several analyses to identify the nucleotides and/or basepairing within SL1 which are necessary for HIV-1 genome dimerization, encapsidation, recombination and infectivity. We unexpectedly found that some nucleotides that are believed to contribute the formation of the stem do not impact dimerization or infectivity. On the other hand, we found that one G-C basepair involved in stem formation may serve as an alternative dimer interactive site. We also report on our further investigation of the roles of the palindromic sequences on viral replication. Collectively, we aim to assemble a more-comprehensive functional map of SL1 on the HIV-1 viral life cycle. We discovered several possibilities for a novel structure of SL1 in HIV-1 DLS. The newly proposed structure model suggested that the hairpin loop of SL1 appeared larger, and genome dimerization process might consist of more complicated mechanism than previously understood. Further investigations would be still required to fully understand the genome packaging and dimerization of HIV.

  8. Successful reprogramming of cellular protein production through mRNA delivered by functionalized lipid nanoparticles.

    Science.gov (United States)

    Yanez Arteta, Marianna; Kjellman, Tomas; Bartesaghi, Stefano; Wallin, Simonetta; Wu, Xiaoqiu; Kvist, Alexander J; Dabkowska, Aleksandra; Székely, Noémi; Radulescu, Aurel; Bergenholtz, Johan; Lindfors, Lennart

    2018-04-10

    The development of safe and efficacious gene vectors has limited greatly the potential for therapeutic treatments based on messenger RNA (mRNA). Lipid nanoparticles (LNPs) formed by an ionizable cationic lipid (here DLin-MC3-DMA), helper lipids (distearoylphosphatidylcholine, DSPC, and cholesterol), and a poly(ethylene glycol) (PEG) lipid have been identified as very promising delivery vectors of short interfering RNA (siRNA) in different clinical phases; however, delivery of high-molecular weight RNA has been proven much more demanding. Herein we elucidate the structure of hEPO modified mRNA-containing LNPs of different sizes and show how structural differences affect transfection of human adipocytes and hepatocytes, two clinically relevant cell types. Employing small-angle scattering, we demonstrate that LNPs have a disordered inverse hexagonal internal structure with a characteristic distance around 6 nm in presence of mRNA, whereas LNPs containing no mRNA do not display this structure. Furthermore, using contrast variation small-angle neutron scattering, we show that one of the lipid components, DSPC, is localized mainly at the surface of mRNA-containing LNPs. By varying LNP size and surface composition we demonstrate that both size and structure have significant influence on intracellular protein production. As an example, in both human adipocytes and hepatocytes, protein expression levels for 130 nm LNPs can differ as much as 50-fold depending on their surface characteristics, likely due to a difference in the ability of LNP fusion with the early endosome membrane. We consider these discoveries to be fundamental and opening up new possibilities for rational design of synthetic nanoscopic vehicles for mRNA delivery. Copyright © 2018 the Author(s). Published by PNAS.

  9. [Risk factors for total hip arthroplasty dislocation and its functional outcomes].

    Science.gov (United States)

    Sanz-Reig, J; Lizaur-Utrilla, A; Miralles-Muñoz, F

    2015-01-01

    To evaluate risk factors for dislocation after primary total hip arthroplasty (THA), and its functional outcomes. A retrospective study was conducted on 22 cases with dislocation and 431 controls without dislocation, all performed with lateral access. The data were collected prospectively with a minimum follow-up of 5 years. Patient related factors, from primary surgery, and position of components on simple radiographs were analyzed. Harris and Merle D'Aubigné hip scores, and short Womac questionnaire were used. Demographic, patient-related or surgical technique factors were not risk predictors of dislocation. With regard to position of components, an acetabular abduction >50° (p = 0.003) and anteversion lower or higher than 10°- 20° (p = 0.044) were risk factors. Controls and dislocation treated conservatively had similar outcomes, and in both were better than in those treated with surgical revision (p = 0.03). Factors relating to the patient and soft tissues status may influence the stability of the arthroplasty, but malposition of the acetabular component seems to be the most important and common risk factor for dislocation. Copyright © 2014. Published by Elsevier Espana.

  10. Functionalized silicon quantum dots tailored for targeted siRNA delivery

    Energy Technology Data Exchange (ETDEWEB)

    Klein, S. [Department Chemistry and Pharmacy, Physical Chemistry I and ICMM, Friedrich-Alexander University of Erlangen-Nuremberg, Egerlandstr. 3, D-91058 Erlangen (Germany); Zolk, O.; Fromm, M.F. [Institute of Experimental and Clinical Pharmacology and Toxicology, Friedrich-Alexander University of Erlangen-Nuremberg, Fahrstr. 17, D-91054 Erlangen (Germany); Schroedl, F. [Institute of Anatomy I, Friedrich-Alexander University of Erlangen-Nuremberg, Krankenhausstr. 9, D-91054 Erlangen (Germany); Departments of Anatomy/Ophthalmology, Paracelsus Medical University, Strubergasse 21, A-5020 Salzburg (Austria); Neuhuber, W. [Institute of Anatomy I, Friedrich-Alexander University of Erlangen-Nuremberg, Krankenhausstr. 9, D-91054 Erlangen (Germany); Kryschi, C., E-mail: kryschi@chemie.uni-erlangen.de [Department Chemistry and Pharmacy, Physical Chemistry I and ICMM, Friedrich-Alexander University of Erlangen-Nuremberg, Egerlandstr. 3, D-91058 Erlangen (Germany)

    2009-09-11

    For RNA interference (RNAi) mediated silencing of the ABCB1 gene in Caco-2 cells biocompatible luminescent silicon quantum dots (SiQDs) were developed to serve as self-tracking transfection tool for ABCB1 siRNA. While the 2-3 nm sized SiQD core exhibits green luminescence, the QD surfaces are completely saturated with covalently linked 2-vinylpyridine that may electrostatically bind siRNA. For down-regulating P-glycoprotein (Pgp) expression of the ABCB1 gene the SiQDs were complexed with siRNA. The cellular uptake and allocation of SiQD-siRNA complexes in Caco-2 cells were monitored using confocal laser scanning microscopy and transmission electron microscopy. The release of siRNA to the cytoplasm was verified through real-time PCR quantification of the reduced ABCB1 mRNA level. Additional evidence was obtained from time-resolved in situ fluorescence spectroscopic monitoring of the Pgp efflux dynamics in transfected Caco-2 cells which yielded significantly reduced transporter efficiencies for the Pgp substrate Rhodamine 123.

  11. A galactose-functionalized dendritic siRNA-nanovector to potentiate hepatitis C inhibition in liver cells

    Science.gov (United States)

    Lakshminarayanan, Abirami; Reddy, B. Uma; Raghav, Nallani; Ravi, Vijay Kumar; Kumar, Anuj; Maiti, Prabal K.; Sood, A. K.; Jayaraman, N.; Das, Saumitra

    2015-10-01

    A RNAi based antiviral strategy holds the promise to impede hepatitis C viral (HCV) infection overcoming the problem of emergence of drug resistant variants, usually encountered in the interferon free direct-acting antiviral therapy. Targeted delivery of siRNA helps minimize adverse `off-target' effects and maximize the efficacy of therapeutic response. Herein, we report the delivery of siRNA against the conserved 5'-untranslated region (UTR) of HCV RNA using a liver-targeted dendritic nano-vector functionalized with a galactopyranoside ligand (DG). Physico-chemical characterization revealed finer details of complexation of DG with siRNA, whereas molecular dynamic simulations demonstrated sugar moieties projecting ``out'' in the complex. Preferential delivery of siRNA to the liver was achieved through a highly specific ligand-receptor interaction between dendritic galactose and the asialoglycoprotein receptor. The siRNA-DG complex exhibited perinuclear localization in liver cells and co-localization with viral proteins. The histopathological studies showed the systemic tolerance and biocompatibility of DG. Further, whole body imaging and immunohistochemistry studies confirmed the preferential delivery of the nucleic acid to mice liver. Significant decrease in HCV RNA levels (up to 75%) was achieved in HCV subgenomic replicon and full length HCV-JFH1 infectious cell culture systems. The multidisciplinary approach provides the `proof of concept' for restricted delivery of therapeutic siRNAs using a target oriented dendritic nano-vector.A RNAi based antiviral strategy holds the promise to impede hepatitis C viral (HCV) infection overcoming the problem of emergence of drug resistant variants, usually encountered in the interferon free direct-acting antiviral therapy. Targeted delivery of siRNA helps minimize adverse `off-target' effects and maximize the efficacy of therapeutic response. Herein, we report the delivery of siRNA against the conserved 5'-untranslated

  12. RNA polymerase II transcriptional fidelity control and its functional interplay with DNA modifications.

    Science.gov (United States)

    Xu, Liang; Wang, Wei; Chong, Jenny; Shin, Ji Hyun; Xu, Jun; Wang, Dong

    2015-01-01

    Accurate genetic information transfer is essential for life. As a key enzyme involved in the first step of gene expression, RNA polymerase II (Pol II) must maintain high transcriptional fidelity while it reads along DNA template and synthesizes RNA transcript in a stepwise manner during transcription elongation. DNA lesions or modifications may lead to significant changes in transcriptional fidelity or transcription elongation dynamics. In this review, we will summarize recent progress toward understanding the molecular basis of RNA Pol II transcriptional fidelity control and impacts of DNA lesions and modifications on Pol II transcription elongation.

  13. A functional screen implicates microRNA-138-dependent regulation of the depalmitoylation enzyme APT1 in dendritic spine morphogenesis

    DEFF Research Database (Denmark)

    Siegel, Gabriele; Obernosterer, Gregor; Fiore, Roberto

    2009-01-01

    The microRNA pathway has been implicated in the regulation of synaptic protein synthesis and ultimately in dendritic spine morphogenesis, a phenomenon associated with long-lasting forms of memory. However, the particular microRNAs (miRNAs) involved are largely unknown. Here we identify specific mi......RNAs that function at synapses to control dendritic spine structure by performing a functional screen. One of the identified miRNAs, miR-138, is highly enriched in the brain, localized within dendrites and negatively regulates the size of dendritic spines in rat hippocampal neurons. miR-138 controls the expression...... of acyl protein thioesterase 1 (APT1), an enzyme regulating the palmitoylation status of proteins that are known to function at the synapse, including the alpha(13) subunits of G proteins (Galpha(13)). RNA-interference-mediated knockdown of APT1 and the expression of membrane-localized Galpha(13) both...

  14. Functional characterization of a juvenile hormone esterase related gene in the moth Sesamia nonagrioides through RNA interference.

    Directory of Open Access Journals (Sweden)

    Dimitrios Kontogiannatos

    Full Text Available Juvenile hormone esterase (JHE is a carboxylesterase that has attracted great interest because of its critical role in regulating larval to adult transition in insects and other arthropods. Previously, we characterized an ecdysteroid sensitive and juvenile hormone non-susceptible juvenile hormone esterase related gene (SnJHER in the corn stalk borer, Sesamia nonagrioides. SnJHER was rhythmically up-regulated close to each molt during the corn stalk borer's larval development. In this paper we attempted to functionally characterize SnJHER using several reverse genetics techniques. To functionally characterize SnJHER, we experimented with different dsRNA administration methods, including hemolymph, bacterial or baculovirus-mediated RNA interference, (RNAi. Our findings indicate the potential implication of SnJHER in the developmental programming of Sesamia nonagrioides. It is still unclear whether SnJHER is closely related to the authentic JHE gene, with different or similar biological functions.

  15. The human nucleolar protein FTSJ3 associates with NIP7 and functions in pre-rRNA processing.

    Directory of Open Access Journals (Sweden)

    Luis G Morello

    Full Text Available NIP7 is one of the many trans-acting factors required for eukaryotic ribosome biogenesis, which interacts with nascent pre-ribosomal particles and dissociates as they complete maturation and are exported to the cytoplasm. By using conditional knockdown, we have shown previously that yeast Nip7p is required primarily for 60S subunit synthesis while human NIP7 is involved in the biogenesis of 40S subunit. This raised the possibility that human NIP7 interacts with a different set of proteins as compared to the yeast protein. By using the yeast two-hybrid system we identified FTSJ3, a putative ortholog of yeast Spb1p, as a human NIP7-interacting protein. A functional association between NIP7 and FTSJ3 is further supported by colocalization and coimmunoprecipitation analyses. Conditional knockdown revealed that depletion of FTSJ3 affects cell proliferation and causes pre-rRNA processing defects. The major pre-rRNA processing defect involves accumulation of the 34S pre-rRNA encompassing from site A' to site 2b. Accumulation of this pre-rRNA indicates that processing of sites A0, 1 and 2 are slower in cells depleted of FTSJ3 and implicates FTSJ3 in the pathway leading to 18S rRNA maturation as observed previously for NIP7. The results presented in this work indicate a close functional interaction between NIP7 and FTSJ3 during pre-rRNA processing and show that FTSJ3 participates in ribosome synthesis in human cells.

  16. Analysis and prediction of translation rate based on sequence and functional features of the mRNA.

    Directory of Open Access Journals (Sweden)

    Tao Huang

    Full Text Available Protein concentrations depend not only on the mRNA level, but also on the translation rate and the degradation rate. Prediction of mRNA's translation rate would provide valuable information for in-depth understanding of the translation mechanism and dynamic proteome. In this study, we developed a new computational model to predict the translation rate, featured by (1 integrating various sequence-derived and functional features, (2 applying the maximum relevance & minimum redundancy method and incremental feature selection to select features to optimize the prediction model, and (3 being able to predict the translation rate of RNA into high or low translation rate category. The prediction accuracies under rich and starvation condition were 68.8% and 70.0%, respectively, evaluated by jackknife cross-validation. It was found that the following features were correlated with translation rate: codon usage frequency, some gene ontology enrichment scores, number of RNA binding proteins known to bind its mRNA product, coding sequence length, protein abundance and 5'UTR free energy. These findings might provide useful information for understanding the mechanisms of translation and dynamic proteome. Our translation rate prediction model might become a high throughput tool for annotating the translation rate of mRNAs in large-scale.

  17. AGO6 functions in RNA-mediated transcriptional gene silencing in shoot and root meristems in Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Changho Eun

    Full Text Available RNA-directed DNA methylation (RdDM is a small interfering RNA (siRNA-mediated epigenetic modification that contributes to transposon silencing in plants. RdDM requires a complex transcriptional machinery that includes specialized RNA polymerases, named Pol IV and Pol V, as well as chromatin remodelling proteins, transcription factors, RNA binding proteins, and other plant-specific proteins whose functions are not yet clarified. In Arabidopsis thaliana, DICER-LIKE3 and members of the ARGONAUTE4 group of ARGONAUTE (AGO proteins are involved, respectively, in generating and using 24-nt siRNAs that trigger methylation and transcriptional gene silencing of homologous promoter sequences. AGO4 is the main AGO protein implicated in the RdDM pathway. Here we report the identification of the related AGO6 in a forward genetic screen for mutants defective in RdDM and transcriptional gene silencing in shoot and root apical meristems in Arabidopsis thaliana. The identification of AGO6, and not AGO4, in our screen is consistent with the primary expression of AGO6 in shoot and root growing points.

  18. Novel functions for chromatin dynamics in mRNA biogenesis beyond transcription.

    Science.gov (United States)

    Dargemont, Catherine; Babour, Anna

    2017-09-03

    The first step of gene expression results in the production of mRNA ribonucleoparticles (mRNPs) that are exported to the cytoplasm via the NPC for translation into the cytoplasm. During this process, the mRNA molecule synthesized by RNA polymerase II (Pol II) undergoes extensive maturation, folding and packaging events that are intimately coupled to its synthesis. All these events take place in a chromatin context and it is therefore not surprising that a growing number of studies recently reported specific contributions of chromatin dynamics to various steps of mRNP biogenesis. In this extra view, we replace our recent findings highlighting the contribution of the yeast chromatin remodeling complex ISW1 to nuclear mRNA quality control in the context of the recent literature.

  19. Developmental and Functional Expression of miRNA-Stability Related Genes in the Nervous System

    OpenAIRE

    de Sousa, ?rica; Walter, Lais Takata; Higa, Guilherme Shigueto Vilar; Casado, Ot?vio Augusto Nocera; Kihara, Alexandre Hiroaki

    2013-01-01

    In the nervous system, control of gene expression by microRNAs (miRNAs) has been investigated in fundamental processes, such as development and adaptation to ambient demands. The action of these short nucleotide sequences on specific genes depends on intracellular concentration, which in turn reflects the balance of biosynthesis and degradation. Whereas mechanisms underlying miRNA biogenesis has been investigated in recent studies, little is known about miRNA-stability related proteins. We fi...

  20. Total sequence decomposition distinguishes functional modules, "molegos" in apurinic/apyrimidinic endonucleases

    Directory of Open Access Journals (Sweden)

    Braun Werner

    2002-11-01

    Full Text Available Abstract Background Total sequence decomposition, using the web-based MASIA tool, identifies areas of conservation in aligned protein sequences. By structurally annotating these motifs, the sequence can be parsed into individual building blocks, molecular legos ("molegos", that can eventually be related to function. Here, the approach is applied to the apurinic/apyrimidinic endonuclease (APE DNA repair proteins, essential enzymes that have been highly conserved throughout evolution. The APEs, DNase-1 and inositol 5'-polyphosphate phosphatases (IPP form a superfamily that catalyze metal ion based phosphorolysis, but recognize different substrates. Results MASIA decomposition of APE yielded 12 sequence motifs, 10 of which are also structurally conserved within the family and are designated as molegos. The 12 motifs include all the residues known to be essential for DNA cleavage by APE. Five of these molegos are sequentially and structurally conserved in DNase-1 and the IPP family. Correcting the sequence alignment to match the residues at the ends of two of the molegos that are absolutely conserved in each of the three families greatly improved the local structural alignment of APEs, DNase-1 and synaptojanin. Comparing substrate/product binding of molegos common to DNase-1 showed that those distinctive for APEs are not directly involved in cleavage, but establish protein-DNA interactions 3' to the abasic site. These additional bonds enhance both specific binding to damaged DNA and the processivity of APE1. Conclusion A modular approach can improve structurally predictive alignments of homologous proteins with low sequence identity and reveal residues peripheral to the traditional "active site" that control the specificity of enzymatic activity.

  1. The structure and function of an RNA polymerase interaction domain in the PcrA/UvrD helicase.

    Science.gov (United States)

    Sanders, Kelly; Lin, Chia-Liang; Smith, Abigail J; Cronin, Nora; Fisher, Gemma; Eftychidis, Vasileios; McGlynn, Peter; Savery, Nigel J; Wigley, Dale B; Dillingham, Mark S

    2017-04-20

    The PcrA/UvrD helicase functions in multiple pathways that promote bacterial genome stability including the suppression of conflicts between replication and transcription and facilitating the repair of transcribed DNA. The reported ability of PcrA/UvrD to bind and backtrack RNA polymerase (1,2) might be relevant to these functions, but the structural basis for this activity is poorly understood. In this work, we define a minimal RNA polymerase interaction domain in PcrA, and report its crystal structure at 1.5 Å resolution. The domain adopts a Tudor-like fold that is similar to other RNA polymerase interaction domains, including that of the prototype transcription-repair coupling factor Mfd. Removal or mutation of the interaction domain reduces the ability of PcrA/UvrD to interact with and to remodel RNA polymerase complexes in vitro. The implications of this work for our understanding of the role of PcrA/UvrD at the interface of DNA replication, transcription and repair are discussed. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  2. MicroRNA-373 functions as an oncogene and targets YOD1 gene in cervical cancer

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Luo-Qiao; Zhang, Yue; Yan, Huan; Liu, Kai-Jiang, E-mail: liukaijiang@126.com; Zhang, Shu, E-mail: drzhangshu@126.com

    2015-04-10

    miR-373 was reported to be elevated in several tumors; however, the role of miR-373 in cervical cancer has not been investigated. In this study we aimed to investigate the role of miR-373 in tumorigenicity of cervical cancer cells in vivo and in vitro. The expression of miR-373 was investigated using real-time reverse transcription-polymerase chain reaction assay in 45 cervical specimens and cervical cancer cell lines. The role of miR-373 in tumorigenicity of cervical cancer cells was assessed by cell proliferation, colony formation in vitro as well as tumor growth assays in vivo with the overexpression of miR-373 or gene silencing. The functional target gene of miR-373 in cervical cancer cells was identified using integrated bioinformatics analysis, gene expression arrays, and luciferase assay. We founded that the expression of miR-373 is upregulated in human cervical cancer tissues and cervical carcinoma cell lines when compared to the corresponding noncancerous tissues. Ectopic overexpression of miR-373 in human cervical cancer cells promoted cell growth in vitro and tumorigenicity in vivo, whereas silencing the expression of miR-373 decreased the rate of cell growth. YOD1 was identified as a direct and functional target of miR-373 in cervical cancer cells. Expression levels of miR-373 were inversely correlated with YOD1 levels in human cervical cancer tissues. RNAi-mediated knockdown of YOD1 phenocopied the proliferation-promoting effect of miR-373. Moreover, overexpression of YOD1 abrogated miR-373-induced proliferation of cervical cancer cells. These results demonstrate that miR-373 increases proliferation by directly targeting YOD1, a new potential therapeutic target in cervical cancer. - Highlights: • The expression of miR-373 is upregulated in human cervical cancer tissues. • miR-373 effects as oncogenic miRNA in cervical cancer in vitro and in vivo. • miR-373 increases proliferation of cervical cancer cells by directly targeting YOD1.

  3. MicroRNA mediated network motifs in autoimmune diseases and its crosstalk between genes, functions and pathways.

    Science.gov (United States)

    Prabahar, Archana; Natarajan, Jeyakumar

    2017-01-01

    Autoimmune diseases (AIDs) are incurable but suppressible diseases whose molecular mechanisms are yet to be elucidated. In this work, we selected five systemic autoimmune diseases such as Rheumatoid Arthritis (RA), Type 1 Diabetes (T1D), Inflammatory Bowel Disease (IBD), Autoimmune Thyroid Disease (ATD) and Systemic Lupus Erythematosus (SLE). Heterogeneous data such as miRNA, transcription factor (TF), target genes and protein-protein interactions involved in these AIDs were integrated to understand their roles at different functional levels of miRNA such as transcription initiation, gene regulatory network formation and post transcriptional regulation. To understand the functional characteristics of these complex biological networks, they can be simplified as network motifs (sub networks) and motif-motif interacting pairs (MMIs). The network motif patterns and motif-motif interacting pairs that occur for the selected five diseases were identified. To further understand the functional association between AIDs, functions and pathways were determined using gene set enrichment analysis and five selected immune signaling pathways (ISPs). The crosstalk within AIDs and between the immune signaling pathways (ISPs) could provide novel insights in deciphering disease mechanisms. This study represents the first investigation of miRNA-TF regulatory network for AIDs and its association with ISPs using sub-network motifs. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Risk factors for delayed inpatient functional recovery after total knee arthroplasty

    NARCIS (Netherlands)

    Hoogeboom, T.J.; Meeteren, N.L. van; Schank, K.; Kim, R.H.; Miner, T.; Stevens-Lapsley, J.E.

    2015-01-01

    Purpose. To determine the predictive value of surgery-related variables for delayed inpatient functional recovery (i.e., >/=3 days to reach functional independence) after TKA. Method. 193 consecutive people undergoing TKA were included in this prospective cohort study. Inpatient functional recovery

  5. Risk factors for delayed inpatient functional recovery after total knee arthroplasty

    NARCIS (Netherlands)

    Hoogeboom, T.J.; Meeteren, N.L.U. van; Schank, K.; Kim, R.H.; Miner, T.; Stevens-Lapsley, J.E.

    2015-01-01

    Purpose. To determine the predictive value of surgery-related variables for delayed inpatient functional recovery (i.e., ≥3 days to reach functional independence) after TKA. Method. 193 consecutive people undergoing TKA were included in this prospective cohort study. Inpatient functional recovery

  6. RNA recognition by a human antibody against brain cytoplasmic 200 RNA.

    Science.gov (United States)

    Jung, Euihan; Lee, Jungmin; Hong, Hyo Jeong; Park, Insoo; Lee, Younghoon

    2014-06-01

    Diverse functional RNAs participate in a wide range of cellular processes. The RNA structure is critical for function, either on its own or as a complex form with proteins and other ligands. Therefore, analysis of the RNA conformation in cells is essential for understanding their functional mechanisms. However, no appropriate methods have been established as yet. Here, we developed an efficient strategy for panning and affinity maturation of anti-RNA human monoclonal antibodies from a naïve antigen binding fragment (Fab) combinatorial phage library. Brain cytoplasmic 200 (BC200) RNA, which is also highly expressed in some tumors, was used as an RNA antigen. We identified MabBC200-A3 as the optimal binding antibody. Mutagenesis and SELEX experiments showed that the antibody recognized a domain of BC200 in a structure- and sequence-dependent manner. Various breast cancer cell lines were further examined for BC200 RNA expression using conventional hybridization and immunoanalysis with MabBC200-A3 to see whether the antibody specifically recognizes BC200 RNA among the total purified RNAs. The amounts of antibody-recognizable BC200 RNA were consistent with hybridization signals among the cell lines. Furthermore, the antibody was able to discriminate BC200 RNA from other RNAs, supporting the utility of this antibody as a specific RNA structure-recognizing probe. Intriguingly, however, when permeabilized cells were subjected to immunoanalysis instead of purified total RNA, the amount of antibody-recognizable RNA was not correlated with the cellular level of BC200 RNA, indicating that BC200 RNA exists as two distinct forms (antibody-recognizable and nonrecognizable) in breast cancer cells and that their distribution depends on the cell type. Our results clearly demonstrate that anti-RNA antibodies provide an effective novel tool for detecting and analyzing RNA conformation. © 2014 Jung et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  7. Mobile-bearing total ankle arthroplasty : a fundamental assessment of the clinical, radiographic and functional outcomes

    NARCIS (Netherlands)

    Doets, Hendrik Cornelis

    2009-01-01

    Ankle arthritis often leads to significant impairments for the patient. As total ankle arthroplasty (TAA) with use of fixed-bearing (2-component) total ankle prostheses has a high rate of early failures, fusion of the ankle joint is, until today, considered to be the standard surgical treatment for

  8. Integrated genome-wide analysis of transcription factor occupancy, RNA polymerase II binding and steady-state RNA levels identify differentially regulated functional gene classes

    NARCIS (Netherlands)

    Mokry, M.; Hatzis, P.; Schuijers, J.; Lansu, N.; Ruzius, F.P.; Clevers, H.; Cuppen, E.

    2012-01-01

    Routine methods for assaying steady-state mRNA levels such as RNA-seq and micro-arrays are commonly used as readouts to study the role of transcription factors (TFs) in gene expression regulation. However, cellular RNA levels do not solely depend on activity of TFs and subsequent transcription by

  9. A Convenient In Vivo Model Using Small Interfering RNA Silencing to Rapidly Assess Skeletal Gene Function.

    Directory of Open Access Journals (Sweden)

    Wen Zhang

    Full Text Available It is difficult to study bone in vitro because it contains various cell types that engage in cross-talk. Bone biologically links various organs, and it has thus become increasingly evident that skeletal physiology must be studied in an integrative manner in an intact animal. We developed a model using local intraosseous small interfering RNA (siRNA injection to rapidly assess the effects of a target gene on the local skeletal environment. In this model, 160-g male Sprague-Dawley rats were treated for 1-2 weeks. The left tibia received intraosseous injection of a parathyroid hormone 1 receptor (Pth1r or insulin-like growth factor 1 receptor (Igf-1r siRNA transfection complex loaded in poloxamer 407 hydrogel, and the right tibia received the same volume of control siRNA. All the tibias received an intraosseous injection of recombinant human parathyroid hormone (1-34 (rhPTH (1-34 or insulin-like growth factor-1 (IGF-1. Calcein green and alizarin red were injected 6 and 2 days before euthanasia, respectively. IGF-1R and PTH1R expression levels were detected via RT-PCR assays and immunohistochemistry. Bone mineral density (BMD, microstructure, mineral apposition rates (MARs, and strength were determined by dual-energy X-ray absorptiometry, micro-CT, histology and biomechanical tests. The RT-PCR and immunohistochemistry results revealed that IGF-1R and PTH1R expression levels were dramatically diminished in the siRNA-treated left tibias compared to the right tibias (both p<0.05. Using poloxamer 407 hydrogel as a controlled-release system prolonged the silencing effect of a single dose of siRNA; the mRNA expression levels of IGF-1R were lower at two weeks than at one week (p<0.01. The BMD, bone microstructure parameters, MAR and bone strength were significantly decreased in the left tibias compared to the right tibias (all p<0.05. This simple and convenient local intraosseous siRNA injection model achieved gene silencing with very small quantities of

  10. DDX5 and its associated lncRNA Rmrp modulate Th17 cell effector functions

    Science.gov (United States)

    Huang, Wendy; Thomas, Benjamin; Flynn, Ryan A.; Gavzy, Samuel J.; Wu, Lin; Kim, Sangwon V.; Hall, Jason A.; Miraldi, Emily R.; Ng, Charles P.; Rigo, Frank; Meadows, Sarah; Montoya, Nina R.; Herrera, Natalia G.; Domingos, Ana I.; Rastinejad, Fraydoon; Myers, Richard M.; Fuller-Pace, Frances V.; Bonneau, Richard; Chang, Howard Y.; Acuto, Oreste; Littman, Dan R.

    2016-01-01

    Th17 lymphocytes protect mucosal barriers from infections, but also contribute to multiple chronic inflammatory diseases. Their differentiation is controlled by RORγt, a ligand-regulated nuclear receptor. We identified the DEAD-box RNA helicase DDX5 as a RORγt partner that coordinates transcription of selective Th17 genes and is required for Th17-mediated inflammatory pathologies. Surprisingly, the ability of DDX5 to interact with RORγt and co-activate its targets depends on its intrinsic RNA helicase activity and binding of a conserved nuclear long noncoding RNA (lncRNA), Rmrp, which is mutated in Cartilage-Hair Hypoplasia (CHH) patients. A targeted Rmrp mutation in mice, corresponding to one in CHH patients, abrogated the lncRNA’s chromatin recruitment, ability to potentiate DDX5-RORγt interaction and RORγt target gene transcription. Elucidation of the link between Rmrp and the DDX5-RORγt complex reveals a role for RNA helicases and lncRNAs in tissue-specific transcriptional regulation and promises new opportunities for therapeutic intervention in Th17-dependent diseases. PMID:26675721

  11. The effects of MicroRNA transfections on global patterns of gene expression in ovarian cancer cells are functionally coordinated

    Directory of Open Access Journals (Sweden)

    Shahab Shubin W

    2012-08-01

    Full Text Available Abstract Background MicroRNAs (miRNAs are a class of small RNAs that have been linked to a number of diseases including cancer. The potential application of miRNAs in the diagnostics and therapeutics of ovarian and other cancers is an area of intense interest. A current challenge is the inability to accurately predict the functional consequences of exogenous modulations in the levels of potentially therapeutic miRNAs. Methods In an initial effort to systematically address this issue, we conducted miRNA transfection experiments using two miRNAs (miR-7, miR-128. We monitored the consequent changes in global patterns of gene expression by microarray and quantitative (real-time polymerase chain reaction. Network analysis of the expression data was used to predict the consequence of each transfection on cellular function and these predictions were experimentally tested. Results While ~20% of the changes in expression patterns of hundreds to thousands of genes could be attributed to direct miRNA-mRNA interactions, the majority of the changes are indirect, involving the downstream consequences of miRNA-mediated changes in regulatory gene expression. The changes in gene expression induced by individual miRNAs are functionally coordinated but distinct between the two miRNAs. MiR-7 transfection into ovarian cancer cells induces changes in cell adhesion and other developmental networks previously associated with epithelial-mesenchymal transitions (EMT and other processes linked with metastasis. In contrast, miR-128 transfection induces changes in cell cycle control and other processes commonly linked with cellular replication. Conclusions The functionally coordinated patterns of gene expression displayed by different families of miRNAs have the potential to provide clinicians with a strategy to treat cancers from a systems rather than a single gene perspective.

  12. Pseudoknot in domain II of 23 S rRNA is essential for ribosome function

    DEFF Research Database (Denmark)

    Rosendahl, G; Hansen, L H; Douthwaite, S

    1995-01-01

    The structure of domain II in all 23 S (and 23 S-like) rRNAs is constrained by a pseudoknot formed between nucleotides 1005 and 1138, and between 1006 and 1137 (Escherichia coli numbering). These nucleotides are exclusively conserved as 1005C.1138G and 1006C.1137G pairs in all Bacteria, Archaea...... increased accessibility in the rRNA structure close to the sites of the mutations. The degree to which the mutations increase rRNA accessibility correlates with the severity of their phenotypic effects. Nucleotide 1131G is extremely reactive to dimethyl sulphate modification in wild-type subunits...

  13. RNA Polymerase Structure, Function, Regulation, Dynamics, Fidelity, and Roles in GENE EXPRESSION | Center for Cancer Research

    Science.gov (United States)

    Multi-subunit RNA polymerases (RNAP) are ornate molecular machines that translocate on a DNA template as they generate a complementary RNA chain. RNAPs are highly conserved in evolution among eukarya, eubacteria, archaea, and some viruses. As such, multi-subunit RNAPs appear to be an irreplaceable advance in the evolution of complex life on earth. Because of their stepwise movement on DNA, RNAPs are considered to be molecular motors, and because RNAPs catalyze a templated polymerization reaction, they are central to biological information flow.

  14. IMP2 axonal localization, RNA interactome, and function in the development of axon trajectories

    DEFF Research Database (Denmark)

    Preitner, Nicolas; Quan, Jie; Li, Xinmin

    2016-01-01

    RNA-based regulatory mechanisms play important roles in the development and plasticity of neural circuits and neurological disease. Developing axons provide a model well suited to the study of RNA-based regulation, and contain specific subsets of mRNAsthat are locally translated and have roles...... to strong defects in commissural axon trajectories at the midline intermediate target. These results reveal a highly distinctive axonal enrichment of IMP2, show that it interacts with a network of axon guidance-related mRNAs, and reveal that it is required for normal axon pathfinding during vertebrate...

  15. Correction of Mutant p63 in EEC Syndrome Using siRNA Mediated Allele-Specific Silencing Restores Defective Stem Cell Function.

    Science.gov (United States)

    Barbaro, Vanessa; Nasti, Annamaria A; Del Vecchio, Claudia; Ferrari, Stefano; Migliorati, Angelo; Raffa, Paolo; Lariccia, Vincenzo; Nespeca, Patrizia; Biasolo, Mariangela; Willoughby, Colin E; Ponzin, Diego; Palù, Giorgio; Parolin, Cristina; Di Iorio, Enzo

    2016-06-01

    Ectrodactyly-Ectodermal dysplasia-Clefting (EEC) syndrome is a rare autosomal dominant disease caused by heterozygous mutations in the p63 gene and characterized by limb defects, orofacial clefting, ectodermal dysplasia, and ocular defects. Patients develop progressive total bilateral limbal stem cell deficiency, which eventually results in corneal blindness. Medical and surgical treatments are ineffective and of limited benefit. Oral mucosa epithelial stem cells (OMESCs) represent an alternative source of stem cells capable of regenerating the corneal epithelium and, combined with gene therapy, could provide an attractive therapeutic avenue. OMESCs from EEC patients carrying the most severe p63 mutations (p.R279H and p.R304Q) were characterized and the genetic defect of p.R279H silenced using allele-specific (AS) small interfering RNAs (siRNAs). Systematic screening of locked nucleic acid (LNA)-siRNAs against R279H-p63 allele in (i) stable WT-ΔNp63α-RFP and R279H-ΔNp63α-EGFP cell lines, (ii) transient doubly transfected cell lines, and (iii) p.R279H OMESCs, identified a number of potent siRNA inhibitors for the mutant allele, which had no effect on wild-type p63. In addition, siRNA treatment led to longer acquired life span of mutated stem cells compared to controls, less accelerated stem cell differentiation in vitro, reduced proliferation properties, and effective ability in correcting the epithelial hypoplasia, thus giving rise to full thickness stratified and differentiated epithelia. This study demonstrates the phenotypic correction of mutant stem cells (OMESCs) in EEC syndrome by means of siRNA mediated AS silencing with restoration of function. The application of siRNA, alone or in combination with cell-based therapies, offers a therapeutic strategy for corneal blindness in EEC syndrome. Stem Cells 2016;34:1588-1600. © 2016 AlphaMed Press.

  16. The decapping activator Edc3 and the Q/N-rich domain of Lsm4 function together to enhance mRNA stability and alter mRNA decay pathway dependence in Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Susanne Huch

    2016-10-01

    Full Text Available The rate and regulation of mRNA decay are major elements in the proper control of gene expression. Edc3 and Lsm4 are two decapping activator proteins that have previously been shown to function in the assembly of RNA granules termed P bodies. Here, we show that deletion of edc3, when combined with a removal of the glutamine/asparagine rich region of Lsm4 (edc3Δ lsm4ΔC reduces mRNA stability and alters pathways of mRNA degradation. Multiple tested mRNAs exhibited reduced stability in the edc3Δ lsm4ΔC mutant. The destabilization was linked to an increased dependence on Ccr4-mediated deadenylation and mRNA decapping. Unlike characterized mutations in decapping factors that either are neutral or are able to stabilize mRNA, the combined edc3Δ lsm4ΔC mutant reduced mRNA stability. We characterized the growth and activity of the major mRNA decay systems and translation in double mutant and wild-type yeast. In the edc3Δ lsm4ΔC mutant, we observed alterations in the levels of specific mRNA decay factors as well as nuclear accumulation of the catalytic subunit of the decapping enzyme Dcp2. Hence, we suggest that the effects on mRNA stability in the edc3Δ lsm4ΔC mutant may originate from mRNA decay protein abundance or changes in mRNPs, or alternatively may imply a role for P bodies in mRNA stabilization.

  17. Pancreatic Endocrine and Exocrine Function in Children following Near-Total Pancreatectomy for Diffuse Congenital Hyperinsulinism

    Science.gov (United States)

    Arya, Ved Bhushan; Senniappan, Senthil; Demirbilek, Huseyin; Alam, Syeda; Flanagan, Sarah E.; Ellard, Sian; Hussain, Khalid

    2014-01-01

    Context Congenital hyperinsulinism (CHI), the commonest cause of persistent hypoglycaemia, has two main histological subtypes: diffuse and focal. Diffuse CHI, if medically unresponsive, is managed with near-total pancreatectomy. Post-pancreatectomy, in addition to persistent hypoglycaemia, there is a very high risk of diabetes mellitus and pancreatic exocrine insufficiency. Setting International referral centre for the management of CHI. Patients Medically unresponsive diffuse CHI patients managed with near-total pancreatectomy between 1994 and 2012. Intervention Near-total pancreatectomy. Main Outcome Measures Persistent hypoglycaemia post near-total pancreatectomy, insulin-dependent diabetes mellitus, clinical and biochemical (faecal elastase 1) pancreatic exocrine insufficiency. Results Of more than 300 patients with CHI managed during this time period, 45 children had medically unresponsive diffuse disease and were managed with near-total pancreatectomy. After near-total pancreatectomy, 60% of children had persistent hypoglycaemia requiring medical interventions. The incidence of insulin dependent diabetes mellitus was 96% at 11 years after surgery. Thirty-two patients (72%) had biochemical evidence of severe pancreatic exocrine insufficiency (Faecal elastase 1pancreatectomy is very unsatisfactory. The incidence of persistent hypoglycaemia and insulin-dependent diabetes mellitus is very high. The presence of clinical rather than biochemical pancreatic exocrine insufficiency should inform decisions about pancreatic enzyme supplementation. PMID:24840042

  18. Data in support of transcriptional regulation and function of Fas-antisense long noncoding RNA during human erythropoiesis

    Directory of Open Access Journals (Sweden)

    Olga Villamizar

    2016-06-01

    Full Text Available This paper describes data related to a research article titled, “Fas-antisense long noncoding RNA is differentially expressed during maturation of human erythrocytes and confers resistance to Fas-mediated cell death” [1]. Long noncoding RNAs (lncRNAs are increasingly appreciated for their capacity to regulate many steps of gene expression. While recent studies suggest that many lncRNAs are functional, the scope of their actions throughout human biology is largely undefined including human red blood cell development (erythropoiesis. Here we include expression data for 82 lncRNAs during early, intermediate and late stages of human erythropoiesis using a commercial qPCR Array. From these data, we identified lncRNA Fas-antisense 1 (Fas-AS1 or Saf described in the research article. Also included are 5′ untranslated sequences (UTR for lncRNA Saf with transcription factor target sequences identified. Quantitative RT-PCR data demonstrate relative levels of critical erythroid transcription factors, GATA-1 and KLF1, in K562 human erythroleukemia cells and maturing erythroblasts derived from human CD34+ cells. End point and quantitative RT-PCR data for cDNA prepared using random hexamers versus oligo(dT18 revealed that lncRNA Saf is not effectively polyadenylated. Finally, we include flow cytometry histograms demonstrating Fas levels on maturing erythroblasts derived from human CD34+ cells transduced using mock conditions or with lentivirus particles encoding for Saf.

  19. Mutational analysis of vaccinia virus E3 protein: the biological functions do not correlate with its biochemical capacity to bind double-stranded RNA.

    Science.gov (United States)

    Dueck, Kevin J; Hu, YuanShen Sandy; Chen, Peter; Deschambault, Yvon; Lee, Jocelyn; Varga, Jessie; Cao, Jingxin

    2015-05-01

    Vaccinia E3 protein has the biochemical capacity of binding to double-stranded RNA (dsRNA). The best characterized biological functions of the E3 protein include its host range function, suppression of cytokine expression, and inhibition of interferon (IFN)-induced antiviral activity. Currently, the role of the dsRNA binding capacity in the biological functions of the E3 protein is not clear. To further understand the mechanism of the E3 protein biological functions, we performed alanine scanning of the entire dsRNA binding domain of the E3 protein to examine the link between its biochemical capacity of dsRNA binding and biological functions. Of the 115 mutants examined, 20 were defective in dsRNA binding. Although the majority of the mutants defective in dsRNA binding also showed defective replication in HeLa cells, nine mutants (I105A, Y125A, E138A, F148A, F159A, K171A, L182A, L183A, and I187/188A) retained the host range function to various degrees. Further examination of a set of representative E3L mutants showed that residues essential for dsRNA binding are not essential for the biological functions of E3 protein, such as inhibition of protein kinase R (PKR) activation, suppression of cytokine expression, and apoptosis. Thus, data described in this communication strongly indicate the E3 protein performs its biological functions via a novel mechanism which does not correlate with its dsRNA binding activity. dsRNAs produced during virus replication are important pathogen-associated molecular patterns (PAMPs) for inducing antiviral immune responses. One of the strategies used by many viruses to counteract such antiviral immune responses is achieved by producing dsRNA binding proteins, such as poxvirus E3 family proteins, influenza virus NS1, and Ebola virus V35 proteins. The most widely accepted model for the biological functions of this class of viral dsRNA binding proteins is that they bind to and sequester viral dsRNA PAMPs; thus, they suppress the related

  20. Effect of preoperative neuromuscular training (NEMEX-TJR) on functional outcome after total knee replacement

    DEFF Research Database (Denmark)

    Huber, Erika O; Roos, Ewa M.; Meichtry, André

    2015-01-01

    BACKGROUND: Improving functional status preoperatively through exercise may improve postoperative outcome. Previous knowledge on preoperative exercise in knee osteoarthritis is insufficient. The aim of the study was to compare the difference in change between groups in lower extremity function from...... sessions of KS or 3 sessions of KS alone. Function was assessed with the Chair Stand Test (CST, primary endpoint) and the Knee Injury and Osteoarthritis Outcome Score (KOOS) subscales focusing on daily living function (ADL) and pain (secondary endpoints). Assessments were performed immediately before...... pain the treatment effect was -4.9 points (-16.3, 6.5) and -3.3 points (-13.5, 6.8) respectively. CONCLUSIONS: A median (IQR) of 10 (8, 14) exercise sessions before surgery showed an additional small but non-significant improvement in all functional assessments compared to patient education alone...

  1. Pseudoknot in domain II of 23 S rRNA is essential for ribosome function

    DEFF Research Database (Denmark)

    Rosendahl, G; Hansen, L H; Douthwaite, S

    1995-01-01

    The structure of domain II in all 23 S (and 23 S-like) rRNAs is constrained by a pseudoknot formed between nucleotides 1005 and 1138, and between 1006 and 1137 (Escherichia coli numbering). These nucleotides are exclusively conserved as 1005C.1138G and 1006C.1137G pairs in all Bacteria, Archaea...... and chloroplasts, whereas 1005G.1138C and 1006U.1137A pairs occur in Eukarya. We have mutagenized nucleotides 1005C-->G, 1006C-->U, 1137G-->A and 1138G-->C, both individually and in combinations, in a 23 S rRNA gene from the bacterium E. coli. The ability of 23 S rRNA to support cell growth is reduced when either...... increased accessibility in the rRNA structure close to the sites of the mutations. The degree to which the mutations increase rRNA accessibility correlates with the severity of their phenotypic effects. Nucleotide 1131G is extremely reactive to dimethyl sulphate modification in wild-type subunits...

  2. Value of a simple technique for the measurement of total renal function and each kidney functions without requiring blood or urine samples

    International Nuclear Information System (INIS)

    Meyers, A.; Chachati, A.; Godon, J.P.; Rigo, P.

    1985-01-01

    The determination of renal uptake of 99m Tc DTPA and of 131 I Hippuran (as a percentage of the administred dose) at a time interval 1-3 minutes after tracer injection, is a rapid, accurate method for the measurement of total renal function and each kidney functions. Its clinical validity has been confirmed [fr

  3. Allele variants in functional MicroRNA target sites of the neurotrophin-3 receptor gene (NTRK3) as susceptibility factors for anxiety disorders

    OpenAIRE

    Muiños Gimeno, Margarita; Guidi, Mònica; Kagerbauer, Birgit; Martín Santos, Rocío; Navinés, Ricard; Alonso, Pino; Menchón, José M.; Gratacós Mayora, Mònica; Estivill, Xavier, 1955-; Espinosa Parrilla, Yolanda

    2009-01-01

    Genetic and functional data indicate that variation in the expression of the neurotrophin-3 receptor gene (NTRK3) may have an impact on neuronal plasticity, suggesting a role for NTRK3 in the pathophysiology of anxiety disorders. MicroRNA (miRNA) posttranscriptional gene regulators act by base-pairing to specific sequence sites, usually at the 3'UTR of the target mRNA. Variants at these sites might result in gene expression changes contributing to disease susceptibility. We investigated genet...

  4. MysiRNA-designer: a workflow for efficient siRNA design.

    Directory of Open Access Journals (Sweden)

    Mohamed Mysara

    Full Text Available The design of small interfering RNA (siRNA is a multi factorial problem that has gained the attention of many researchers in the area of therapeutic and functional genomics. MysiRNA score was previously introduced that improves the correlation of siRNA activity prediction considering state of the art algorithms. In this paper, a new program, MysiRNA-Designer, is described which integrates several factors in an automated work-flow considering mRNA transcripts variations, siRNA and mRNA target accessibility, and both near-perfect and partial off-target matches. It also features the MysiRNA score, a highly ranked correlated siRNA efficacy prediction score for ranking the designed siRNAs, in addition to top scoring models Biopredsi, DISR, Thermocomposition21 and i-Score, and integrates them in a unique siRNA score-filtration technique. This multi-score filtration layer filters siRNA that passes the 90% thresholds calculated from experimental dataset features. MysiRNA-Designer takes an accession, finds conserved regions among its transcript space, finds accessible regions within the mRNA, designs all possible siRNAs for these regions, filters them based on multi-scores thresholds, and then performs SNP and off-target filtration. These strict selection criteria were tested against human genes in which at least one active siRNA was designed from 95.7% of total genes. In addition, when tested against an experimental dataset, MysiRNA-Designer was found capable of rejecting 98% of the false positive siRNAs, showing superiority over three state of the art siRNA design programs. MysiRNA is a freely accessible (Microsoft Windows based desktop application that can be used to design siRNA with a high accuracy and specificity. We believe that MysiRNA-Designer has the potential to play an important role in this area.

  5. Total average diastolic longitudinal displacement by colour tissue doppler imaging as an assessment of diastolic function

    DEFF Research Database (Denmark)

    de Knegt, Martina Chantal; Biering-Sørensen, Tor; Søgaard, Peter

    2016-01-01

    BACKGROUND: The current method for a non-invasive assessment of diastolic dysfunction is complex with the use of algorithms of many different echocardiographic parameters. Total average diastolic longitudinal displacement (LD), determined by colour tissue Doppler imaging (TDI) via the measurement...

  6. Functional capacity and actual daily activity do not contribute to patient satisfaction after total knee arthroplasty

    NARCIS (Netherlands)

    M.M. Vissers (Maaike); I.B. de Groot (Ingrid); M. Reijman (Max); J.B.J. Bussmann (Hans); H.J. Stam (Henk); J.A.N. Verhaar (Jan)

    2010-01-01

    textabstractAbstract Background: After total knee arthroplasty (TKA) only 75-89% of patients are satisfied. Because patient satisfaction is a prime goal of all orthopaedic procedures, optimization of patient satisfaction is of major importance. Factors related to patient satisfaction after TKA

  7. Risk Factors for Delayed Inpatient Functional Recovery after Total Knee Arthroplasty

    Directory of Open Access Journals (Sweden)

    Thomas J. Hoogeboom

    2015-01-01

    Full Text Available Purpose. To determine the predictive value of surgery-related variables for delayed inpatient functional recovery (i.e., ≥3 days to reach functional independence after TKA. Method. 193 consecutive people undergoing TKA were included in this prospective cohort study. Inpatient functional recovery was measured daily using the Iowa Level of Assistance scale (ILAS. Two persons reviewed medical records to extract patient characteristics (i.e., age, sex, and BMI and surgical factors (i.e., blood loss, tourniquet time, postoperative morphine use, and surgical experience. Odds ratios (OR and area under the curves (AUC were calculated to determine the predictive value of the putative factors and of the model on delayed functional recovery, respectively. Results. Delayed functional recovery was apparent in 76 (39% people. Higher age, female sex, and higher BMI were all independent risk factors for delayed functional recovery (AUC (95%-CI; 0.72 (0.65–0.80, whereas blood loss (OR (95%-CI; 1.00 (0.99–1.01, tourniquet time (OR = 1.00 (0.98–1.02, and postoperative morphine use (OR = 0.88 (0.37–2.06 did not statistically improve the predictive value of the model, while surgical experience did (OR = 0.31 (0.16–0.64; AUC = 0.76 (0.69–83. Conclusions. Surgery-related factors contribute little to the patient-related characteristics in a predictive model explaining delayed functional recovery after TKA in daily orthopaedic practice.

  8. Development of functional genomic tools in trematodes: RNA interference and luciferase reporter gene activity in Fasciola hepatica.

    Directory of Open Access Journals (Sweden)

    Gabriel Rinaldi

    2008-07-01

    Full Text Available The growing availability of sequence information from diverse parasites through genomic and transcriptomic projects offer new opportunities for the identification of key mediators in the parasite-host interaction. Functional genomics approaches and methods for the manipulation of genes are essential tools for deciphering the roles of genes and to identify new intervention targets in parasites. Exciting advances in functional genomics for parasitic helminths are starting to occur, with transgene expression and RNA interference (RNAi reported in several species of nematodes, but the area is still in its infancy in flatworms, with reports in just three species. While advancing in model organisms, there is a need to rapidly extend these technologies to other parasites responsible for several chronic diseases of humans and cattle. In order to extend these approaches to less well studied parasitic worms, we developed a test method for the presence of a viable RNAi pathway by silencing the exogenous reporter gene, firefly luciferase (fLUC. We established the method in the human blood fluke Schistosoma mansoni and then confirmed its utility in the liver fluke Fasciola hepatica. We transformed newly excysted juveniles of F. hepatica by electroporation with mRNA of fLUC and three hours later were able to detect luciferase enzyme activity, concentrated mainly in the digestive ceca. Subsequently, we tested the presence of an active RNAi pathway in F. hepatica by knocking down the exogenous luciferase activity by introduction into the transformed parasites of double-stranded RNA (dsRNA specific for fLUC. In addition, we tested the RNAi pathway targeting an endogenous F. hepatica gene encoding leucine aminopeptidase (FhLAP, and observed a significant reduction in specific mRNA levels. In summary, these studies demonstrated the utility of RNAi targeting reporter fLUC as a reporter gene assay to establish the presence of an intact RNAi pathway in helminth

  9. Development of Functional Genomic Tools in Trematodes: RNA Interference and Luciferase Reporter Gene Activity in Fasciola hepatica

    Science.gov (United States)

    Rinaldi, Gabriel; Morales, Maria E.; Cancela, Martín; Castillo, Estela; Brindley, Paul J.; Tort, José F.

    2008-01-01

    The growing availability of sequence information from diverse parasites through genomic and transcriptomic projects offer new opportunities for the identification of key mediators in the parasite–host interaction. Functional genomics approaches and methods for the manipulation of genes are essential tools for deciphering the roles of genes and to identify new intervention targets in parasites. Exciting advances in functional genomics for parasitic helminths are starting to occur, with transgene expression and RNA interference (RNAi) reported in several species of nematodes, but the area is still in its infancy in flatworms, with reports in just three species. While advancing in model organisms, there is a need to rapidly extend these technologies to other parasites responsible for several chronic diseases of humans and cattle. In order to extend these approaches to less well studied parasitic worms, we developed a test method for the presence of a viable RNAi pathway by silencing the exogenous reporter gene, firefly luciferase (fLUC). We established the method in the human blood fluke Schistosoma mansoni and then confirmed its utility in the liver fluke Fasciola hepatica. We transformed newly excysted juveniles of F. hepatica by electroporation with mRNA of fLUC and three hours later were able to detect luciferase enzyme activity, concentrated mainly in the digestive ceca. Subsequently, we tested the presence of an active RNAi pathway in F. hepatica by knocking down the exogenous luciferase activity by introduction into the transformed parasites of double-stranded RNA (dsRNA) specific for fLUC. In addition, we tested the RNAi pathway targeting an endogenous F. hepatica gene encoding leucine aminopeptidase (FhLAP), and observed a significant reduction in specific mRNA levels. In summary, these studies demonstrated the utility of RNAi targeting reporter fLUC as a reporter gene assay to establish the presence of an intact RNAi pathway in helminth parasites. These

  10. Delivery of siRNA silencing P-gp in peptide-functionalized nanoparticles causes efflux modulation at the blood-brain barrier

    DEFF Research Database (Denmark)

    Gomes, Maria João; Kennedy, Patrick J; Martins, Susana

    2017-01-01

    AIM: Explore the use of transferrin-receptor peptide-functionalized nanoparticles (NPs) targeting blood-brain barrier (BBB) as siRNA carriers to silence P-glycoprotein (P-gp). MATERIALS & METHODS: Permeability experiments were assessed through a developed BBB cell-based model; P-gp mRNA expressio...

  11. OPTIMAL IMPROVEMENT IN FUNCTION AFTER TOTAL HIP AND KNEE REPLACEMENT: HOW DEEP DO YOU KNOW YOUR PATIENT’S MIND?

    Science.gov (United States)

    De Caro, M F; Vicenti, G; Abate, A; Picca, G; Leoncini, V; Lomuscio, M; Casalino, A; Solarino, G; Moretti, B

    2015-01-01

    Osteoarthritis (OA) of the hip and knee causes pain and loss of joint mobility, leading to limitations in physical function. When conservative treatment fails total hip and knee replacement is a cost-effective surgical option. Patients have high expectations regarding functional outcome after these procedures. If such expectations are not met, they may still be dissatisfied with the outcome of a technically successful procedure. Recently, numerous studies reported that psychological factors can influence the outcome of total knee replacement (tkr) and total hip artrhoplasty with total hip replacement (thr). We conducted a prospective study on a consecutive sample of 280 patients affected by hip or knee OA who underwent total joint replacement. At patients’ admission, Harris Hip Score (HHS) and Knee Society Score (KSS) were used to assess pain and function. Furthermore, SF-36, Mini-Mental Status Examination (MMSE), Symptom Checklist-90-R (SCL-90-R), Coping Orientation to Problems Experienced (BRIEF-COPE) and the Amsterdam Preoperative Anxiety and Information Scale (APAIS) were administered. Patients had clinical and radio graphical follow up at 1, 3 and 6 months post-operatively. The HHS and KSS values before surgery showed a linear correlation with both SCL-90-R and MMSE. None of the investigated variables influenced post-operative HHS and KSS scores; however, the improvement of functional scores resulted conditioned by SCL-90-R values, VAS score, schooling and MMSE. Psychological factors and mental status in primary total hip and knee replacement can affect outcome and patient satisfaction. Strategies focused on identification and facing of these conditions must be considered to improve outcome of total replacement.

  12. Increasing Total Serum IgE, Allergic Bronchopulmonary Aspergillosis, and Lung Function in Cystic Fibrosis.

    Science.gov (United States)

    Gothe, Florian; Kappler, Matthias; Griese, Matthias

    Allergic bronchopulmonary aspergillosis (ABPA) is a hypersensitivity disorder contributing to lung disease in cystic fibrosis (CF) and challenging to diagnose. This study analyzed the predictive value of increasing total IgE (t-IgE) levels in a CF cohort alongside with clinical and serologic data. A total of 387 children and young adults were followed from 2000 to 2006 and retrospectively classified into 6 groups. Patients with t-IgE levels IgE were classified as "Naïve," those with Af-specific IgE (Af-sIgE) as "Sensitized." Patients with elevated t-IgE at entrance and Af-sIgE were labeled "Former ABPA," and those without, as "High t-IgE." Patients whose t-IgE values started normal and exceeded the 95th percentile during the study were labeled either "ABPA at risk" if Af-sIgE-positive or "Rising t-IgE" if not. Courses of t-IgE over time were divided into episodes with increasing IgE (ΔIgE) and related to pulmonary outcome. A total of 125 patients were classified Naïve (32%), 64 Sensitized (17%), 49 ABPA at risk (13%), 32 Rising t-IgE (8%), 102 Former ABPA (26%), and 15 High t-IgE (4%). A total of 874 ΔIgE episodes were accompanied by forced expiratory volume in 1 second (FEV 1 ) declines (r = -0.21, P IgE levels at least doubled within 3 months and exceeded the 95th age-specific percentile (P IgE levels may be helpful in diagnosing treatment requiring ABPA and predicts the effect of systemic steroid treatment on pulmonary outcome. Copyright © 2017 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  13. Effect of halophilic conditions in stabilisation of RNA structure and function at high temperature under radiations.

    Science.gov (United States)

    Maurel, M.-C.

    We have already shown the structural integrity of tRNA at high temperature - 82C for 30h - in high salt concentrations (Tehei et al, 2002). Stability were also performed by measuring the residual specific tRNA charge capacity after heat treatment for 30 h at 82C. RNA molecules are selected (in vitro selection) at high temperature at high salt concentration. We are undergoing studies of such molecules submitted to several stressful conditions, in particular high radiations. These studies provide support for the importance of salt to protect macromolecules against severe cosmic conditions. These could be useful for searching traces of life in planetary objects and space exploration. References : ElAmri, C., Baron, M-H., Maurel, M.-C. ``Adenine adsorption onto and release from meteorite specimens assessed by Surface Enhanced Raman Spectroscopy ''. Journal of Raman Spectroscopy (2004) in press. Meli, M., Vergne, J. and Maurel, M-C. "In vitro selection of adenine-dependent hairpin ribozymes" J. Biol. Chem., (2003), 278, 11, 9835-9842. ElAmri, C., Baron, M-H., Maurel, M.-C. ``Adenine in mineral samples : development of a methodology based on Surface Enhanced Raman Spectroscopy (SERS) for picomole detections ''. Spectrochimica Acta, A, 59, 2645-2654. Tehei, M., Franzetti, B., Maurel, M-C., Vergne, J., Hountondji, C. , Zaccai, G. ``Salt and the Search for Traces of Life '', Extremophiles, (2002), 6 : 427-430. Meli, M., Vergne, J., Décout, J.L., and Maurel, M-C. ``Adenine-Aptamer Complexes. A bipartite RNA site which binds the adenine nucleic base '', J. Biol. Chem., (2002), 277, 3, 2104-2111.

  14. Microarray Meta-Analysis of RNA-Binding Protein Functions in Alternative Polyadenylation

    Science.gov (United States)

    Hu, Wenchao; Liu, Yuting; Yan, Jun

    2014-01-01

    Alternative polyadenylation (APA) is a post-transcriptional mechanism to generate diverse mRNA transcripts with different 3′UTRs from the same gene. In this study, we systematically searched for the APA events with differential expression in public mouse microarray data. Hundreds of genes with over-represented differential APA events and the corresponding experiments were identified. We further revealed that global APA differential expression occurred prevalently in tissues such as brain comparing to peripheral tissues, and biological processes such as development, differentiation and immune responses. Interestingly, we also observed widespread differential APA events in RNA-binding protein (RBP) genes such as Rbm3, Eif4e2 and Elavl1. Given the fact that RBPs are considered as the main regulators of differential APA expression, we constructed a co-expression network between APAs and RBPs using the microarray data. Further incorporation of CLIP-seq data of selected RBPs showed that Nova2 represses and Mbnl1 promotes the polyadenylation of closest poly(A) sites respectively. Altogether, our study is the first microarray meta-analysis in a mammal on the regulation of APA by RBPs that integrated massive mRNA expression data under a wide-range of biological conditions. Finally, we present our results as a comprehensive resource in an online website for the research community. PMID:24622240

  15. Molecular Properties, Functional Mechanisms, and Applications of Sliced siRNA

    Directory of Open Access Journals (Sweden)

    Guihua Sun

    2015-01-01

    Full Text Available Using pre-miR-451 as a model molecule, we have characterized the general molecular properties of small hairpin RNAs that are processed into potent small interfering RNAs (siRNA by Argonaute2 (Ago2. The Ago2-sliced siRNAs (sli-siRNAs have the same silencing potency as the classical Dicer diced siRNAs (di-siRNAs but have dramatically reduced unwanted sense strand activities. We have built vectors with the constitutive or inducible U6 promoter that can express sli-siRNAs in mammalian cells, in which the sli-siRNAs can be correctly processed to repress target genes. As a proof of principle for potential applications of sli-siRNAs in vivo, we show that the expression of one Ago2 shRNA-1148 in HCT-116 colon cancer cells knocked down RRM2 expression and reduced the proliferation and invasiveness of the cells. The defined sli-siRNA model molecules and the expression systems established in this study will facilitate the design and application of sli-siRNAs as novel potent RNAi triggers with reduced off-target effects.

  16. Identification of functional sequences in the pregenomic RNA promoter of the Banana streak virus Cavendish strain (BSV-Cav).

    Science.gov (United States)

    Remans, Tony; Grof, Christopher P L; Ebert, Paul R; Schenk, Peer M

    2005-03-01

    The promoter regions of plant pararetroviruses direct transcription of the full-length viral genome into a pregenomic RNA that is an intermediate in the replication of the virus. It serves as template for reverse transcription and as polycistronic mRNA for translation to viral proteins. We have identified functional promoter elements in the intergenic region of the Cavendish isolate of Banana streak virus (BSV-Cav), a member of the genus Badnavirus. Potential binding sites for plant transcription factors were found both upstream and downstream of the transcription start site by homology search in the PLACE database of plant cis-acting elements. The functionality of these putative cis-acting elements was tested by constructing loss-of-function and "regain"-of-function mutant promoters whose activity was quantified in embryogenic sugarcane suspension cells. Four regions that are important for activity of the BSV-Cav promoter were identified: the region containing an as-1-like element, the region around -141 and down to -77, containing several putative transcription factor binding sites, the region including the CAAT-box, and the leader region. The results could help explain the high BSV-Cav promoter activity that was observed previously in transgenic sugarcane plants and give more insight into the plant cell-mediated replication of the viral genome in banana streak disease.

  17. Towards Total Quality Management in Universities: Quality Function Deployment Paradigm and Beyond

    Science.gov (United States)

    Al-Fuqaha, Isam Najib

    2014-01-01

    This paper is an endeavor to develop a customised and computerized matrix of Quality Function Deployment paradigm (QFD) that has been applied in industry, with the aim of probing quality assurance and enhancement in Universities. Results of testing the new matrix proved that, it is efficient and time-saving while compared with a detailed field…

  18. Component alignment and functional outcome following computer assisted and jig based total knee arthroplasty

    Directory of Open Access Journals (Sweden)

    Dnyanesh G Lad

    2013-01-01

    Conclusions: A significantly improved placement of the component was found in the coronal and sagittal planes of the tibial component by CAS. The placement of the components in the other planes was comparable with the values recorded in the jig-based surgery group. Functional outcome was not significantly different.

  19. The Effects of Total Sleep Deprivation and Recovery Sleep on Cognitive Performance and Brain Function

    National Research Council Canada - National Science Library

    Gillin, J

    2003-01-01

    ..... Even less is known about the cerebral effects of recovery sleep. The objective of this study is to investigate the effects of 2 full nights of sleep loss and 2 full nights of recovery sleep on cognitive performance and brain function...

  20. Multi-Product Total Cost Functions for Higher Education: The Case of Chinese Research Universities

    Science.gov (United States)

    Longlong, Hou; Fengliang, Li; Weifang, Min

    2009-01-01

    This paper empirically investigates the economies of scale and economies of scope for the Chinese research universities by employing the flexible fixed cost quadratic (FFCQ) function. The empirical results show that both economies of scale and economies of scope exist in the Chinese higher education system and support the common belief of…

  1. The Effects of Total Sleep Deprivation and Recovery Sleep on Cognitive Performance and Brain Function

    National Research Council Canada - National Science Library

    Drummond, Sean P

    2004-01-01

    .... Even less is known about the cerebral effects of recovery sleep. The overarching objective of this study is to investigate the effects of 2 full nights of sleep loss and 2 full nights of recovery sleep on cognitive performance and brain function...

  2. Manipulations with Multiple Choice Tests: A Probability Function of a Total Testscore

    Science.gov (United States)

    Dyck, Walter; Plancke-Schuyten, Gilberte

    1976-01-01

    Previous knowledge of the difficulty index and the intercorrelations of the items will allow group results to be predicted and manipulated. A compound bionomial probability function of a testscore is established for which a computer program has been written. Three item selections and the appropriate probability distributions are given which give…

  3. B Cell Receptor Activation Predominantly Regulates AKT-mTORC1/2 Substrates Functionally Related to RNA Processing.

    Directory of Open Access Journals (Sweden)

    Dara K Mohammad

    Full Text Available Protein kinase B (AKT phosphorylates numerous substrates on the consensus motif RXRXXpS/T, a docking site for 14-3-3 interactions. To identify novel AKT-induced phosphorylation events following B cell receptor (BCR activation, we performed proteomics, biochemical and bioinformatics analyses. Phosphorylated consensus motif-specific antibody enrichment, followed by tandem mass spectrometry, identified 446 proteins, containing 186 novel phosphorylation events. Moreover, we found 85 proteins with up regulated phosphorylation, while in 277 it was down regulated following stimulation. Up regulation was mainly in proteins involved in ribosomal and translational regulation, DNA binding and transcription regulation. Conversely, down regulation was preferentially in RNA binding, mRNA splicing and mRNP export proteins. Immunoblotting of two identified RNA regulatory proteins, RBM25 and MEF-2D, confirmed the proteomics data. Consistent with these findings, the AKT-inhibitor (MK-2206 dramatically reduced, while the mTORC-inhibitor PP242 totally blocked phosphorylation on the RXRXXpS/T motif. This demonstrates that this motif, previously suggested as an AKT target sequence, also is a substrate for mTORC1/2. Proteins with PDZ, PH and/or SH3 domains contained the consensus motif, whereas in those with an HMG-box, H15 domains and/or NF-X1-zinc-fingers, the motif was absent. Proteins carrying the consensus motif were found in all eukaryotic clades indicating that they regulate a phylogenetically conserved set of proteins.

  4. Probing Amorphous Components in High Temperature TE Materials by in situ Total Scattering and the Pair Distribution Function (PDF) Method

    DEFF Research Database (Denmark)

    Reardon, Hazel; Iversen, Bo Brummerstedt; Blichfeld, Anders Bank

    .e., by measuring both the Bragg and diffuse scattering from a sample. This method has rarely been exploited by the non-oxide thermoelectrics community. , , Treating total scattering data by the Pair Distribution Function method is a logical approach to understanding defects, disorder and amorphous components...

  5. Effects of preoperative neuromuscular electrical stimulation on quadriceps strength and functional recovery in total knee arthroplasty. A pilot study.

    LENUS (Irish Health Repository)

    Walls, Raymond J

    2010-01-01

    Supervised preoperative muscle strengthening programmes (prehabilitation) can improve recovery after total joint arthroplasty but are considered resource intensive. Neuromuscular electrical stimulation (NMES) has been shown to improve quadriceps femoris muscle (QFM) strength and clinical function in subjects with knee osteoarthritis (OA) however it has not been previously investigated as a prehabilitation modality.

  6. Preoperative prediction of inpatient recovery of function after total hip arthroplasty using performance-based tests: a prospective cohort study

    NARCIS (Netherlands)

    Oosting, E.; Hoogeboom, T.J.; Appelman-de Vries, S.A.; Swets, A.; Dronkers, J.J.; Meeteren, N.L. van

    2016-01-01

    PURPOSE: The aim of this study was to evaluate the value of conventional factors, the Risk Assessment and Predictor Tool (RAPT) and performance-based functional tests as predictors of delayed recovery after total hip arthroplasty (THA). METHOD: A prospective cohort study in a regional hospital in

  7. The efficacy of tourniquet assisted total knee arthroplasty on patient-reported and performance-based physical function

    DEFF Research Database (Denmark)

    Lohmann-Jensen, Rasmus; Holsgaard-Larsen, Anders; Emmeluth, Claus

    2014-01-01

    BACKGROUND: Surgical treatment of osteoarthritis with total knee arthroplasty (TKA) usually takes place in a complete bloodless field using a tourniquet. However, doing the surgery without a tourniquet may reduce muscle damage, post-surgery pain and led to improved functional rehabilitation and m...

  8. A Banach algebra of functions of several variables of finite total variation and Lipschitzian superposition operators. I

    NARCIS (Netherlands)

    Chistyakov, VV

    2005-01-01

    It is shown that the space of functions of n real variables with finite total variation in the sense of Vitali, Hardy and Krause, defined on a rectangle I-a(b) C R-n, is a Banach algebra under the pointwise operations and Hildebrandt-Leonov's norm. This result generalizes the classical case of

  9. A Banach algebra of functions of several variables of finite total variation and Lipschitzian superposition operators. II

    NARCIS (Netherlands)

    Chistyakov, VV

    2005-01-01

    We characterize superposition Nemytskii operators, which map the Banach algebra of functions of n real variables with finite total variation in the sense of Vitali, Hardy and Krause into itself and satisfy the global Lipschitz condition. Our results extend previous results in this direction by

  10. The Human dsRNA binding protein PACT is unable to functionally substitute for the Drosophila dsRNA binding protein R2D2 [v1; ref status: indexed, http://f1000r.es/201

    Directory of Open Access Journals (Sweden)

    Benjamin K Dickerman

    2013-10-01

    Full Text Available The primary function of the dsRNA binding protein (dsRBP PACT/RAX is to activate the dsRNA dependent protein kinase PKR in response to stress signals.  Additionally, it has been identified as a component of the small RNA processing pathway.  A role for PACT/RAX in this pathway represents an important interplay between two modes of post-transcriptional gene regulation.  The function of PACT/RAX in this context is poorly understood.  Thus, additional models are required to clarify the mechanism by which PACT/RAX functions.  In this study, Drosophila melanogaster was employed to identify functionally orthologous dsRNA-binding proteins.  Transgenic Drosophila expressing human PACT were generated to determine whether PACT is capable of functionally substituting for the Drosophila dsRBP R2D2, which has a well-defined role in small RNA biogenesis.  Results presented here indicate that PACT is unable to substitute for R2D2 at the whole organism level.

  11. Topical Anti-Nuclear Factor-Kappa B Small Interfering RNA with Functional Peptides Containing Sericin-Based Hydrogel for Atopic Dermatitis

    Directory of Open Access Journals (Sweden)

    Takanori Kanazawa

    2015-09-01

    Full Text Available The small interfering RNA (siRNA is suggested to offer a novel means of treating atopic dermatitis (AD because it allows the specific silencing of genes related to AD pathogenesis. In our previous study, we found that siRNA targeted against RelA, an important nuclear factor-kappa B (NF-κB subdomain, with functional peptides, showed therapeutic effects in a mouse model of AD. In the present study, to develop a topical skin application against AD, we prepared a hydrogel containing anti-RelA siRNA and functional peptides and determined the intradermal permeation and the anti-AD effects in an AD mouse model. We selected the silk protein, sericin (SC, which is a versatile biocompatible biomaterial to prepare hydrogel as an aqueous gel base. We found that the siRNA was more widely delivered to the site of application in AD-induced ear skin of mice after topical application via the hydrogel containing functional peptides than via the preparation without functional peptides. In addition, the ear thickness and clinical skin severity of the AD-induced mice treated with hydrogel containing anti-RelA siRNA with functional peptides improved more than that of mice treated with the preparation formulated with negative siRNA.

  12. Total and Segmental Colon Transit Time Study in Functional Constipation: Comparison With Healthy Subjects

    OpenAIRE

    Bhate, Prasad A.; Patel, Jatin A.; Parikh, Pathik; Ingle, Meghraj A.; Phadke, Anniruddha; Sawant, Prabha D.

    2015-01-01

    Background Constipation is a common problem worldwide. Constipation can be primary or secondary. Primary constipation is subdivided in slow transit constipation, normal transit constipation, and dyssynergic defecation. Colon transit time (CTT) is the most basic and primary tool in evaluating disorders of colonic motility. CTT helps to differentiate between types of constipation and plan the treatment. Methods Fifty functional constipation patients and 25 healthy controls were asked to ingest ...

  13. Functional and Structural Analysis of a Highly-Expressed Yersinia pestis Small RNA following Infection of Cultured Macrophages.

    Directory of Open Access Journals (Sweden)

    Nan Li

    Full Text Available Non-coding small RNAs (sRNAs are found in practically all bacterial genomes and play important roles in regulating gene expression to impact bacterial metabolism, growth, and virulence. We performed transcriptomics analysis to identify sRNAs that are differentially expressed in Yersinia pestis that invaded the human macrophage cell line THP-1, compared to pathogens that remained extracellular in the presence of host. Using ultra high-throughput sequencing, we identified 37 novel and 143 previously known sRNAs in Y. pestis. In particular, the sRNA Ysr170 was highly expressed in intracellular Yersinia and exhibited a log2 fold change ~3.6 higher levels compared to extracellular bacteria. We found that knock-down of Ysr170 expression attenuated infection efficiency in cell culture and growth rate in response to different stressors. In addition, we applied selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE analysis to determine the secondary structure of Ysr170 and observed structural changes resulting from interactions with the aminoglycoside antibiotic gentamycin and the RNA chaperone Hfq. Interestingly, gentamicin stabilized helix 4 of Ysr170, which structurally resembles the native gentamicin 16S ribosomal binding site. Finally, we modeled the tertiary structure of Ysr170 binding to gentamycin using RNA motif modeling. Integration of these experimental and structural methods can provide further insight into the design of small molecules that can inhibit function of sRNAs required for pathogen virulence.

  14. Acquisition of functions on the outer capsid surface during evolution of double-stranded RNA fungal viruses.

    Science.gov (United States)

    Mata, Carlos P; Luque, Daniel; Gómez-Blanco, Josué; Rodríguez, Javier M; González, José M; Suzuki, Nobuhiro; Ghabrial, Said A; Carrascosa, José L; Trus, Benes L; Castón, José R

    2017-12-01

    Unlike their counterparts in bacterial and higher eukaryotic hosts, most fungal viruses are transmitted intracellularly and lack an extracellular phase. Here we determined the cryo-EM structure at 3.7 Å resolution of Rosellinia necatrix quadrivirus 1 (RnQV1), a fungal double-stranded (ds)RNA virus. RnQV1, the type species of the family Quadriviridae, has a multipartite genome consisting of four monocistronic segments. Whereas most dsRNA virus capsids are based on dimers of a single protein, the ~450-Å-diameter, T = 1 RnQV1 capsid is built of P2 and P4 protein heterodimers, each with more than 1000 residues. Despite a lack of sequence similarity between the two proteins, they have a similar α-helical domain, the structural signature shared with the lineage of the dsRNA bluetongue virus-like viruses. Domain insertions in P2 and P4 preferential sites provide additional functions at the capsid outer surface, probably related to enzyme activity. The P2 insertion has a fold similar to that of gelsolin and profilin, two actin-binding proteins with a function in cytoskeleton metabolism, whereas the P4 insertion suggests protease activity involved in cleavage of the P2 383-residue C-terminal region, absent in the mature viral particle. Our results indicate that the intimate virus-fungus partnership has altered the capsid genome-protective and/or receptor-binding functions. Fungal virus evolution has tended to allocate enzyme activities to the virus capsid outer surface.

  15. Applications of Total Scattering & Pair Distribution Function Analysis in Metal-Organic Framework Materials

    DEFF Research Database (Denmark)

    Xu, Hui; Birgisson, Steinar; Sommer, Sanna

    Metal-Organic Frameworks (MOFs) is constructed by metal-oxide nodes and organic ligands. The formation of different structures of metal-oxide nodes (also called secondary building units, SBU) is crucial for MOF final structures, because the connectivity of SBU greatly influence the final MOF...... structure. At the same time, there is an ongoing debate on whether the SBU is present prior, or during MOF crystallization in MOF chemistry. However, little is known about MOFs formation mechanism. Currently techniques to study the in situ MOF formation process mainly focused on after......-crystallization process, for example in situ XRD and SAXS/WAXS study on MOF formation. However, the pre-crystallization process in the early stage of MOF formation is still unexplored. In this project, total scattering and PDF study will be carried out to explore the MOF formation process in early stage. This includes...

  16. Morphological and Functional Changes in Rat Brain under Total Sleep Deprivation

    Directory of Open Access Journals (Sweden)

    Babak ABUSHOV

    2015-07-01

    Full Text Available In this article the effect of 36 h total sleep deprivation (TSD on behavioural reactions of rats (rearing, grooming and sexual activity and on ultrastructure of brain neurons have been studied. A group of somnogenic structures (III-V layers of the frontal limbic cortex, CA1 area of the dorsal hippocampus, reticular formation of pons varolli, nucleus raphe dorsalis and locus coeruleus of the brain of 6 month-old Wistar rats has been analized in this study. It has been revealed that dystrophic changes (chromatolysis of cytoplasm and vacuolization enveloping a group of medium-sized neurons (20-30 (μ micron in diameter are accompanied by decrease of numbers of rearing, grooming and sexual activity. It is supposed that dystrophic changes revealed in medium-sized neurons result in partial disturbances of integrative and behavioral reactions of animals.

  17. Comparison of Free Total Amino Acid Compositions and Their Functional Classifications in 13 Wild Edible Mushrooms.

    Science.gov (United States)

    Sun, Liping; Liu, Qiuming; Bao, Changjun; Fan, Jian

    2017-02-24

    Thirteen popular wild edible mushroom species in Yunnan Province, Boletus bicolor , Boletus speciosus , Boletus sinicus , Boletus craspedius , Boletus griseus , Boletus ornatipes , Xerocomus , Suillus placidus , Boletinus pinetorus , Tricholoma terreum , Tricholomopsis lividipileata , Termitomyces microcarpus , and Amanita hemibapha , were analyzed for their free amino acid compositions by online pre-column derivazation reversed phase high-performance liquid chromatography (RP-HPLC) analysis. Twenty free amino acids, aspartic acid, glutamic acid, serine, glycine, alanine, praline, cysteine, valine, methionine, phenylalanine, isoleucine, leucine, lysine, histidine, threonine, asparagines, glutamine, arginine, tyrosine, and tryptophan, were determined. The total free amino acid (TAA) contents ranged from 1462.6 mg/100 g in B. craspedius to 13,106.2 mg/100 g in T. microcarpus . The different species showed distinct free amino acid profiles. The ratio of total essential amino acids (EAA) to TAA was 0.13-0.41. All of the analyzed species showed high contents of hydrophobic amino acids, at 33%-54% of TAA. Alanine, cysteine, glutamine, and glutamic acid were among the most abundant amino acids present in all species. The results showed that the analyzed mushrooms possessed significant free amino acid contents, which may be important compounds contributing to the typical mushroom taste, nutritional value, and potent antioxidant properties of these wild edible mushrooms. Furthermore, the principal component analysis (PCA) showed that the accumulative variance contribution rate of the first four principal components reached 94.39%. Cluster analysis revealed EAA composition and content might be an important parameter to separate the mushroom species, and T. microcarpus and A. hemibapha showed remarkable EAA content among the 13 species.

  18. Comparison of Free Total Amino Acid Compositions and Their Functional Classifications in 13 Wild Edible Mushrooms

    Directory of Open Access Journals (Sweden)

    Liping Sun

    2017-02-01

    Full Text Available Thirteen popular wild edible mushroom species in Yunnan Province, Boletus bicolor, Boletus speciosus, Boletus sinicus, Boletus craspedius, Boletus griseus, Boletus ornatipes, Xerocomus, Suillus placidus, Boletinus pinetorus, Tricholoma terreum, Tricholomopsis lividipileata, Termitomyces microcarpus, and Amanita hemibapha, were analyzed for their free amino acid compositions by online pre-column derivazation reversed phase high-performance liquid chromatography (RP-HPLC analysis. Twenty free amino acids, aspartic acid, glutamic acid, serine, glycine, alanine, praline, cysteine, valine, methionine, phenylalanine, isoleucine, leucine, lysine, histidine, threonine, asparagines, glutamine, arginine, tyrosine, and tryptophan, were determined. The total free amino acid (TAA contents ranged from 1462.6 mg/100 g in B. craspedius to 13,106.2 mg/100 g in T. microcarpus. The different species showed distinct free amino acid profiles. The ratio of total essential amino acids (EAA to TAA was 0.13–0.41. All of the analyzed species showed high contents of hydrophobic amino acids, at 33%–54% of TAA. Alanine, cysteine, glutamine, and glutamic acid were among the most abundant amino acids present in all species. The results showed that the analyzed mushrooms possessed significant free amino acid contents, which may be important compounds contributing to the typical mushroom taste, nutritional value, and potent antioxidant properties of these wild edible mushrooms. Furthermore, the principal component analysis (PCA showed that the accumulative variance contribution rate of the first four principal components reached 94.39%. Cluster analysis revealed EAA composition and content might be an important parameter to separate the mushroom species, and T. microcarpus and A. hemibapha showed remarkable EAA content among the 13 species.

  19. T7-RNA Polymerase

    Science.gov (United States)

    1997-01-01

    T7-RNA Polymerase grown on STS-81. Structure-Function Relationships of RNA Polymerase: DNA-dependent RNA polymerase is the key enzyme responsible for the biosynthesis of RNA, a process known as transcription. Principal Investigator's include Dr. Dan Carter, Dr. B.C. Wang, and Dr. John Rose of New Century Pharmaceuticals.

  20. Low genetic diversity and functional constraint of miRNA genes participating pollen-pistil interaction in rice.

    Science.gov (United States)

    Wang, Kun; Wang, Xin; Li, Ming; Shi, Tao; Yang, Pingfang

    2017-09-01

    In this study, we sequenced and analyzed the expression and evolution of rice miRNA genes participating pollen-pistil interaction that is crucial to rice yield. Pollen-pistil interaction is an essential reproductive process for all flowering plants. While microRNAs (miRNAs) are important noncoding small RNAs that regulate mRNA levels in eukaryotic cells, there is little knowledge about which miRNAs involved in the early stages of pollen-pistil interaction in rice and how they evolve under this conserved process. In this study, we sequenced the small RNAs in rice from unpollinated pistil (R0), pistil from 5 min and 15 min after pollination, respectively, to identify known and novel miRNAs that are involved in this process. By comparing the corresponding mRNA-seq dataset, we identified a group of miRNAs with strong negative expression pattern with their target genes. Further investigation of all miRNA loci (MIRNAs) across 1083 public rice accessions revealed significantly reduced genetic diversity in MIRNAs with strong negative expression of their targets when comparing to those with little or no impact on targets during pollen-pistil interaction. Annotation of targets suggested that those MIRNAs with strong impact on targets were pronounced in cell wall related processes such as xylan metabolism. Additionally, plant conserved miRNAs, such as those with functions in gibberellic acid, auxin and nitrate signaling, were also with strong negative expression of their targets. Overall, our analyses identified key miRNAs participating pollen-pistil interaction and their evolutionary patterns in rice, which can facilitate the understanding of molecular mechanisms associated with seed setting.

  1. An amphipathic alpha-helix controls multiple roles of brome mosaic virus protein 1a in RNA replication complex assembly and function.

    Directory of Open Access Journals (Sweden)

    Ling Liu

    2009-03-01

    Full Text Available Brome mosaic virus (BMV protein 1a has multiple key roles in viral RNA replication. 1a localizes to perinuclear endoplasmic reticulum (ER membranes as a peripheral membrane protein, induces ER membrane invaginations in which RNA replication complexes form, and recruits and stabilizes BMV 2a polymerase (2a(Pol and RNA replication templates at these sites to establish active replication complexes. During replication, 1a provides RNA capping, NTPase and possibly RNA helicase functions. Here we identify in BMV 1a an amphipathic alpha-helix, helix A, and use NMR analysis to define its structure and propensity to insert in hydrophobic membrane-mimicking micelles. We show that helix A is essential for efficient 1a-ER membrane association and normal perinuclear ER localization, and that deletion or mutation of helix A abolishes RNA replication. Strikingly, mutations in helix A give rise to two dramatically opposite 1a function phenotypes, implying that helix A acts as a molecular switch regulating the intricate balance between separable 1a functions. One class of helix A deletions and amino acid substitutions markedly inhibits 1a-membrane association and abolishes ER membrane invagination, viral RNA template recruitment, and replication, but doubles the 1a-mediated increase in 2a(Pol accumulation. The second class of helix A mutations not only maintains efficient 1a-membrane association but also amplifies the number of 1a-induced membrane invaginations 5- to 8-fold and enhances viral RNA template recruitment, while failing to stimulate 2a(Pol accumulation. The results provide new insights into the pathways of RNA replication complex assembly and show that helix A is critical for assembly and function of the viral RNA replication complex, including its central role in targeting replication components and controlling modes of 1a action.

  2. Relationship between total testosterone, cognitive function, depressive behavior, and sleep quality in chronic kidney disease patients not on dialysis.

    Science.gov (United States)

    Afsar, Baris

    2013-02-01

    Studies show that testosterone levels are associated with cognitive function, depression, and sleep quality in the general population. However, these relationships in chronic kidney disease (CKD) patients not on dialysis have not yet been evaluated before. All patients underwent history taking, physical examination, blood pressure measurement, routine urine and biochemical analysis, 24-h urine collection to measure urinary protein excretion and creatinine clearance, and evaluation of cognitive function, depressive behavior, and sleep quality. In total, 109 CKD patients were enrolled. Total testosterone levels in stage 3, 4, and 5 CKD patients were 8.32 ± 4.35, 6.71 ± 3.12, and 4.22 ± 1.28 ng/ml, respectively (p Sleep Quality Index (PSQI) score, and Beck Depression Inventory (BDI) score were 26.2 ± 1.9, 7.1 ± 3.4, and 8.6 ± 6.4, respectively. In linear regression analysis, total testosterone levels were independently associated with SMMSE score [b 0.170, confidence interval (CI) 0.047-0.293, p 0.008] and BDI score (b -0.750, CI -1.283 to -0.216, p 0.006) but not with sleep quality. Total serum testosterone levels were independently associated with cognitive function and depressive behavior but not with sleep disorders in stage 3-5 CKD patients not on dialysis.

  3. Early self-managed focal sensorimotor rehabilitative training enhances functional mobility and sensorimotor function in patients following total knee replacement: a controlled clinical trial.

    Science.gov (United States)

    Moutzouri, Maria; Gleeson, Nigel; Coutts, Fiona; Tsepis, Elias; John, Gliatis

    2018-02-01

    To assess the effects of early self-managed focal sensorimotor training compared to functional exercise training after total knee replacement on functional mobility and sensorimotor function. A single-blind controlled clinical trial. University Hospital of Rion, Greece. A total of 52 participants following total knee replacement. The primary outcome was the Timed Up and Go Test and the secondary outcomes were balance, joint position error, the Knee Outcome Survey Activities of Daily Living Scale, and pain. Patients were assessed on three separate occasions (presurgery, 8 weeks post surgery, and 14 weeks post surgery). Participants were randomized to either focal sensorimotor exercise training (experimental group) or functional exercise training (control group). Both groups received a 12-week home-based programme prescribed for 3-5 sessions/week (35-45 minutes). Consistently greater improvements ( F 2,98  = 4.3 to 24.8; P functional mobility and sensorimotor function endorses using focal sensorimotor training as an effective mode of rehabilitation following knee replacement.

  4. MicroRNA repertoire for functional genome research in tilapia identified by deep sequencing.

    Science.gov (United States)

    Yan, Biao; Wang, Zhen-Hua; Zhu, Chang-Dong; Guo, Jin-Tao; Zhao, Jin-Liang

    2014-08-01

    The Nile tilapia (Oreochromis niloticus; Cichlidae) is an economically important species in aquaculture and occupies a prominent position in the aquaculture industry. MicroRNAs (miRNAs) are a class of noncoding RNAs that post-transcriptionally regulate gene expression involved in diverse biological and metabolic processes. To increase the repertoire of miRNAs characterized in tilapia, we used the Illumina/Solexa sequencing technology to sequence a small RNA library using pooled RNA sample isolated from the different developmental stages of tilapia. Bioinformatic analyses suggest that 197 conserved and 27 novel miRNAs are expressed in tilapia. Sequence alignments indicate that all tested miRNAs and miRNAs* are highly conserved across many species. In addition, we characterized the tissue expression patterns of five miRNAs using real-time quantitative PCR. We found that miR-1/206, miR-7/9, and miR-122 is abundantly expressed in muscle, brain, and liver, respectively, implying a potential role in the regulation of tissue differentiation or the maintenance of tissue identity. Overall, our results expand the number of tilapia miRNAs, and the discovery of miRNAs in tilapia genome contributes to a better understanding the role of miRNAs in regulating diverse biological processes.

  5. Total Cerebral Small Vessel Disease MRI Score Is Associated With Cognitive Decline In Executive Function In Patients With Hypertension

    Directory of Open Access Journals (Sweden)

    Renske Uiterwijk

    2016-12-01

    Full Text Available Objectives: Hypertension is a major risk factor for white matter hyperintensities, lacunes, cerebral microbleeds and perivascular spaces, which are MRI markers of cerebral small vessel disease (SVD. Studies have shown associations between these individual MRI markers and cognitive functioning and decline. Recently, a total SVD score was proposed in which the different MRI markers were combined into one measure of SVD, to capture total SVD-related brain damage. We investigated if this SVD score was associated with cognitive decline over 4 years in patients with hypertension. Methods: In this longitudinal cohort study, 130 hypertensive patients (91 patients with uncomplicated hypertension and 39 hypertensive patients with a lacunar stroke were included. They underwent a neuropsychological assessment at baseline and after 4 years. The presence of white matter hyperintensities, lacunes, cerebral microbleeds, and perivascular spaces were rated on baseline MRI. Presence of each individual marker was added to calculate the total SVD score (range 0-4 in each patient. Results: Uncorrected linear regression analyses showed associations between SVD score and decline in overall cognition (p=0.017, executive functioning (p<0.001 and information processing speed (p=0.037, but not with memory (p=0.911. The association between SVD score and decline in overall cognition and executive function remained significant after adjustment for age, sex, education, anxiety and depression score, potential vascular risk factors, patient group and baseline cognitive performance.Conclusions: Our study shows that a total SVD score can predict cognitive decline, specifically in executive function, over 4 years in hypertensive patients. This emphasizes the importance of considering total brain damage due to SVD.

  6. The total scattering atomic pair distribution function: New methodology for nanostructure analysis

    Science.gov (United States)

    Masadeh, Ahmad

    The conventional xray diffration (XRD) methods probe for the presence of long-range order (periodic structure) which are reflected in the Bragg peaks. Local structural deviations or disorder mainly affect the diffuse scattering intensity. In order to obtain structural information about both long-range order and local structure disorder, a technique that takes in account both Bragg and diffuse scattering need to be employed, such as the atomic pair distribution function (PDF) technique. This work introduces a PDF based methodology to quantitatively investigate nanostructure materials in general. The introduced methodology can be applied to extract quantitatively structural information about structure, crystallinity level, core/shell size, nanoparticle size, and inhomogeneous internal strain in the measured nanoparticles. This method is generally applicable to the characterization of the nano-scale solid, many of which may exhibit complex disorder and strain

  7. Total cervical disk replacement with a prestige LP® prosthesis: clinical and functional outcomes

    Directory of Open Access Journals (Sweden)

    Rafael Osório Rocha

    2014-01-01

    Full Text Available Objective: To determine the clinical and functional results of short- and medium-term cervical arthroplasty with the Prestige LP® prosthesis for the treatment of compressive myelopathy, radiculopathy and axial pain with radiculopathty. Methods: This retrospective study, conducted from 2009 to 2012, included 18 patients. Only 16 were found for the second stage of research, conducted in 2011 and 2012. Pre- and postoperative assessments were carried out using the CSOQ (Cervical Spine Outcomes Questionnaire. Odom criteria were used only in the postoperative evaluation. Both were translated and adapted to the local culture. Results: There was no postoperative radiculopathy or other complications requiring prolonged hospitalization. In most patients, there was a significant improvement in axial pain and radiculopathy, and there was only one indication of conversion to fusion. Conclusions: In selected cases of cervical degenerative disc disease, herniated cervical disc and compressive myeolopathy, cervical arthroplasty proved to be an effective and safe treatment in the short and medium terms.

  8. Similar range of motion and function after resurfacing large-head or standard total hip arthroplasty

    DEFF Research Database (Denmark)

    Penny, Jeannette Østergaard; Ovesen, Ole; Varmarken, Jens-Erik

    2013-01-01

    BACKGROUND AND PURPOSE: Large-size hip articulations may improve range of motion (ROM) and function compared to a 28-mm THA, and the low risk of dislocation allows the patients more activity postoperatively. On the other hand, the greater extent of surgery for resurfacing hip arthroplasty (RHA......° (35), 232° (36), and 225° (30) respectively, but the differences were not statistically significant. The 3 groups were similar regarding Harris hip score, UCLA activity score, step rate, and sick leave. INTERPRETATION: Head size had no influence on range of motion. The lack of restriction allowed...... for large articulations did not improve the clinical and patient-perceived outcomes. The more extensive surgical procedure of RHA did not impair the rehabilitation. This project is registered at ClinicalTrials.gov under # NCT01113762....

  9. Irreversible Total Loss of Brain Function and Organ Donation in Patients with Aneurysmal Subarachnoid Hemorrhage.

    Science.gov (United States)

    Mohme, Malte; Sauvigny, Thomas; Grensemann, Jörn; Söffker, Gerold; Kluge, Stefan; Westphal, Manfred; Czorlich, Patrick

    2017-09-01

    Aneurysmal subarachnoid hemorrhage (aSAH) is a life-threatening disease with an often fatal clinical course leading to irreversible loss of brain function (ILBF) (i.e., brain death). The purpose of this study was to assess the prevalence and characteristics of patients with aSAH who became organ donors after diagnosis of ILBF. Anonymized clinical data sets of 395 patients treated for aSAH at a university medical center from January 2011 to December 2016 were retrospectively analyzed. Prevalence of consent for organ donation and clinical characteristics, including parameters for diagnosis of irreversible loss of brain function, were assessed. After initial admission to the intensive care unit, 18.0% of patients (n = 71) died (Glasgow Outcome Scale score 1). Intracerebral hemorrhage occurred in 42.3% of patients who died, aneurysmal rebleeding occurred in 19.7%, and intraventricular hemorrhage occurred in 87.3%. In 50.7% of patients who died (n = 36), ILBF was diagnosed, and 32.4% (n = 23) of these patients became organ donors. In 55.6%, additional diagnostic electroencephalography was performed. Male patients significantly more often became organ donors than female patients (P = 0.008). ILBF with subsequent organ donation was predominantly seen in patients organs were explanted for donation, including 42 kidneys, 21 livers, 3 pancreas, 11 hearts and 8 lungs. ILBF in the setting of fatal aSAH is a prevalent diagnosis with complex demands for neurointensive care physicians. We demonstrated the clinical characteristics and epidemiologic factors of patients with aSAH converting to organ donors. Copyright © 2017 Elsevier Inc. All rights reserved.

  10. Influence of black cohosh (Cimicifuga racemosa) use by postmenopausal women on total hepatic perfusion and liver functions.

    Science.gov (United States)

    Nasr, Ahmed; Nafeh, Hanan

    2009-11-01

    In this prospective longitudinal clinical trial, 87 postmenopausal women receiving for 12 consecutive months a daily dose of 40 mg of a dry extract preparation of Cimicifuga racemosa (Klimadynon) for relief of vasomotor symptoms were followed up by evaluation of total hepatic blood flow, assessed by color Doppler ultrasound, as well as prothrombin time and concentration, serum albumin, bilirubin, gamma-glutamyltransferase, alkaline phosphatase, alanine aminotransferase, and aspartate aminotransferase. Because no significant changes in total hepatic blood flow or any of the liver functions tested were reported, we concluded that use of C. racemosa for 1 year by healthy postmenopausal women without evidence of liver disease does not seem to influence the liver.

  11. Pre-operative functional mobility as an independent determinant of inpatient functional recovery after total knee arthroplasty during three periods that coincided with changes in clinical pathways

    NARCIS (Netherlands)

    Sluis, G. van der; Goldbohm, R.A.; Elings, J.E.; Nijhuis-Van der Sanden, M.W.G.; Akkermans, R.P.; Bimmel, R.; Hoogeboom, T.J.; Meeteren, N.L. van

    2017-01-01

    AIMS: To investigate whether pre-operative functional mobility is a determinant of delayed inpatient recovery of activities (IRoA) after total knee arthroplasty (TKA) in three periods that coincided with changes in the clinical pathway. PATIENTS AND METHODS: All patients (n = 682, 73% women, mean

  12. Identification of active methanotrophs in a landfill cover soil through detection of expression of 16S rRNA and functional genes.

    Science.gov (United States)

    Chen, Yin; Dumont, Marc G; Cébron, Aurélie; Murrell, J Colin

    2007-11-01

    Active methanotrophs in a landfill soil were revealed by detecting the 16S rRNA of methanotrophs and the mRNA transcripts of key genes involved in methane oxidation. New 16S rRNA primers targeting type I and type II methanotrophs were designed and optimized for analysis by denaturing gradient gel electrophoresis. Direct extraction of RNA from soil enabled the analysis of the expression of the functional genes: mmoX, pmoA and mxaF, which encode subunits of soluble methane monooxygenase, particulate methane monooxygenase and methanol dehydrogenase respectively. The 16S rRNA polymerase chain reaction (PCR) primers for type I methanotrophs detected Methylomonas, Methylosarcina and Methylobacter sequences from both soil DNA and cDNA which was generated from RNA extracted directly from the landfill cover soil. The 16S rRNA primers for type II methanotrophs detected primarily Methylocella and some Methylocystis 16S rRNA genes. Phylogenetic analysis of mRNA recovered from the soil indicated that Methylobacter, Methylosarcina, Methylomonas, Methylocystis and Methylocella were actively expressing genes involved in methane and methanol oxidation. Transcripts of pmoA but not mmoX were readily detected by reverse transcription polymerase chain reaction (RT-PCR), indicating that particulate methane monooxygenase may be largely responsible for methane oxidation in situ.

  13. The function of the RNA-binding protein TEL1 in moss reveals ancient regulatory mechanisms of shoot development.

    Science.gov (United States)

    Vivancos, Julien; Spinner, Lara; Mazubert, Christelle; Charlot, Florence; Paquet, Nicolas; Thareau, Vincent; Dron, Michel; Nogué, Fabien; Charon, Céline

    2012-03-01

    The shoot represents the basic body plan in land plants. It consists of a repeated structure composed of stems and leaves. Whereas vascular plants generate a shoot in their diploid phase, non-vascular plants such as mosses form a shoot (called the gametophore) in their haploid generation. The evolution of regulatory mechanisms or genetic networks used in the development of these two kinds of shoots is unclear. TERMINAL EAR1-like genes have been involved in diploid shoot development in vascular plants. Here, we show that disruption of PpTEL1 from the moss Physcomitrella patens, causes reduced protonema growth and gametophore initiation, as well as defects in gametophore development. Leafy shoots formed on ΔTEL1 mutants exhibit shorter stems with more leaves per shoot, suggesting an accelerated leaf initiation (shortened plastochron), a phenotype shared with the Poaceae vascular plants TE1 and PLA2/LHD2 mutants. Moreover, the positive correlation between plastochron length and leaf size observed in ΔTEL1 mutants suggests a conserved compensatory mechanism correlating leaf growth and leaf initiation rate that would minimize overall changes in plant biomass. The RNA-binding protein encoded by PpTEL1 contains two N-terminus RNA-recognition motifs, and a third C-terminus non-canonical RRM, specific to TEL proteins. Removal of the PpTEL1 C-terminus (including this third RRM) or only 16-18 amino acids within it seriously impairs PpTEL1 function, suggesting a critical role for this third RRM. These results show a conserved function of the RNA-binding PpTEL1 protein in the regulation of shoot development, from early ancestors to vascular plants, that depends on the third TEL-specific RRM.

  14. The expanding functions of cellular helicases: the tombusvirus RNA replication enhancer co-opts the plant eIF4AIII-like AtRH2 and the DDX5-like AtRH5 DEAD-box RNA helicases to promote viral asymmetric RNA replication.

    Directory of Open Access Journals (Sweden)

    Nikolay Kovalev

    2014-04-01

    Full Text Available Replication of plus-strand RNA viruses depends on recruited host factors that aid several critical steps during replication. Several of the co-opted host factors bind to the viral RNA, which plays multiple roles, including mRNA function, as an assembly platform for the viral replicase (VRC, template for RNA synthesis, and encapsidation during infection. It is likely that remodeling of the viral RNAs and RNA-protein complexes during the switch from one step to another requires RNA helicases. In this paper, we have discovered a second group of cellular RNA helicases, including the eIF4AIII-like yeast Fal1p and the DDX5-like Dbp3p and the orthologous plant AtRH2 and AtRH5 DEAD box helicases, which are co-opted by tombusviruses. Unlike the previously characterized DDX3-like AtRH20/Ded1p helicases that bind to the 3' terminal promoter region in the viral minus-strand (-RNA, the other class of eIF4AIII-like RNA helicases bind to a different cis-acting element, namely the 5' proximal RIII(- replication enhancer (REN element in the TBSV (-RNA. We show that the binding of AtRH2 and AtRH5 helicases to the TBSV (-RNA could unwind the dsRNA structure within the RIII(- REN. This unique characteristic allows the eIF4AIII-like helicases to perform novel pro-viral functions involving the RIII(- REN in stimulation of plus-strand (+RNA synthesis. We also show that AtRH2 and AtRH5 helicases are components of the tombusvirus VRCs based on co-purification experiments. We propose that eIF4AIII-like helicases destabilize dsRNA replication intermediate within the RIII(- REN that promotes bringing the 5' and 3' terminal (-RNA sequences in close vicinity via long-range RNA-RNA base pairing. This newly formed RNA structure promoted by eIF4AIII helicase together with AtRH20 helicase might facilitate the recycling of the viral replicases for multiple rounds of (+-strand synthesis, thus resulting in asymmetrical viral replication.

  15. The expanding functions of cellular helicases: the tombusvirus RNA replication enhancer co-opts the plant eIF4AIII-like AtRH2 and the DDX5-like AtRH5 DEAD-box RNA helicases to promote viral asymmetric RNA replication.

    Science.gov (United States)

    Kovalev, Nikolay; Nagy, Peter D

    2014-04-01

    Replication of plus-strand RNA viruses depends on recruited host factors that aid several critical steps during replication. Several of the co-opted host factors bind to the viral RNA, which plays multiple roles, including mRNA function, as an assembly platform for the viral replicase (VRC), template for RNA synthesis, and encapsidation during infection. It is likely that remodeling of the viral RNAs and RNA-protein complexes during the switch from one step to another requires RNA helicases. In this paper, we have discovered a second group of cellular RNA helicases, including the eIF4AIII-like yeast Fal1p and the DDX5-like Dbp3p and the orthologous plant AtRH2 and AtRH5 DEAD box helicases, which are co-opted by tombusviruses. Unlike the previously characterized DDX3-like AtRH20/Ded1p helicases that bind to the 3' terminal promoter region in the viral minus-strand (-)RNA, the other class of eIF4AIII-like RNA helicases bind to a different cis-acting element, namely the 5' proximal RIII(-) replication enhancer (REN) element in the TBSV (-)RNA. We show that the binding of AtRH2 and AtRH5 helicases to the TBSV (-)RNA could unwind the dsRNA structure within the RIII(-) REN. This unique characteristic allows the eIF4AIII-like helicases to perform novel pro-viral functions involving the RIII(-) REN in stimulation of plus-strand (+)RNA synthesis. We also show that AtRH2 and AtRH5 helicases are components of the tombusvirus VRCs based on co-purification experiments. We propose that eIF4AIII-like helicases destabilize dsRNA replication intermediate within the RIII(-) REN that promotes bringing the 5' and 3' terminal (-)RNA sequences in close vicinity via long-range RNA-RNA base pairing. This newly formed RNA structure promoted by eIF4AIII helicase together with AtRH20 helicase might facilitate the recycling of the viral replicases for multiple rounds of (+)-strand synthesis, thus resulting in asymmetrical viral replication.

  16. The origin, function, and diagnostic potential of RNA within extracellular vesicles present in human biological fluids

    Science.gov (United States)

    Taylor, Douglas D.; Gercel-Taylor, Cicek

    2013-01-01

    We have previously demonstrated that tumor cells release membranous structures into their extracellular environment, which are termed exosomes, microvesicles or extracellular vesicles depending on specific characteristics, including size, composition and biogenesis pathway. These cell-derived vesicles can exhibit an array of proteins, lipids and nucleic acids derived from the originating tumor. This review focuses of the transcriptome (RNA) of these extracellular vesicles. Based on current data, these vesicular components play essential roles as conveyers of intercellular communication and mediators of many of the pathological conditions associated with cancer development, progression and therapeutic failures. These extracellular vesicles express components responsible for angiogenesis promotion, stromal remodeling, signal pathway activation through growth factor/receptor transfer, chemoresistance, and genetic exchange. These tumor-derived extracellular vesicles not only to represent a central mediator of the tumor microenvironment, but their presence in the peripheral circulation may serve as a surrogate for tumor biopsies, enabling real-time diagnosis and disease monitoring. PMID:23908664

  17. The origin, function and diagnostic potential of RNA within extracellular vesicles present in human biological fluids

    Directory of Open Access Journals (Sweden)

    Douglas D. Taylor

    2013-07-01

    Full Text Available We have previously demonstrated that tumor cells release membranous structures into their extracellular environment, which are termed exosomes, microvesicles or extracellular vesicles depending on specific characteristics, including size, composition and biogenesis pathway. These cell-derived vesicles can exhibit an array of proteins, lipids and nucleic acids derived from the originating tumor. This review focuses of the transcriptome (RNA of these extracellular vesicles. Based on current data, these vesicular components play essential roles as conveyers of intercellular communication and mediators of many of the pathological conditions associated with cancer development, progression and therapeutic failures. These extracellular vesicles express components responsible for angiogenesis promotion, stromal remodeling, signal pathway activation through growth factor/receptor transfer, chemoresistance, and genetic exchange. These tumor-derived extracellular vesicles not only to represent a central mediator of the tumor microenvironment, but their presence in the peripheral circulation may serve as a surrogate for tumor biopsies, enabling real-time diagnosis and disease monitoring.

  18. Triggering of RNA interference with RNA-RNA, RNA-DNA, and DNA-RNA nanoparticles.

    Science.gov (United States)

    Afonin, Kirill A; Viard, Mathias; Kagiampakis, Ioannis; Case, Christopher L; Dobrovolskaia, Marina A; Hofmann, Jen; Vrzak, Ashlee; Kireeva, Maria; Kasprzak, Wojciech K; KewalRamani, Vineet N; Shapiro, Bruce A

    2015-01-27

    Control over cellular delivery of different functionalities and their synchronized activation is a challenging task. We report several RNA and RNA/DNA-based nanoparticles designed to conditionally activate the RNA interference in various human cells. These nanoparticles allow precise control over their formulation, stability in blood serum, and activation of multiple functionalities. Importantly, interferon and pro-inflammatory cytokine activation assays indicate the significantly lower responses for DNA nanoparticles compared to the RNA counterparts, suggesting greater potential of these molecules for therapeutic use.

  19. Study of the Effects of Total Modulation Transfer Function Changes on Observer Performance Using Clinical Mammograms.

    Science.gov (United States)

    Bencomo, Jose Antonio Fagundez

    The main goal of this study was to relate physical changes in image quality measured by Modulation Transfer Function (MTF) to diagnostic accuracy. One Hundred and Fifty Kodak Min-R screen/film combination conventional craniocaudal mammograms obtained with the Pfizer Microfocus Mammographic system were selected from the files of the Department of Radiology, at M.D. Anderson Hospital and Tumor Institute. The mammograms included 88 cases with a variety of benign diagnosis and 62 cases with a variety of malignant biopsy diagnosis. The average age of the patient population was 55 years old. 70 cases presented calcifications with 30 cases having calcifications smaller than 0.5mm. 46 cases presented irregular bordered masses larger than 1 cm. 30 cases presented smooth bordered masses with 20 larger than 1 cm. Four separated copies of the original images were made each having a different change in the MTF using a defocusing technique whereby copies of the original were obtained by light exposure through different thicknesses (spacing) of transparent film base. The mammograms were randomized, and evaluated by three experienced mammographers for the degree of visibility of various anatomical breast structures and pathological lesions (masses and calicifications), subjective image quality, and mammographic interpretation. 3,000 separate evaluations were anayzed by several statistical techniques including Receiver Operating Characteristic curve analysis, McNemar test for differences between proportions and the Landis et al. method of agreement weighted kappa for ordinal categorical data. Results from the statistical analysis show: (1) There were no statistical significant differences in the diagnostic accuracy of the observers when diagnosing from mammograms with the same MTF. (2) There were no statistically significant differences in diagnostic accuracy for each observer when diagnosing from mammograms with the different MTF's used in the study. (3) There statistical

  20. Functional Response of Tumor Vasculature to PaCO2: Determination of Total and Microvascular Blood Volume by MRI

    Directory of Open Access Journals (Sweden)

    Scott D. Packard

    2003-07-01

    Full Text Available In order to identify differences in functional activity, we compared the reactivity of glioma vasculature and the native cerebral vasculature to both dilate and constrict in response to altered PaCO2. Gliomas were generated by unilateral implantation of U87MGdEGFR human glioma tumor cells into the striatum of adult female athymic rats. Relative changes in total and microvascular cerebral blood volume were determined by steady state contrast agent-enhanced magnetic resonance imaging for transitions from normocarbia to hypercarbia and hypocarbia. Although hypercarbia induced a significant increase in both total and microvascular blood volume in normal brain and glioma, reactivity of glioma vasculature was significantly blunted in comparison to normal striatum; glioma total CBV increased by 0.6±0.1%/mm Hg CO2 whereas normal striatum increased by 1.5±0.2%/mm Hg CO2, (P < .0001, group t-test. Reactivity of microvascular blood volume was also significantly blunted. In contrast, hypocarbia decreased both total and microvascular blood volumes more in glioma than in normal striatum. These results indicate that cerebral blood vessels derived by tumor-directed angiogenesis do retain reactivity to CO2. Furthermore, reduced reactivity of tumor vessels to a single physiological perturbation, such as hypercarbia, should not be construed as a generalized reduction of functional activity of the tumor vascular bed.

  1. Dissection of functional lncRNAs in Alzheimer's disease by construction and analysis of lncRNA-mRNA networks based on competitive endogenous RNAs.

    Science.gov (United States)

    Wang, Lian-Kun; Chen, Xiao-Feng; He, Dan-Dan; Li, You; Fu, Jin

    2017-04-08

    Alzheimer's disease (AD) is a neurodegenerative disorder that is the most common cause of dementia in the elderly, and intracellular neurofibrillary tangles (NFTs) are one of the pathological features of AD. Recent studies have suggested long noncoding RNAs (lncRNAs) play important roles in AD. Competing endogenous RNAs (ceRNAs) is a mechanism that has recently been proposed, in which lncRNAs compete for common miRNA-binding sites with mRNAs. However, the roles of lncRNAs and ceRNA in AD NFTs is limited. In this study, we constructed a global triple network based on ceRNA theory, then an AD NFT lncRNA-mRNA network (NFTLMN) was generated. By analyzing the NFTLMN, three lncRNAs (AP000265.1, KB-1460A1.5 and RP11-145M9.4), which are highly related with AD NFTs were identified. To further explore the cross-talk between mRNAs and lncRNAs, a clustering module analysis was performed on the NFTLMN and two AD NFT related modules were identified. Our study provides a better understanding of the molecular basis of AD NFTs and may offer novel treatment strategies for AD. Copyright © 2016. Published by Elsevier Inc.

  2. Comparison of Early Functional Recovery After Total Hip Arthroplasty Using a Direct Anterior or Posterolateral Approach: A Randomized Controlled Trial.

    Science.gov (United States)

    Zhao, Hai-Yan; Kang, Peng-De; Xia, Ya-Yi; Shi, Xiao-Jun; Nie, Yong; Pei, Fu-Xing

    2017-11-01

    Controversy exists as to whether early functional outcomes differ after total hip arthroplasty performed using the direct anterior approach (DAA) or the posterolateral approach (PLA). One hundred twenty patients were enrolled in this study and were divided into 2 groups based on surgical approach. Group A included patients who had a total hip arthroplasty with a DAA, whereas group B included those with a PLA. Patients were randomized into the DAA or PLA groups (n = 60), and perioperative and postoperative outcomes were recorded. When compared with the PLA, the DAA had a shorter incision length (9.1 vs 13.1 cm; P group. However, the PLA had shorter operative times (65.5 vs 83.3 min, P = .03) and less intraoperative blood loss (123.8 vs 165.9 mL, P = .04). The DAA had significantly lower variance in cup inclination and anteversion. Similar rates of intraoperative complications were identified in the 2 groups. The DAA was associated with better functional recovery at 3 months based on the Harris hip score, University of California Los Angeles activity score, and gait analysis; however, functional recovery at 6 months was similar between the 2 groups. We found functional advantages in early recovery after the DAA compared with the PLA. The DAA can offer rapid functional recovery with less muscle damage, greater pain relief, and lower variance in cup inclination and anteversion. However, no functional difference was found at 6 months follow-up. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Functional Role of the microRNA-200 Family in Breast Morphogenesis and Neoplasia

    Directory of Open Access Journals (Sweden)

    Bylgja Hilmarsdottir

    2014-09-01

    Full Text Available Branching epithelial morphogenesis is closely linked to epithelial-to-mesenchymal transition (EMT, a process important in normal development and cancer progression. The miR-200 family regulates epithelial morphogenesis and EMT through a negative feedback loop with the ZEB1 and ZEB2 transcription factors. miR-200 inhibits expression of ZEB1/2 mRNA, which in turn can down-regulate the miR-200 family that further results in down-regulation of E-cadherin and induction of a mesenchymal phenotype. Recent studies show that the expression of miR-200 genes is high during late pregnancy and lactation, thereby indicating that these miRs are important for breast epithelial morphogenesis and differentiation. miR-200 genes have been studied intensively in relation to breast cancer progression and metastasis, where it has been shown that miR-200 members are down-regulated in basal-like breast cancer where the EMT phenotype is prominent. There is growing evidence that the miR-200 family is up-regulated in distal breast metastasis indicating that these miRs are important for colonization of metastatic breast cancer cells through induction of mesenchymal to epithelial transition. The dual role of miR-200 in primary and metastatic breast cancer is of interest for future therapeutic interventions, making it important to understand its role and interacting partners in more detail.

  4. Translational recognition of the 5'-terminal 7-methylguanosine of globin messenger RNA as a function of ionic strength.

    Science.gov (United States)

    Chu, L Y; Rhoads, R E

    1978-06-13

    The translation of rabbit globin mRNA in cell-free systems derived from either wheat germ or rabbit reticulocyte was studied in the presence of various analogues of the methylated 5' terminus (cap) as a function of ionic strength. Inhibition by these analogues was strongly enhanced by increasing concentrations of KCl, K(OAc), Na(OAc), or NH4(OAc). At appropriate concentrations of K(OAc), both cell-free systems were equally sensitive to inhibition by m7GTP. At 50 mM K(OAc), the reticulocyte system was not sensitive to m7GMP or m7GTP, but at higher concentrations up to 200 mM K(OAc), both nucleotides caused strong inhibition. The compound in m7G5'ppp5'Am was inhibitory at all concentrations of K(OAc) ranging from 50 to 200 mM, although more strongly so at the higher concentrations. Over the same range of nucleotide concentrations, the compounds GMP, GTP, and G5'ppp5'Am were not inhibitors. The mobility on sodium dodecyl sulfate-polyacrylamide electrophoresis of the translation product was that of globin at all K(OAc) concentrations in the presence of m7GTP. Globin mRNA from which the terminal m7GTP group had been removed by chemical treatment (periodate-cyclohexylamine-alkaline phosphatase) or enzymatic treatment (tobacco acid pyrophosphatase-alkaline phosphatase) was translated less efficiently than untreated globin mRNA at higher K(OAc) concentrations, but retained appreciable activity at low K(OAc) concentrations.

  5. Functional and structural insights revealed by molecular dynamics simulations of an essential RNA editing ligase in Trypanosoma brucei.

    Directory of Open Access Journals (Sweden)

    Rommie E Amaro

    2007-11-01

    Full Text Available RNA editing ligase 1 (TbREL1 is required for the survival of both the insect and bloodstream forms of Trypanosoma brucei, the parasite responsible for the devastating tropical disease African sleeping sickness. The type of RNA editing that TbREL1 is involved in is unique to the trypanosomes, and no close human homolog is known to exist. In addition, the high-resolution crystal structure revealed several unique features of the active site, making this enzyme a promising target for structure-based drug design. In this work, two 20 ns atomistic molecular dynamics (MD simulations are employed to investigate the dynamics of TbREL1, both with and without the ATP substrate present. The flexibility of the active site, dynamics of conserved residues and crystallized water molecules, and the interactions between TbREL1 and the ATP substrate are investigated and discussed in the context of TbREL1's function. Differences in local and global motion upon ATP binding suggest that two peripheral loops, unique to the trypanosomes, may be involved in interdomain signaling events. Notably, a significant structural rearrangement of the enzyme's active site occurs during the apo simulations, opening an additional cavity adjacent to the ATP binding site that could be exploited in the development of effective inhibitors directed against this protozoan parasite. Finally, ensemble averaged electrostatics calculations over the MD simulations reveal a novel putative RNA binding site, a discovery that has previously eluded scientists. Ultimately, we use the insights gained through the MD simulations to make several predictions and recommendations, which we anticipate will help direct future experimental studies and structure-based drug discovery efforts against this vital enzyme.

  6. Improved method for prioritization of disease associated lncRNAs based on ceRNA theory and functional genomics data.

    Science.gov (United States)

    Wang, Peng; Guo, Qiuyan; Gao, Yue; Zhi, Hui; Zhang, Yan; Liu, Yue; Zhang, Jizhou; Yue, Ming; Guo, Maoni; Ning, Shangwei; Zhang, Guangmei; Li, Xia

    2017-01-17

    Although several computational models that predict disease-associated lncRNAs (long non-coding RNAs) exist, only a limited number of disease-associated lncRNAs are known. In this study, we mapped lncRNAs to their functional genomics context using competing endogenous RNAs (ceRNAs) theory. Based on the criteria that similar lncRNAs are likely involved in similar diseases, we proposed a disease lncRNA prioritization method, DisLncPri, to identify novel disease-lncRNA associations. Using a leave-one-out cross validation (LOOCV) strategy, DisLncPri achieved reliable area under curve (AUC) values of 0.89 and 0.87 for the LncRNADisease and Lnc2Cancer datasets that further improved to 0.90 and 0.89 by integrating a multiple rank fusion strategy. We found that DisLncPri had the highest rank enrichment score and AUC value in comparison to several other methods for case studies of alzheimer's disease, ovarian cancer, pancreatic cancer and gastric cancer. Several novel lncRNAs in the top ranks of these diseases were found to be newly verified by relevant databases or reported in recent studies. Prioritization of lncRNAs from a microarray (GSE53622) of oesophageal cancer patients highlighted ENSG00000226029 (top 2), a previously unidentified lncRNA as a potential prognostic biomarker. Our analysis thus indicates that DisLncPri is an excellent tool for identifying lncRNAs that could be novel biomarkers and therapeutic targets in a variety of human diseases.

  7. The rRNA methyltransferase Bud23 shows functional interaction with components of the SSU processome and RNase MRP.

    Science.gov (United States)

    Sardana, Richa; White, Joshua P; Johnson, Arlen W

    2013-06-01

    Bud23 is responsible for the conserved methylation of G1575 of 18S rRNA, in the P-site of the small subunit of the ribosome. bud23Δ mutants have severely reduced small subunit levels and show a general failure in cleavage at site A2 during rRNA processing. Site A2 is the primary cleavage site for separating the precursors of 18S and 25S rRNAs. Here, we have taken a genetic approach to identify the functional environment of BUD23. We found mutations in UTP2 and UTP14, encoding components of the SSU processome, as spontaneous suppressors of a bud23Δ mutant. The suppressors improved growth and subunit balance and restored cleavage at site A2. In a directed screen of 50 ribosomal trans-acting factors, we identified strong positive and negative genetic interactions with components of the SSU processome and strong negative interactions with components of RNase MRP. RNase MRP is responsible for cleavage at site A3 in pre-rRNA, an alternative cleavage site for separating the precursor rRNAs. The strong negative genetic interaction between RNase MRP mutants and bud23Δ is likely due to the combined defects in cleavage at A2 and A3. Our results suggest that Bud23 plays a role at the time of A2 cleavage, earlier than previously thought. The genetic interaction with the SSU processome suggests that Bud23 could be involved in triggering disassembly of the SSU processome, or of particular subcomplexes of the processome.

  8. MicroRNA-31 functions as an oncogenic microRNA in mouse and human lung cancer cells by repressing specific tumor suppressors

    DEFF Research Database (Denmark)

    Liu, Xi; Sempere, Lorenzo F; Ouyang, Haoxu

    2010-01-01

    MicroRNAs (miRNAs) regulate gene expression. It has been suggested that obtaining miRNA expression profiles can improve classification, diagnostic, and prognostic information in oncology. Here, we sought to comprehensively identify the miRNAs that are overexpressed in lung cancer by conducting mi......RNA microarray expression profiling on normal lung versus adjacent lung cancers from transgenic mice. We found that miR-136, miR-376a, and miR-31 were each prominently overexpressed in murine lung cancers. Real-time RT-PCR and in situ hybridization (ISH) assays confirmed these miRNA expression profiles in paired...... normal-malignant lung tissues from mice and humans. Engineered knockdown of miR-31, but not other highlighted miRNAs, substantially repressed lung cancer cell growth and tumorigenicity in a dose-dependent manner. Using a bioinformatics approach, we identified miR-31 target mRNAs and independently...

  9. Supplemental single shot femoral nerve block for total hip arthroplasty: impact on early postoperative care, pain management and lung function.

    Science.gov (United States)

    Wiesmann, T; Steinfeldt, T; Wagner, G; Wulf, H; Schmitt, J; Zoremba, M

    2014-01-01

    Peripheral regional anesthesia is beneficial in the management of postoperative pain in hip surgery, and can also reduce post-operative care unit (PACU) stay. Its opioid-sparing actions may also be beneficial for respiratory mechanics and pulmonary function. The aim of our pilot study was to evaluate the effect of a supplemental single shot femoral block for elective total hip arthroplasty on early respiratory function and postoperative management within the first 24 postoperative hours. We prospectively studied 80 patients undergoing total hip arthroplasty. Written informed consent was obtained after ethics committee approval. Forty patients were randomLy assigned to receive single shot femoral nerve block (FNB) using 15mL bupivacaine 0.25% and 20 mg clonidine while the remainder received standard treatment without nerve block (STN). Premedication and general anesthesia were standardized. Pulse oximetry saturation and spirometric lung function were measured preoperatively (baseline) and at 0.5 h, 2 h, 6 h and 24 h, after extubation breathing room air. PACU surveillance and postoperative pain therapy was standardized. Oxygen saturation and spirometry results were significantly better within the FNB group during the first six postoperative hours. Although VAS scores during the PACU stay did not significantly differ between the study groups, PACU discharge criteria were met earlier in the FNB group (116±40 min [mean±SD] vs. 152±47 min in the STN group). The FNB group exhibited significantly lower VAS scores at 6 and 24 hours. Supplemental single shot femoral nerve block for total hip arthroplasty resulted in earlier PACU discharge capability, improved lung function during the first six hours and better pain control within the first 24 postoperative hours.

  10. Comparison of MI, Chromocult®coliform, and Compass CC chromogenic culture-based methods to detect Escherichia coli and total coliforms in water using 16S rRNA sequencing for colony identification.

    Science.gov (United States)

    Maheux, Andrée F; Bouchard, Sébastien; Bérubé, Ève; Bergeron, Michel G

    2017-06-01

    The MI, Chromocult ® coliform, and Compass CC chromogenic culture-based methods used to assess water quality by the detection of Escherichia coli and total coliforms were compared in terms of their specificity and sensitivity, using 16S rRNA sequencing for colony identification. A sewage water sample was divided in 2-μL subsamples for testing by all three culture-based methods. All growing colonies were harvested and subjected to 16S rRNA sequencing. Test results showed that all E. coli colonies were correctly identified by all three methods, for a specificity and a sensitivity of 100%. However, for the total coliform detection, the MI agar, Chromocult ® coliform agar, and Compass CC agar were specific for only 69.2% (9/13), 47.2% (25/53), and 40.5% (17/42), whereas sensitive for 97.8% (45/46), 97.5% (39/40), and 85.7% (24/28), respectively. Thus, given the low level of specificity of these methods for the detection of total coliforms, confirming the identity of total coliform colonies could help to take public health decisions, in particular for cities connected to a public drinking water distribution system since the growth of few putative total coliform colonies on chromogenic agar is problematic and can lead to unnecessary and costly boiling notices from public health authorities.

  11. Xenopus Xp54 and human RCK/p54 helicases functionally replace yeast Dhh1p in brome mosaic virus RNA replication.

    Science.gov (United States)

    Alves-Rodrigues, Isabel; Mas, Antonio; Díez, Juana

    2007-04-01

    By using a Brome mosaic virus (BMV)-Saccharomyces cerevisiae system, we previously showed that the cellular Lsm1p-7p/Pat1p/Dhh1p decapping-activator complex functions in BMV RNA translation and replication. As a first approach in investigating whether the corresponding human homologues play a similar role, we expressed human Lsm1p (hLsm1p) and RCK/p54 in yeast. Expression of RCK/p54 but not hLsm1p restored the defect in BMV RNA translation and replication observed in the dhh1Delta and lsm1Delta strains, respectively. This functional conservation, together with the common replication strategies of positive-stranded RNA viruses, suggests that RCK/p54 may also play a role in the replication of positive-stranded RNA viruses that infect humans.

  12. The functional role of long non-coding RNA in digestive system carcinomas.

    Science.gov (United States)

    Wang, Guang-Yu; Zhu, Yuan-Yuan; Zhang, Yan-Qiao

    2014-09-01

    In recent years, long non-coding RNAs (lncRNAs) are emerging as either oncogenes or tumor suppressor genes. Recent evidences suggest that lncRNAs play a very important role in digestive system carcinomas. However, the biological function of lncRNAs in the vast majority of digestive system carcinomas remains unclear. Recently, increasing studies has begun to explore their molecular mechanisms and regulatory networks that they are implicated in tumorigenesis. In this review, we highlight the emerging functional role of lncRNAs in digestive system carcinomas. It is becoming clear that lncRNAs will be exciting and potentially useful for diagnosis and treatment of digestive system carcinomas, some of these lncRNAs might function as both diagnostic markers and the treatment targets of digestive system carcinomas.

  13. Importance of exosome depletion protocols to eliminate functional and RNA-containing extracellular vesicles from fetal bovine serum

    Directory of Open Access Journals (Sweden)

    Ganesh Vilas Shelke

    2014-09-01

    Full Text Available Extracellular vesicles (EVs, including the nano-sized exosomes, have the capacity to transfer multiple functional molecules between cells. In cell culture experiments, fetal bovine serum (FBS is often used to supplement cell culture medium as a nutrient, but it is important to know that the FBS also contain significant quantities of EVs. The aim of the current study was to determine whether the FBS EVs can influence cultured cell phenotype, and secondly to determine the efficiency of FBS-EV elimination protocols. Firstly, FBS that had not been depleted of EVs induced a migratory phenotype in a lung cancer epithelial cell line (A549 cells, an effect that could be mimicked by isolated FBS EVs alone. FBS-derived EVs also contained RNA, which was protected from consecutive proteinase K and RNase A treatment. Comparison of common isolation protocols suggested that an 18-hour centrifugation period eliminates approximately 95% of RNA-containing FBS EVs, whereas a 1.5-hour protocol is insufficient. In conclusion, this study shows that FBS EVs substantially influence cultured cell behaviour, but also that they can be virtually removed by an 18-hour ultracentrifugation protocol.

  14. Structure and Functional Characterization of the RNA-Binding Element of the NLRX1 Innate Immune Modulator

    Energy Technology Data Exchange (ETDEWEB)

    Hong, Minsun; Yoon, Sung-il; Wilson, Ian A. (Scripps)

    2012-06-20

    Mitochondrial NLRX1 is a member of the family of nucleotide-binding domain and leucine-rich-repeat-containing proteins (NLRs) that mediate host innate immunity as intracellular surveillance sensors against common molecular patterns of invading pathogens. NLRX1 functions in antiviral immunity, but the molecular mechanism of its ligand-induced activation is largely unknown. The crystal structure of the C-terminal fragment (residues 629975) of human NLRX1 (cNLRX1) at 2.65 {angstrom} resolution reveals that cNLRX1 consists of an N-terminal helical (LRRNT) domain, central leucine-rich repeat modules (LRRM), and a C-terminal three-helix bundle (LRRCT). cNLRX1 assembles into a compact hexameric architecture that is stabilized by intersubunit and interdomain interactions of LRRNT and LRRCT in the trimer and dimer components of the hexamer, respectively. Furthermore, we find that cNLRX1 interacts directly with RNA and supports a role for NLRX1 in recognition of intracellular viral RNA in antiviral immunity.

  15. Cas9/sgRNA-based genome editing and other reverse genetic approaches for functional genomic studies in rice.

    Science.gov (United States)

    Moin, Mazahar; Bakshi, Achala; Madhav, M S; Kirti, P B

    2018-03-22

    One of the important and direct ways of investigating the function of a gene is to characterize the phenotypic consequences associated with loss or gain-of-function of the corresponding gene. These mutagenesis strategies have been successfully deployed in Arabidopsis, and subsequently extended to crop species including rice. Researchers have made vast advancements in the area of rice genomics and functional genomics, as it is a diploid plant with a relatively smaller genome size unlike other cereals. The advent of rice genome research and the annotation of high-quality genome sequencing along with the developments in databases and computer searches have enabled the functional characterization of unknown genes in rice. Further, with the improvements in the efficiency of regeneration and transformation protocols, it has now become feasible to produce sizable mutant populations in indica rice varieties also. In this review, various mutagenesis methods, the current status of the mutant resources, limitations and strengths of insertional mutagenesis approaches and also results obtained with suitable screens for stress tolerance in rice are discussed. In addition, targeted genome editing using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) or Cas9/single-guide RNA system and its potential applications in generating transgene-free rice plants through genome engineering as an efficient alternative to classical transgenic technology are also discussed.

  16. Thigh and knee circumference, knee-extension strength, and functional performance after fast-track total hip arthroplasty

    DEFF Research Database (Denmark)

    Holm, Bente; Kristensen, Morten Tange; Husted, Henrik

    2011-01-01

    OBJECTIVE: To (1) quantify changes in knee-extension strength and functional-performance at discharge after fast-track total hip arthroplasty (THA) and (2) investigate whether these changes correlate to changes in thigh and knee circumference (ie, swelling) or pain. DESIGN: A prospective, descrip......OBJECTIVE: To (1) quantify changes in knee-extension strength and functional-performance at discharge after fast-track total hip arthroplasty (THA) and (2) investigate whether these changes correlate to changes in thigh and knee circumference (ie, swelling) or pain. DESIGN: A prospective......, descriptive, hypothesis-generating study. SETTING: A special unit for fast-track hip and knee arthroplasty operations at a university hospital. PARTICIPANTS: Twenty-four patients (20 women and 4 men; ages 69 ± 6.1 years) scheduled for primary unilateral THA. METHODS: All patients were evaluated before surgery...... and on the day of hospital discharge. MAIN OUTCOME MEASURES: Knee-extension strength, thigh and knee joint circumference, hip pain, and functional performance (Timed Up & Go, 30-Second Chair Stand, and 10-Meter Walk tests). RESULTS: All investigated variables changed significantly from before to after surgery...

  17. Alteration of protein function by a silent polymorphism linked to tRNA abundance

    NARCIS (Netherlands)

    Kirchner, Sebastian; Cai, Zhiwei; Rauscher, Robert; Kastelic, Nicolai; Anding, Melanie; Czech, Andreas; Kleizen, Bertrand; Ostedgaard, Lynda S.; Braakman, Ineke; Sheppard, David N.; Ignatova, Zoya

    2017-01-01

    Synonymous single nucleotide polymorphisms (sSNPs) are considered neutral for protein function, as by definition they exchange only codons, not amino acids. We identified an sSNP that modifies the local translation speed of the cystic fibrosis transmembrane conduc-tance regulator (CFTR), leading to

  18. Early days of tRNA research: Discovery, function, purification and ...

    Indian Academy of Sciences (India)

    Madhu

    2006-10-04

    Oct 4, 2006 ... acids attached to the tRNAs are transferred to protein. The. tRNAs turned out to be much larger than the small molecules proposed by Crick to function as adaptors, in his well known albeit unpublished “Adaptor Hypothesis” designed to explain how DNA sequences might be translated to amino.

  19. Indocyanine green fluorescence angiography for quantitative evaluation of in situ parathyroid gland perfusion and function after total thyroidectomy.

    Science.gov (United States)

    Lang, Brian Hung-Hin; Wong, Carlos K H; Hung, Hing Tsun; Wong, Kai Pun; Mak, Ka Lun; Au, Kin Bun

    2017-01-01

    Because the fluorescent light intensity on an indocyanine green fluorescence angiography reflects the blood perfusion within a focused area, the fluorescent light intensity in the remaining in situ parathyroid glands may predict postoperative hypocalcemia risk after total thyroidectomy. Seventy patients underwent intraoperative indocyanine green fluorescence angiography after total thyroidectomy. Any parathyroid glands with a vascular pedicle was left in situ while any parathyroid glands without pedicle or inadvertently removed was autotransplanted. After total thyroidectomy, an intravenous 2.5 mg indocyanine green fluorescence angiography was given and real-time fluorescent images of the thyroid bed were recorded using the SPY imaging system (Novadaq, Ontario, Canada). The fluorescent light intensity of each indocyanine green fluorescence angiography as well as the average and greatest fluorescent light intensity in each patient were calculated. Postoperative hypocalcemia was defined as adjusted calcium 150% developed postoperative hypocalcemia while 9 (81.8%) patients with a greatest fluorescent light intensity ≤150% did. Similarly, no patients with an average fluorescent light intensity >109% developed PH while 9 (30%) with an average fluorescent light intensity ≤109% did. The greatest fluorescent light intensity was more predictive than day-0 postoperative hypocalcemia (P = .027) and % PTH drop day-0 to 1 (P parathyroid glands function and predicting postoperative hypocalcemia risk after total thyroidectomy. Copyright © 2016 Elsevier Inc. All rights reserved.

  20. The use of clinical analysis of movements in evaluation of motor functional status of patients after total hip replacement

    Directory of Open Access Journals (Sweden)

    Romakina N.A.

    2016-06-01

    Full Text Available Aim: to estimate functional status of coxarthrosis in patients requiring total hip replacement of the two hip joints. Material and methods. The biomechanical examination of 94 patients with bilateral primary coxarthrosis before and after total hip replacement was performed using clinical stabilometric software complex. The ability to perceive the mechanical load during standing and walking was evaluated at different stages of the treatment. The difference between the samples was estimated with the use of Mann — Whitney U-test. The rank correlation of biomechanical parameters was measured by Spearman coefficient. Results. It was revealed that the most responsive indicators are the transfer period, the first and second periods of double support and the deviation of the center of pressure relative to the average position in the frontal plane. Conclusion. After surgical treatment there was observed some improvement: the-left-and-right-step asymmetry decrease, rhythm rate increase, improved ability to maintain body balance, jog reactions increase.

  1. Diversity of total and functional microbiome of anammox reactors fed with complex and synthetic nitrogen-rich wastewaters

    DEFF Research Database (Denmark)

    Gülay, Arda; Pellicer i Nàcher, Carles; Mutlu, Ayten Gizem

    implementations treating complex nitrogen-rich wastewaters and 14 were lab-scale implementations treating synthetic wastewaters. We found that nitritation/anammox bioreactors treating complex nitrogen-rich wastewaters were more diverse in terms of total microbial diversity but less diverse at anammox functional...... diversity than the bioreactors treating synthetic wastewaters inferred from observed OTUs0.03, Chao1, Shannon index and Phylogenetic distance calculations. Differences in total microbial diversity agreed with the ecological theory concerning the positive correlation between substrate complexity...... weighted UniFrac algorithm explained 29% of the variance where the bioreactor samples of complex nitrogen-rich wastewater feeding was clearly separated from the bioreactor samples of synthetic feeding. Here we examined and compared for the first time microbial diversity of nitritation-anammox reactors...

  2. New method to determine the total carbonyl functional group content in extractable particulate organic matter by tandem mass spectrometry.

    Science.gov (United States)

    Dron, J; Zheng, W; Marchand, N; Wortham, H

    2008-08-01

    A functional group analysis method was developed to determine the quantitative content of carbonyl functional groups in atmospheric particulate organic matter (POM) using constant neutral loss scanning-tandem mass spectrometry (CNLS-MS/MS). The neutral loss method consists in monitoring the loss of a neutral fragment produced by the fragmentation of a precursor ion in a collision cell. The only ions detected are the daughter ions resulting from the loss of the neutral fragment under study. Then, scanning the loss of a neutral fragment characteristic of a functional group enables the selective detection of the compounds bearing the chemical function under study within a complex mixture. The selective detection of carbonyl functional groups was achieved after derivatization with pentafluorophenylhydrazine (PFPH) by monitoring the neutral loss of C(6)F(5)N (181 amu), which was characteristic of a large panel of derivatized carbonyl compounds. The method was tested on 25 reference mixtures of different composition, all containing 24 carbonyl compounds at randomly determined concentrations. The repeatability and calibration tests were satisfying as they resulted in a relative standard deviation below 5% and a linear range between 0.01 and 0.65 mM with a calculated detection limit of 0.0035 mM. Also, the relative deviation induced by changing the composition of the mixture while keeping the total concentration of carbonyl functional groups constant was less than 20%. These reliability experiments demonstrate the high robustness of the developed procedure for accurate carbonyl functional group measurement, which was applied to atmospheric POM samples. Copyright (c) 2008 John Wiley & Sons, Ltd.

  3. Circulating microRNA levels predict residual beta cell function and glycaemic control in children with type 1 diabetes mellitus

    DEFF Research Database (Denmark)

    Samandari, Nasim; Mirza, Aashiq H.; Nielsen, Lotte B.

    2017-01-01

    from the Danish Remission Phase Cohort, and profiled for miRNAs. At the same time points, meal-stimulated C-peptide and HbA1c levels were measured and insulin-dose adjusted HbA1c (IDAA1c) calculated. miRNAs that at 3 months after diagnosis predicted residual beta cell function and glycaemic control...... in this subgroup were further validated in the remaining cohort (n = 83). Statistical analysis of miRNA prediction for disease progression was performed by multiple linear regression analysis adjusted for age and sex. Results: In the discovery analysis, six miRNAs (hsa-miR-24-3p, hsa-miR-146a-5p, hsa-miR-194-5p...

  4. Multifunctional G-rich and RRM-containing domains of TbRGG2 perform separate yet essential functions in trypanosome RNA editing.

    Science.gov (United States)

    Foda, Bardees M; Downey, Kurtis M; Fisk, John C; Read, Laurie K

    2012-09-01

    Efficient editing of Trypanosoma brucei mitochondrial RNAs involves the actions of multiple accessory factors. T. brucei RGG2 (TbRGG2) is an essential protein crucial for initiation and 3'-to-5' progression of editing. TbRGG2 comprises an N-terminal G-rich region containing GWG and RG repeats and a C-terminal RNA recognition motif (RRM)-containing domain. Here, we perform in vitro and in vivo separation-of-function studies to interrogate the mechanism of TbRGG2 action in RNA editing. TbRGG2 preferentially binds preedited mRNA in vitro with high affinity attributable to its G-rich region. RNA-annealing and -melting activities are separable, carried out primarily by the G-rich and RRM domains, respectively. In vivo, the G-rich domain partially complements TbRGG2 knockdown, but the RRM domain is also required. Notably, TbRGG2's RNA-melting activity is dispensable for RNA editing in vivo. Interactions between TbRGG2 and MRB1 complex proteins are mediated by both G-rich and RRM-containing domains, depending on the binding partner. Overall, our results are consistent with a model in which the high-affinity RNA binding and RNA-annealing activities of the G-rich domain are essential for RNA editing in vivo. The RRM domain may have key functions involving interactions with the MRB1 complex and/or regulation of the activities of the G-rich domain.

  5. Development of an Innovative Intradermal siRNA Delivery System Using a Combination of a Functional Stearylated Cytoplasm-Responsive Peptide and a Tight Junction-Opening Peptide

    Directory of Open Access Journals (Sweden)

    Hisako Ibaraki

    2016-09-01

    Full Text Available As a new category of therapeutics for skin diseases including atopic dermatitis (AD, nucleic acids are gaining importance in the clinical setting. Intradermal administration is noninvasive and improves patients′ quality of life. However, intradermal small interfering RNA (siRNA delivery is difficult because of two barriers encountered in the skin: intercellular lipids in the stratum corneum and tight junctions in the stratum granulosum. Tight junctions are the major barrier in AD; therefore, we focused on functional peptides to devise an intradermal siRNA delivery system for topical skin application. In this study, we examined intradermal siRNA permeability in the tape-stripped (20 times back skin of mice or AD-like skin of auricles treated with 6-carboxyfluorescein-aminohexyl phosphoramidite (FAM-labeled siRNA, the tight junction modulator AT1002, and the functional cytoplasm-responsive stearylated peptide STR-CH2R4H2C by using confocal laser microscopy. We found that strong fluorescence was observed deep and wide in the epidermis and dermis of back skin and AD-like ears after siRNA with STR-CH2R4H2C and AT1002 treatment. After 10 h from administration, brightness of FAM-siRNA was significantly higher for STR-CH2R4H2C + AT1002, compared to other groups. In addition, we confirmed the nontoxicity of STR-CH2R4H2C as a siRNA carrier using PAM212 cells. Thus, our results demonstrate the applicability of the combination of STR-CH2R4H2C and AT1002 for effective intradermal siRNA delivery.

  6. Acute Total and Chronic Partial Sleep Deprivation: Effects on Neurobehavioral Functions, Waking EEG and Renin-Angiotensin System

    Science.gov (United States)

    Dijk, Derk-Jan

    1999-01-01

    Total sleep deprivation leads to decrements in neurobehavioral performance and changes in electroencephalographic (EEG) oscillations as well as the incidence of slow eye movements ad detected in the electro-oculogram (EOG) during wakefulness. Although total sleep deprivation is a powerful tool to investigate the association of EEG/EOG and neurobehavioral decrements, sleep loss during space flight is usual only partial. Furthermore exposure to the microgravity environment leads to changes in sodium and volume homeostasis and associated renal and cardio-endocrine responses. Some of these changes can be induced in head down tilt bedrest studies. We integrate research tools and research projects to enhance the fidelity of the simulated conditions of space flight which are characterized by complexity and mutual interactions. The effectiveness of countermeasures and physiologic mechanisms underlying neurobehavioral changes and renal-cardio endocrine changes are investigated in Project 3 of the Human Performance Team and Project 3 of the Cardiovascular Alterations Team respectively. Although the. specific aims of these two projects are very different, they employ very similar research protocols. Thus, both projects investigate the effects of posture/bedrest and sleep deprivation (total or partial) on outcome measures relevant to their specific aims. The main aim of this enhancement grant is to exploit the similarities in research protocols by including the assessment of outcome variables relevant to the Renal-Cardio project in the research protocol of Project 3 of the Human Performance Team and by including the assessment of outcome variables relevant to the Quantitative EEG and Sleep Deprivation Project in the research protocols of Project 3 of the Cardiovascular Alterations team. In particular we will assess Neurobehavioral Function and Waking EEG in the research protocols of the renal-cardio endocrine project and renin-angiotensin and cardiac function in the research

  7. Early life ozone exposure results in dysregulated innate immune function and altered microRNA expression in airway epithelium.

    Directory of Open Access Journals (Sweden)

    Candice C Clay

    Full Text Available Exposure to ozone has been associated with increased incidence of respiratory morbidity in humans; however the mechanism(s behind the enhancement of susceptibility are unclear. We have previously reported that exposure to episodic ozone during postnatal development results in an attenuated peripheral blood cytokine response to lipopolysaccharide (LPS that persists with maturity. As the lung is closely interfaced with the external environment, we hypothesized that the conducting airway epithelium of neonates may also be a target of immunomodulation by ozone. To test this hypothesis, we evaluated primary airway epithelial cell cultures derived from juvenile rhesus macaque monkeys with a prior history of episodic postnatal ozone exposure. Innate immune function was measured by expression of the proinflammatory cytokines IL-6 and IL-8 in primary cultures established following in vivo LPS challenge or, in response to in vitro LPS treatment. Postnatal ozone exposure resulted in significantly attenuated IL-6 mRNA and protein expression in primary cultures from juvenile animals; IL-8 mRNA was also significantly reduced. The effect of antecedent ozone exposure was modulated by in vivo LPS challenge, as primary cultures exhibited enhanced cytokine expression upon secondary in vitro LPS treatment. Assessment of potential IL-6-targeting microRNAs miR-149, miR-202, and miR-410 showed differential expression in primary cultures based upon animal exposure history. Functional assays revealed that miR-149 is capable of binding to the IL-6 3' UTR and decreasing IL-6 protein synthesis in airway epithelial cell lines. Cumulatively, our findings suggest that episodic ozone during early life contributes to the molecular programming of airway epithelium, such that memory from prior exposures is retained in the form of a dysregulated IL-6 and IL-8 response to LPS; differentially expressed microRNAs such as miR-149 may play a role in the persistent modulation of the

  8. The Ever-Evolving Concept of the Gene: The Use of RNA/Protein Experimental Techniques to Understand Genome Functions

    Directory of Open Access Journals (Sweden)

    Andrea Cipriano

    2018-03-01

    Full Text Available The completion of the human genome sequence together with advances in sequencing technologies have shifted the paradigm of the genome, as composed of discrete and hereditable coding entities, and have shown the abundance of functional noncoding DNA. This part of the genome, previously dismissed as “junk” DNA, increases proportionally with organismal complexity and contributes to gene regulation beyond the boundaries of known protein-coding genes. Different classes of functionally relevant nonprotein-coding RNAs are transcribed from noncoding DNA sequences. Among them are the long noncoding RNAs (lncRNAs, which are thought to participate in the basal regulation of protein-coding genes at both transcriptional and post-transcriptional levels. Although knowledge of this field is still limited, the ability of lncRNAs to localize in different cellular compartments, to fold into specific secondary structures and to interact with different molecules (RNA or proteins endows them with multiple regulatory mechanisms. It is becoming evident that lncRNAs may play a crucial role in most biological processes such as the control of development, differentiation and cell growth. This review places the evolution of the concept of the gene in its historical context, from Darwin's hypothetical mechanism of heredity to the post-genomic era. We discuss how the original idea of protein-coding genes as unique determinants of phenotypic traits has been reconsidered in light of the existence of noncoding RNAs. We summarize the technological developments which have been made in the genome-wide identification and study of lncRNAs and emphasize the methodologies that have aided our understanding of the complexity of lncRNA-protein interactions in recent years.

  9. Comparative Analyses of MicroRNA Microarrays during Cardiogenesis: Functional Perspectives

    OpenAIRE

    Bonet, Fernando; Hernandez-Torres, Francisco; Esteban, Franciso J.; Aranega, Amelia; Franco, Diego

    2013-01-01

    Cardiovascular development is a complex process in which several transcriptional pathways are operative, providing instructions to the developing cardiomyocytes, while coping with contraction and morphogenetic movements to shape the mature heart. The discovery of microRNAs has added a new layer of complexity to the molecular mechanisms governing the formation of the heart. Discrete genetic ablation of the microRNAs processing enzymes, such as Dicer and Drosha, has highlighted the functional r...

  10. Microbial community functioning at hypoxic sediments revealed by targeted metagenomics and RNA stable isotope probing

    OpenAIRE

    Pavloudi, Christina

    2017-01-01

    Microorganisms are instrumental to the structure and functioning of marine ecosystems and to the chemistry of the ocean due to their essential part in the cycling of the elements and in the recycling of the organic matter. Two of the most critical ocean biogeochemical cycles are those of nitrogen and sulfur, since they can influence the synthesis of nucleic acids and proteins, primary productivity and microbial community structure. Oxygen concentration in marine environments is one of the env...

  11. The tumor suppressive role of eIF3f and its function in translation inhibition and rRNA degradation.

    Directory of Open Access Journals (Sweden)

    Fushi Wen

    Full Text Available Deregulated translation plays an important role in human cancer. We previously reported decreased eukaryotic initiation factor 3 subunit f (eIF3f expression in pancreatic cancer. Whether decreased eIF3f expression can transform normal epithelial cells is not known. In our current study, we found evidence that stable knockdown of eIF3f in normal human pancreatic ductal epithelial cells increased cell size, nuclear pleomorphism, cytokinesis defects, cell proliferation, clonogenicity, apoptotic resistance, migration, and formation of 3-dimensional irregular masses. Our findings support the tumor suppressive role of eIF3f in pancreatic cancer. Mechanistically, we found that eIF3f inhibited both cap-dependent and cap-independent translation. An increase in the ribosomal RNA (rRNA level was suggested to promote the generation of cancer. The regulatory mechanism of rRNA degradation in mammals is not well understood. We demonstrated here that eIF3f promotes rRNA degradation through direct interaction with heterogeneous nuclear ribonucleoprotein (hnRNP K. We showed that hnRNP K is required for maintaining rRNA stability: under stress conditions, eIF3f dissociates hnRNP K from rRNA, thereby preventing it from protecting rRNA from degradation. We also demonstrated that rRNA degradation occurred in non-P body, non-stress granule cytoplasmic foci that contain eIF3f. Our findings established a new mechanism of rRNA decay regulation mediated by hnRNP K/eIF3f and suggest that the tumor suppressive function of eIF3f may link to impaired rRNA degradation and translation.

  12. Possible effects of mobilisation on acute post-operative pain and nociceptive function after total knee arthroplasty

    DEFF Research Database (Denmark)

    Lunn, T H; Kristensen, B B; Gaarn-Larsen, L

    2012-01-01

    BACKGROUND: Experimental studies in animals, healthy volunteers, and patients with chronic pain suggest exercise to provide analgesia in several types of pain conditions and after various nociceptive stimuli. To our knowledge, there is no data on the effects of exercise on pain and nociceptive...... function in surgical patients despite early mobilisation being an important factor to enhance recovery. We therefore investigated possible effects of mobilisation on post-operative pain and nociceptive function after total knee arthroplasty (TKA). METHODS: Thirty patients undergoing TKA under standardised...... anaesthesia and analgesia underwent an exercise (mobilisation) strategy on the first post-operative morning consisting of 25-m walking twice, with a 20-min interval. Pain was assessed at rest and during passive hip and knee flexion before, and 5 and 20 min after walk, as well as during walk. Nociceptive...

  13. A daily adjustable progressive resistance exercise protocol and functional training to increase quadriceps muscle strength and functional performance in an elderly homebound patient following a total knee arthroplasty.

    Science.gov (United States)

    Ardali, Gunay

    2014-05-01

    There is no routinely prescribed protocol to address quadriceps weakness and functional impairments following a total knee arthroplasty (TKA). The purpose of this case report is to introduce and describe the early use of a daily adjustable progressive resistance exercise (DAPRE) protocol as an adjunct to standard rehabilitation to maximize quadriceps muscle strength and functional performance in an elderly homebound patient following a TKA. A 61-year-old female was referred to home care physical therapy for 6 weeks following left TKA due to functional deficits and inability to activate the weak left quadriceps muscle. In phase I, the patient received three visits with emphasis on edema management, improving left knee range of motion, and reducing pain. Phase II consisted of two main components: (1) a DAPRE protocol aimed at maximizing the quadriceps strength and (2) functional training aimed at improving normal gait patterns, transfers, and dynamic balance. The patient made substantial improvements in both quadriceps muscle strength and functional performance in the first seven weeks following the TKA. The patient had a pain free return to daily living activities. The results suggest that early initiation of a DAPRE protocol was free of adverse events and improved quadriceps strength and functional performance for this patient.

  14. Hydro-isomerization of n-hexane on bi-functional catalyst: Effect of total and hydrogen partial pressures

    Science.gov (United States)

    Thoa, Dao Thi Kim; Loc, Luu Cam

    2017-09-01

    The effect of both total pressure and hydrogen partial pressure during n-hexane hydro-isomerization over platinum impregnated on HZSM-5 was studied. n-Hexane hydro-isomerization was conducted at atmospheric pressure and 0.7 MPa to observe the influence of total pressure. In order to see the effect of hydrogen partial pressure, the reaction was taken place at different partial pressure of hydrogen varied from 307 hPa to 718 hPa by dilution with nitrogen to keep the total pressure at 0.1 MPa. Physico-chemical characteristics of catalyst were determined by the methods of nitrogen physi-sorption BET, SEM, XRD, TEM, NH3-TPD, TPR, and Hydrogen Pulse Chemi-sorption. Activity of catalyst in the hydro-isomerization of n-hexane was studied in a micro-flow reactor in the temperature range of 225-325 °C; the molar ratio H2/ hydrocarbon: 5.92, concentration of n-hexane: 9.2 mol.%, GHSV 2698 h-1. The obtained catalyst expressed high acid density, good reducing property, high metal dispersion, and good balance between metallic and acidic sites. It is excellent contact for n-hexane hydro-isomerization. At 250 °C, n-hexane conversion and selectivity were as high as 59-76 % and 85-99 %, respectively. It was found that catalytic activity was promoted either by total pressure or hydrogen partial pressure. At total pressure of 0.7 MPa while hydrogen partial pressure of 718 hPa, catalyst produced 63 RON liquid product containing friendly environmental iso-paraffins which is superior blending stock for green gasoline. Hydrogen did not only preserve catalyst actives by depressing hydrocracking and removing coke precursors but also facilitated hydride transfer step in the bi-functional bi-molecular mechanism.

  15. Functional substitution of a eukaryotic glycyl-tRNA synthetase with an evolutionarily unrelated bacterial cognate enzyme.

    Directory of Open Access Journals (Sweden)

    Chin-I Chien

    Full Text Available Two oligomeric types of glycyl-tRNA synthetase (GlyRS are found in nature: a α2 type and a α2β2 type. The former has been identified in all three kingdoms of life and often pairs with tRNAGly that carries an A73 discriminator base, while the latter is found only in bacteria and chloroplasts and is almost always coupled with tRNAGly that contains U73. In the yeast Saccharomyces cerevisiae, a single GlyRS gene, GRS1, provides both the cytoplasmic and mitochondrial functions, and tRNAGly isoacceptors in both compartments possess A73. We showed herein that Homo sapiens and Arabidopsis thaliana cytoplasmic GlyRSs (both α2-type enzymes can rescue both the cytoplasmic and mitochondrial defects of a yeast grs1- strain, while Escherichia coli GlyRS (a α2β2-type enzyme and A. thaliana organellar GlyRS (a (αβ2-type enzyme failed to rescue either defect of the yeast mull allele. However, a head-to-tail αβ fusion of E. coli GlyRS effectively supported the mitochondrial function. Our study suggests that a α2-type eukaryotic GlyRS may be functionally substituted with a α2β2-type bacterial cognate enzyme despite their remote evolutionary relationships.

  16. Functional Diets Modulate lncRNA-Coding RNAs and Gene Interactions in the Intestine of Rainbow Trout Oncorhynchus mykiss.

    Science.gov (United States)

    Núñez-Acuña, Gustavo; Détrée, Camille; Gallardo-Escárate, Cristian; Gonçalves, Ana Teresa

    2017-06-01

    The advent of functional genomics has sparked the interest in inferring the function of non-coding regions from the transcriptome in non-model species. However, numerous biological processes remain understudied from this perspective, including intestinal immunity in farmed fish. The aim of this study was to infer long non-coding RNA (lncRNAs) expression profiles in rainbow trout (Oncorhynchus mykiss) fed for 30 days with functional diets based on pre- and probiotics. For this, whole transcriptome sequencing was conducted through Illumina technology, and lncRNAs were mined to evaluate transcriptional activity in conjunction with known protein sequences. To detect differentially expressed transcripts, 880 novels and 9067 previously described O. mykiss lncRNAs were used. Expression levels and genome co-localization correlations with coding genes were also analyzed. Significant differences in gene expression were primarily found in the probiotic diet, which had a twofold downregulation of lncRNAs compared to other treatments. Notable differences by diet were also evidenced between the coding genes of distinct metabolic processes. In contrast, genome co-localization of lncRNAs with coding genes was similar for all diets. This study contributes novel knowledge regarding lncRNAs in fish, suggesting key roles in salmons fed with in-feed additives with the capacity to modulate the intestinal homeostasis and host health.

  17. Long noncoding RNA MALAT1 regulates endothelial cell function and vessel growth.

    Science.gov (United States)

    Michalik, Katharina M; You, Xintian; Manavski, Yosif; Doddaballapur, Anuradha; Zörnig, Martin; Braun, Thomas; John, David; Ponomareva, Yuliya; Chen, Wei; Uchida, Shizuka; Boon, Reinier A; Dimmeler, Stefanie

    2014-04-25

    The human genome harbors a large number of sequences encoding for RNAs that are not translated but control cellular functions by distinct mechanisms. The expression and function of the longer transcripts namely the long noncoding RNAs in the vasculature are largely unknown. Here, we characterized the expression of long noncoding RNAs in human endothelial cells and elucidated the function of the highly expressed metastasis-associated lung adenocarcinoma transcript 1 (MALAT1). Endothelial cells of different origin express relative high levels of the conserved long noncoding RNAs MALAT1, taurine upregulated gene 1 (TUG1), maternally expressed 3 (MEG3), linc00657, and linc00493. MALAT1 was significantly increased by hypoxia and controls a phenotypic switch in endothelial cells. Silencing of MALAT1 by small interfering RNAs or GapmeRs induced a promigratory response and increased basal sprouting and migration, whereas proliferation of endothelial cells was inhibited. When angiogenesis was further stimulated by vascular endothelial growth factor, MALAT1 small interfering RNAs induced discontinuous sprouts indicative of defective proliferation of stalk cells. In vivo studies confirmed that genetic ablation of MALAT1 inhibited proliferation of endothelial cells and reduced neonatal retina vascularization. Pharmacological inhibition of MALAT1 by GapmeRs reduced blood flow recovery and capillary density after hindlimb ischemia. Gene expression profiling followed by confirmatory quantitative reverse transcriptase-polymerase chain reaction demonstrated that silencing of MALAT1 impaired the expression of various cell cycle regulators. Silencing of MALAT1 tips the balance from a proliferative to a migratory endothelial cell phenotype in vitro, and its genetic deletion or pharmacological inhibition reduces vascular growth in vivo.

  18. Novel Functions of MicroRNA-17-92 Cluster in the Endocrine System.

    Science.gov (United States)

    Wan, Shan; Chen, Xiang; He, Yuedong; Yu, Xijie

    2018-01-01

    MiR-17-92 cluster is coded by MIR17HG in chromosome 13, which is highly conserved in vertebrates. Published literatures have proved that miR-17-92 cluster critically regulates tumorigenesis and metastasis. Recent researches showed that the miR-17-92 cluster also plays novel functions in the endocrine system. To summarize recent findings on the physiological and pathological roles of miR-17-92 cluster in bone, lipid and glucose metabolisms. MiR-17-92 cluster plays significant regulatory roles in bone development and metabolism through regulating the differentiation and function of osteoblasts and osteoclasts. In addition, miR-17- 92 cluster is nearly involved in every aspect of lipid metabolism. Last but not the least, the miR-17-92 cluster is closely bound up with pancreatic beta cell function, development of type 1 diabetes and insulin resistance. However, whether miR-17-92 cluster is involved in the communication among bone, fat and glucose metabolisms remains unknown. Growing evidence indicates that miR-17-92 cluster plays significant roles in bone, lipid and glucose metabolisms through a variety of signaling pathways. Fully understanding its modulating mechanisms may necessarily facilitate to comprehend the clinical and molecule features of some metabolic disorders such as osteoporosis, arthrosclerosis and diabetes mellitus. It may provide new drug targets to prevent and cure these disorders. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  19. Emerging role of non-coding RNA in neural plasticity, cognitive function, and neuropsychiatric disorders

    Directory of Open Access Journals (Sweden)

    Paola eSpadaro

    2012-07-01

    Full Text Available Non-coding RNAs have emerged as critical regulators of transcription, epigenetic processes, and gene silencing, which make them ideal candidates for insight into molecular evolution and a better understanding of the molecular pathways of neuropsychiatric disease. Here we provide an overview of the current state of knowledge regarding various classes of ncRNAs and their role in neural plasticity and cognitive function, and highlight the potential contribution they may make to the development of a variety of neuropsychiatric disorders, including schizophrenia, addiction and fear-related anxiety disorders.

  20. Screening miRNA for Functional Significance by 3D Cell Culture System.

    Science.gov (United States)

    Han, Bo

    2018-01-01

    Cell-based assays play important roles in cell biology and drug discovery. 3D cell culture, which allows cells to grow or interact with their surrounding in all three dimensions, provides more physiological information for the in vivo tests. Here, we describe a tunable collagen-based 3D cell culture system based on collagen material crosslinked with transgluminase, to study the function of miR. Methods including gel handling, proliferation assays, gene, and protein expressions in a 3D setting are described.

  1. Effect of single base changes and the absence of modified bases in 16S RNA on the reconstitution and function of Escherichia coli 30S ribosomes

    International Nuclear Information System (INIS)

    Denman, R.; Krzyzosiak, W.; Nurse, K.; Ofengand, J.

    1987-01-01

    The gene coding for E. coli 16S rRNA was placed in pUC19 under the control of the strong class III T7 promoter, phi 10, by ligation of the 1490 bp BclI/BstEII fragment of the rrnB operon with appropriate synthetic oligodeoxynucleotides. Such constructs allowed efficient in vitro synthesis of full-length transcripts (up to 900 mol RNA/mol template) free of modified bases. The synthetic RNA could be assembled into 30S subunits upon addition of E. coli 30S ribosomal proteins. The particles co-sedimented with authentic 30S particles and were electron microscopically indistinguishable from them. Upon addition of 50S subunits, codon-dependent P-site binding of tRNA and codon-dependent polypeptide synthesis were >80% of 30S reconstituted from natural 16S RNA and >50% of isolated 30S. UV-induced crosslinking of P-site bound AcVal-tRNA to residue C 1400 was preserved. Changing C 1400 to A had little effect on reconstitution, P-site binding, or polypeptide synthesis. However, the substitution of C 1499 by G markedly inhibited assembly. The effect on P-site binding and polypeptide synthesis is under study. These results show (1) none of the modified bases of 16S RNA are essential for protein synthesis, (2) substitution of A for C 1400 has little functional effect, and (3) position 1400 may be important for ribosome assembly

  2. Prospective study comparing functional outcomes and revision rates between hip resurfacing and total hip arthroplasty: preliminary results for 2 years

    Directory of Open Access Journals (Sweden)

    Régis Pailhé

    2013-07-01

    Full Text Available There is a need of independent prospective studies about modern generation of hip resurfacing implants. The aim of this propective observational study was to compare the functional outcomes and revision rates with hip resurfacing arthroplasty and total hip arthroplasty and to present the preliminary results at 2 years. Patients included were recruited prospectively in the Partial Pelvic Replacement Hip Project by a single surgeon between January 2007 and January 2010. Patients were assessed with the Harris Hip Score (HHS and Postel-Merle d’Aubigné (MDA score and Devane Score. The end point of the study was reoperation for any cause related to the prosthesis. At a mean follow up of 38.6 months there were a total of 142 patients with hip resurfacing (group 1 [100 Durom® (Zimmer Inc., Warsaw, IN, USA and 42 Birmingham Hip Resurfacing® (Smith & Nephew, Memphis, TN, USA] and 278 patients with total hip arthroplasty (group 2. The results showed significantly greater gain of HHS, MDA and Devane score with hip resurfacing procedures. However, considering all the complications, the rate was significantly higher in group 16.4% vs 1.79% in group 2 (P<0.0001. In group 1 we observed 6 complications only concerned males with Durom® implants. The follow up of this cohort is still on going and may deliver more information on the evolution of these results in time.

  3. Pulmonary function in relation to total dust exposure at a bauxite refinery and alumina-based chemical products plant.

    Science.gov (United States)

    Townsend, M C; Enterline, P E; Sussman, N B; Bonney, T B; Rippey, L L

    1985-12-01

    A cross-sectional study of 1,142 male employees at the Arkansas Operations of a large aluminum production company examined the effect on pulmonary function of chronic exposure to total dust produced in the mining and refining of bauxite and the production of alumina chemicals. Never smokers, ex-smokers, and current smokers were analyzed separately. Among never smokers, a pattern of decreasing FEV1 was observed in relation to increasing duration and cumulative total dust exposure. Among never smokers with cumulative total dust exposures of greater than or equal to 100 mg/m3 yr and greater than or equal to 20 yr of exposure, there was a mean reduction from the predicted FEV1 of 0.29 to 0.39 L, in addition to a 3- to 4-fold excess of observed/expected numbers of subjects with FEV1 less than 80% of predicted. These results were observed relative to an external and an internal comparison group. Among current smokers, the deviations from predicted and the excess numbers of subjects with FEV1 less than 80% of predicted were larger in all exposure groups than for the never smokers. However, the quality of the smoking data was inadequate to allow separation of the effects of smoking and dust exposure.

  4. A Mitochondria-Specific Isoform of FASTK Is Present In Mitochondrial RNA Granules and Regulates Gene Expression and Function

    Directory of Open Access Journals (Sweden)

    Alexis A. Jourdain

    2015-02-01

    Full Text Available The mitochondrial genome relies heavily on post-transcriptional events for its proper expression, and misregulation of this process can cause mitochondrial genetic diseases in humans. Here, we report that a novel translational variant of Fas-activated serine/threonine kinase (FASTK co-localizes with mitochondrial RNA granules and is required for the biogenesis of ND6 mRNA, a mitochondrial-encoded subunit of the NADH dehydrogenase complex (complex I. We show that ablating FASTK expression in cultured cells and mice results specifically in loss of ND6 mRNA and reduced complex I activity in vivo. FASTK binds at multiple sites along the ND6 mRNA and its precursors and cooperates with the mitochondrial degradosome to ensure regulated ND6 mRNA biogenesis. These data provide insights into the mechanism and control of mitochondrial RNA processing within mitochondrial RNA granules.

  5. The Crystal Structure of the RNA-Dependent RNA Polymerase from Human Rhinovirus: A Dual Function Target for Common Cold Antiviral Therapy

    Energy Technology Data Exchange (ETDEWEB)

    Love, Robert A.; Maegley, Karen A.; Yu, Xiu; Ferre, RoseAnn; Lingardo, Laura K.; Diehl, Wade; Parge, Hans E.; Dragovich, Peter S.; Fuhrman, Shella A. (Pfizer)

    2010-11-16

    Human rhinoviruses (HRV), the predominant members of the Picornaviridae family of positive-strand RNA viruses, are the major causative agents of the common cold. Given the lack of effective treatments for rhinoviral infections, virally encoded proteins have become attractive therapeutic targets. The HRV genome encodes an RNA-dependent RNA polymerase (RdRp) denoted 3D{sup pol}, which is responsible for replicating the viral genome and for synthesizing a protein primer used in the replication. Here the crystal structures for three viral serotypes (1B, 14, and 16) of HRV 3D{sup pol} have been determined. The three structures are very similar to one another, and to the closely related poliovirus (PV) 3D{sup pol} enzyme. Because the reported PV crystal structure shows significant disorder, HRV 3D{sup pol} provides the first complete view of a picornaviral RdRp. The folding topology of HRV 3D{sup pol} also resembles that of RdRps from hepatitis C virus (HCV) and rabbit hemorrhagic disease virus (RHDV) despite very low sequence homology.

  6. Implication of Ccr4-Not complex function in mRNA quality control in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Assenholt, Jannie; Mouaikel, John; Saguez, Cyril

    2011-01-01

    RNPs are exported to the cytoplasm. The Ccr4-Not complex, which constitutes the major S. cerevisiae cytoplasmic deadenylase, has recently been implied in nuclear exosome–related processes. Consistent with a possible nuclear function of the complex, the deletion or mutation of Ccr4-Not factors also elicits...... transcription phenotypes. Here we use genetic depletion of the Mft1p protein of the THO transcription/mRNP packaging complex as a model system to link the Ccr4-Not complex to nuclear mRNP QC. We reveal strong genetic interactions between alleles of the Ccr4-Not complex with both the exosomal RRP6 and MFT1 genes....... Moreover, Rrp6p-dependent in vivo QC phenotypes of Δmft1 cells can be rescued by codeletion of several Ccr4-Not components. We discuss how the Ccr4-Not complex may connect with the mRNP QC pathway...

  7. Quantification of thymidine kinase (TK1) mRNA in normal and leukemic cells and investigation of structure-function relatiosnhip of recombinant TK1enzyme

    DEFF Research Database (Denmark)

    Kristensen, Tina

    Thymidine kinase (TK) catalyses the ATP-dependent phosphorylation of thymidine to thymidine monophosphate, which is subsequency phosphorylated to thymidine triphosphate and utilized for DNA synthesis. Human cytosolic TK (TKI) is cell cycle regulated, e.g. the TK1 activity increases sharply at the G...... patients with chronic lymphatic leukemia (CLL). 2: Structure-function relationship of recombinant TKI. In the first part a sensitive method (competitive PCR) for quantification of TKI mRNA was established. The TKI mRNA level was quantified in quiescent lymphocytes from control donors (n = 6...... are characterized as being quiescent, the TK activity was in the same range as in quiescent lymphocytes from control donors. However, quantification of the TKI mRNA level shows that all five CLL patients had a very high level (6 to 22 x IO6 copies mg-’ protein) of TKI mRNA, corresponding to the level in dividing...

  8. Negative emotions affect postoperative scores for evaluating functional knee recovery and quality of life after total knee replacement

    Directory of Open Access Journals (Sweden)

    A. Qi

    2016-01-01

    Full Text Available This study aimed to determine whether psychological factors affect health-related quality of life (HRQL and recovery of knee function in total knee replacement (TKR patients. A total of 119 TKR patients (male: 38; female: 81 completed the Beck Anxiety Inventory (BAI, Beck Depression Inventory (BDI, State Trait Anxiety Inventory (STAI, Eysenck Personality Questionnaire-revised (EPQR-S, Knee Society Score (KSS, and HRQL (SF-36. At 1 and 6 months after surgery, anxiety, depression, and KSS scores in TKR patients were significantly better compared with those preoperatively (P<0.05. SF-36 scores at the sixth month after surgery were significantly improved compared with preoperative scores (P<0.001. Preoperative Physical Component Summary Scale (PCS and Mental Component Summary Scale (MCS scores were negatively associated with extraversion (E score (B=-0.986 and -0.967, respectively, both P<0.05. Postoperative PCS and State Anxiety Inventory (SAI scores were negatively associated with neuroticism (N score; B=-0.137 and -0.991, respectively, both P<0.05. Postoperative MCS, SAI, Trait Anxiety Inventory (TAI, and BAI scores were also negatively associated with the N score (B=-0.367, -0.107, -0.281, and -0.851, respectively, all P<0.05. The KSS function score at the sixth month after surgery was negatively associated with TAI and N scores (B=-0.315 and -0.532, respectively, both P<0.05, but positively associated with the E score (B=0.215, P<0.05. The postoperative KSS joint score was positively associated with postoperative PCS (B=0.356, P<0.05. In conclusion, for TKR patients, the scores used for evaluating recovery of knee function and HRQL after 6 months are inversely associated with the presence of negative emotions.

  9. Analysis of bacterial core communities in the central Baltic by comparative RNA-DNA-based fingerprinting provides links to structure-function relationships.

    Science.gov (United States)

    Brettar, Ingrid; Christen, Richard; Höfle, Manfred G

    2012-01-01

    Understanding structure-function links of microbial communities is a central theme of microbial ecology since its beginning. To this end, we studied the spatial variability of the bacterioplankton community structure and composition across the central Baltic Sea at four stations, which were up to 450 km apart and at a depth profile representative for the central part (Gotland Deep, 235 m). Bacterial community structure was followed by 16S ribosomal RNA (rRNA)- and 16S rRNA gene-based fingerprints using single-strand conformation polymorphism (SSCP) electrophoresis. Species composition was determined by sequence analysis of SSCP bands. High similarities of the bacterioplankton communities across several hundred kilometers were observed in the surface water using RNA- and DNA-based fingerprints. In these surface communities, the RNA- and DNA-based fingerprints resulted in very different pattern, presumably indicating large difference between the active members of the community as represented by RNA-based fingerprints and the present members represented by the DNA-based fingerprints. This large discrepancy changed gradually over depth, resulting in highly similar RNA- and DNA-based fingerprints in the anoxic part of the water column below 130 m depth. A conceivable mechanism explaining this high similarity could be the reduced oxidative stress in the anoxic zone. The stable communities on the surface and in the anoxic zone indicate the strong influence of the hydrography on the bacterioplankton community structure. Comparative analysis of RNA- and DNA-based community structure provided criteria for the identification of the core community, its key members and their links to biogeochemical functions.

  10. [LincRNA-ROR functions as a ceRNA to regulate Oct4, Sox2, and Nanog expression by sponging miR-145 and its effect on biologic characteristics of colonic cancer stem cells].

    Science.gov (United States)

    Yan, Z Y; Sun, X C

    2018-04-08

    Objective: To investigate the impact of lincRNA-ROR, a ceRNA by binding miR-145 on the expression of the downstream genes Oct4, Sox2 and Nanog, and related biological characteristics of colon cancer stem cells, and to elucidate the clinical significance of this molecular regulatory network. Methods: Fifty-two cases of colorectal cancer tissue and adjacent tissue were collected at Nanyang City Central Hospital and Nanyang Second Hospital, Henan Province, from 2014 to 2016. Real-time quantitative polymerase chain reaction (qPCR) was used to detect the expression of lincRNA-ROR and miR-145 in colorectal cancer tissue and isolated colon cancer cells. The correlation between the expression of lincRNA-ROR, miR-145 and the clinicopathologic features of colon cancer was performed. CD44(-)CD133(-) and CD44(+) CD133(+) cells were isolated from SW1116 by using flow cytometry. The expression of CD44, CD133, Oct4, Sox2, Nanog, lincRNA-ROR and miR-145 in cells were detected by qPCR. The relationship between lincRNA-ROR, miR-145, Oct4, Sox2 and Nanog was analyzed by bioinformatics, dual luciferase reporter assay, qPCR and Western blot. The effects of silencing lincRNA-ROR on the proliferation and chemosensitivity of colon cancer stem cells were detected by MTT, colony formation. Results: LincRNA-ROR was frequently up-regulated and inversely correlated with miR-145 down-regulation in the colon cancer specimens( P colon cancer stem cells proliferation and increased the sensitivity to chemotherapy. Conclusions: Linc-ROR functions as a key ceRNA to prevent core TFs, e. g., Oct4, Sox2, Nanog, from miR-145-mediated suppression in colon cancer stem cells and regulates cell proliferation and chemosensitivity.The data may provide insights into the pathophysiological interactions of the components of genetic networks in the development of colon cancer and may lead to new therapies in the future.

  11. The Dynamics of microRNA Transcriptome in Bovine Corpus Luteum during Its Formation, Function, and Regression

    Directory of Open Access Journals (Sweden)

    Rreze M. Gecaj

    2017-12-01

    Full Text Available The formation, function, and subsequent regression of the ovarian corpus luteum (CL are dynamic processes that enable ovary cyclical activity. Studies in whole ovary tissue have found microRNAs (miRNAs to by critical for ovary function. However, relatively little is known about the role of miRNAs in the bovine CL. Utilizing small RNA next-generation sequencing we profiled miRNA transcriptome in bovine CL during the entire physiological estrous cycle, by sampling the CL on days: d 1–2, d 3–4, and d 5–7 (early CL, eCL, d 8–12 (mid CL, mCL, d 13–16 (late CL, lCL, and d > 18 (regressed CL, rCL. We characterized patterns of miRNAs abundance and identified 42 miRNAs that were consistent significantly different expressed (DE in the eCL relative to their expression at each of the analyzed stages (mCL, lCL, and rCL. Out of these, bta-miR-210-3p, −2898, −96, −7-5p, −183-5p, −182, and −202 showed drastic up-regulation with a fold-change of ≥2.0 and adjusted P < 0.01 in the eCL, while bta-miR-146a was downregulated at lCL and rCL vs. the eCL. Another 24, 11, and 21 miRNAs were significantly DE only between individual comparisons, eCL vs. the mCL, lCL, and rCL, respectively. Irrespective of cycle stage two miRNAs, bta-miR-21-5p and bta-miR-143 were identified as the most abundant miRNAs species and show opposing expression abundance. Whilst bta-miR-21-5p peaked in number of reads in the eCL and was significantly downregulated in the mCL and lCL, bta-miR-143 reached its peak in the rCL and is significantly downregulated in the eCL. MiRNAs with significant DE in at least one cycle stage (CL class were further grouped into eight distinct clusters by the self-organizing tree algorithm (SOTA. Half of the clusters contain miRNAs with low-expression, whilst the other half contain miRNAs with high-expression levels during eCL. Prediction analysis for significantly DE miRNAs resulted in target genes involved with CL formation

  12. Multidrug resistance mediated by ABC transporters in osteosarcoma cell lines: mRNA analysis and functional radiotracer studies

    Energy Technology Data Exchange (ETDEWEB)

    Gomes, Celia Maria Freitas [Department of Pathology, Leiden University Medical Center, 2300 RC Leiden (Netherlands); Faculty of Medicine, Institute of Biophysics/Biomathematics, IBILI, 3000-354 Coimbra (Portugal)]. E-mail: cgomes@ibili.uc.pt; van Paassen, Heidi [Department of Pathology, Leiden University Medical Center, 2300 RC Leiden (Netherlands); Romeo, Salvatore [Department of Pathology, Leiden University Medical Center, 2300 RC Leiden (Netherlands); Welling, Mick M. [Department of Radiology, Section of Nuclear Medicine, Leiden University Medical Center, 2300 RC Leiden (Netherlands); Feitsma, R.I.J. [Department of Radiology, Section of Nuclear Medicine, Leiden University Medical Center, 2300 RC Leiden (Netherlands); Abrunhosa, Antero J. [Faculty of Medicine, Institute of Biophysics/Biomathematics, IBILI, 3000-354 Coimbra (Portugal); Botelho, M. Filomena [Faculty of Medicine, Institute of Biophysics/Biomathematics, IBILI, 3000-354 Coimbra (Portugal); Hogendoorn, Pancras C.W. [Department of Pathology, Leiden University Medical Center, 2300 RC Leiden (Netherlands); Pauwels, Ernest [Department of Radiology, Section of Nuclear Medicine, Leiden University Medical Center, 2300 RC Leiden (Netherlands); Cleton-Jansen, Anne Marie [Department of Pathology, Leiden University Medical Center, 2300 RC Leiden (Netherlands)

    2006-10-15

    Drug resistance remains a significant impediment to successful chemotherapy and constitutes a major prognostic factor in osteosarcoma (OS) patients. This study was designed to identify the role and prognostic significance of multidrug-resistance (MDR)-related transporters, such as multidrug resistance protein 1 (MDR1), multidrug-resistance-associated protein (MRP1) and breast-cancer-related protein (BCRP), in OS using cationic lipophilic radiotracers. We evaluated the chemosensitivity of four OS cell lines (Saos-2, 143B, MNNG/HOS and U-2OS) to doxorubicin (DOX), cisplatin (CIS) and methotrexate. The expression of MDR-related transporters was analyzed at mRNA level by quantitative polymerase chain reaction and at functional level by {sup 99m}Tc sestamibi and {sup 99m}Tc tetrofosmin. The effectiveness of MDR modulators [cyclosporin A (CsA) and imatinib] on transporter inhibition and on the reversal of resistance was also assessed. MNNG/HOS and U-2OS cells expressing high levels of MDR1 were highly resistant to DOX and showed reduced accumulation and higher efflux for radiotracers. Although MRP1 was uniformly expressed in all cells, only U-2OS was resistant to CIS. CsA restored sensitivity to DOX and CIS, and enhanced the accumulation and efflux half-life of radiotracers in MDR1-expressing cell lines. The chemosensitivity of OS cells to DOX was strongly dependent on mRNA MDR1 expression and could be circumvented by adding CsA. The kinetic parameters of radiotracers correlated with MDR1 expression levels, hence predicting DOX resistance. We concluded that sensitivity to chemotherapy is strongly dependent on the expression of MDR1 transporter and that radiotracer studies could prove clinically useful in predicting chemotherapy response and in evaluating the efficacy of MDR-reversing agents.

  13. Relationship between PPARα mRNA expression and mitochondrial respiratory function and ultrastructure of the skeletal muscle of patients with COPD.

    Science.gov (United States)

    Zhang, Jian-Qing; Long, Xiang-Yu; Xie, Yu; Zhao, Zhi-Huan; Fang, Li-Zhou; Liu, Ling; Fu, Wei-Ping; Shu, Jing-Kui; Wu, Jiang-Hai; Dai, Lu-Ming

    2017-11-02

    Peripheral muscle dysfunction is an important complication in patients with chronic obstructive pulmonary disease (COPD). The objective of this study was to explore the relationship between the levels of peroxisome proliferator-activated receptor α (PPARα) mRNA expression and the respiratory function and ultrastructure of mitochondria in the vastus lateralis of patients with COPD. Vastus lateralis biopsies were performed on 14 patients with COPD and 6 control subjects with normal lung function. PPARα mRNA levels in the muscle tissue were detected by real-time PCR. A Clark oxygen electrode was used to assess mitochondrial respiratory function. Mitochondrial number, fractional area in skeletal muscle cross-sections, and Z-line width were observed via transmission electron microscopy. The PPARα mRNA expression was significantly lower in COPD patients with low body mass index (BMIL) than in both COPD patients with normal body mass index (BMIN) and controls. Mitochondrial respiratory function (assessed by respiratory control ratio) was impaired in COPD patients, particularly in BMIL. Compared with that in the control group, mitochondrial number and fractional area were lower in the BMIL group, but were maintained in the BMIN group. Further, the Z-line became narrow in the BMIL group. PPARα mRNA expression was positively related to mitochondrial respiratory function and volume density. In COPD patients with BMIN, mitochondria volume density was maintained, while respiratory function decreased, whereas both volume density and respiratory function decreased in COPD patients with BMIL. PPARα mRNA expression levels are associated with decreased mitochondrial respiratory function and volume density, which may contribute to muscle dysfunction in COPD patients.

  14. A Genome-Wide mRNA Screen and Functional Analysis Reveal FOXO3 as a Candidate Gene for Chicken Growth.

    Directory of Open Access Journals (Sweden)

    Biao Chen

    Full Text Available Chicken growth performance provides direct economic benefits to the poultry industry. However, the underlying genetic mechanisms are unclear. The objective of this study was to identify candidate genes associated with chicken growth and investigate their potential mechanisms. We used RNA-Seq to study the breast muscle transcriptome in high and low tails of Recessive White Rock (WRRh, WRRl and Xinghua chickens (XHh, XHl. A total of 60, 23, 153 and 359 differentially expressed genes were detected in WRRh vs. WRRl, XHh vs. XHl, WRRh vs. XHh and WRRl vs. XHl, respectively. GO, KEGG pathway and gene network analyses showed that CEBPB, FBXO32, FOXO3 and MYOD1 played key roles in growth. The functions of FBXO32 and FOXO3 were validated. FBXO32 was predominantly expressed in leg muscle, heart and breast muscle. After decreased FBXO32 expression, growth-related genes such as PDK4, IGF2R and IGF2BP3 were significantly down-regulated (P < 0.05. FBXO32 was significantly (P < 0.05 associated with carcass and meat quality traits, but not growth traits. FOXO3 was predominantly expressed in breast and leg muscle. In both of these tissues, the FOXO3 mRNA level in XH was significantly higher than that in WRR chickens with normal body weight (P < 0.05. In DF-1 cells, siRNA knockdown of FOXO3 significantly (P < 0.01 inhibited the MYOD expression and significantly up-regulated (P < 0.01 or P < 0.05 the expression of growth-related genes including CEBPB, FBXO32, GH, GHR, IGF1R, IGF2R, IGF2BP1, IGF2BP3, INSR, PDK1 and PDK4. Moreover, 18 SNPs were identified in FOXO3. G66716193A was significantly (P < 0.05 associated with growth traits. The sites C66716002T, C66716195T and A66716179G were significantly (P < 0.05 associated with growth or carcass traits. These results demonstrated that FOXO3 is a candidate gene influencing chicken growth. Our observations provide new clues to understand the molecular basis of chicken growth.

  15. Duplication and Loss of Function of Genes Encoding RNA Polymerase III Subunit C4 Causes Hybrid Incompatibility in Rice

    Directory of Open Access Journals (Sweden)

    Giao Ngoc Nguyen

    2017-08-01

    Full Text Available Reproductive barriers are commonly observed in both animals and plants, in which they maintain species integrity and contribute to speciation. This report shows that a combination of loss-of-function alleles at two duplicated loci, DUPLICATED GAMETOPHYTIC STERILITY 1 (DGS1 on chromosome 4 and DGS2 on chromosome 7, causes pollen sterility in hybrid progeny derived from an interspecific cross between cultivated rice, Oryza sativa, and an Asian annual wild rice, O. nivara. Male gametes carrying the DGS1 allele from O. nivara (DGS1-nivaras and the DGS2 allele from O. sativa (DGS2-T65s were sterile, but female gametes carrying the same genotype were fertile. We isolated the causal gene, which encodes a protein homologous to DNA-dependent RNA polymerase (RNAP III subunit C4 (RPC4. RPC4 facilitates the transcription of 5S rRNAs and tRNAs. The loss-of-function alleles at DGS1-nivaras and DGS2-T65s were caused by weak or nonexpression of RPC4 and an absence of RPC4, respectively. Phylogenetic analysis demonstrated that gene duplication of RPC4 at DGS1 and DGS2 was a recent event that occurred after divergence of the ancestral population of Oryza from other Poaceae or during diversification of AA-genome species.

  16. Analysis of a predicted nuclear localization signal: implications for the intracellular localization and function of the Saccharomyces cerevisiae RNA-binding protein Scp160.

    Science.gov (United States)

    Brykailo, Melissa A; McLane, Laura M; Fridovich-Keil, Judith; Corbett, Anita H

    2007-01-01

    Gene expression is controlled by RNA-binding proteins that modulate the synthesis, processing, transport and stability of various classes of RNA. Some RNA-binding proteins shuttle between the nucleus and cytoplasm and are thought to bind to RNA transcripts in the nucleus and remain bound during translocation to the cytoplasm. One RNA-binding protein that has been hypothesized to function in this manner is the Saccharomyces cerevisiae Scp160 protein. Although the steady-state localization of Scp160 is cytoplasmic, previous studies have identified putative nuclear localization (NLS) and nuclear export (NES) signals. The goal of this study was to test the hypothesis that Scp160 is a nucleocytoplasmic shuttling protein. We exploited a variety of yeast export mutants to capture any potential nuclear accumulation of Scp160 and found no evidence that Scp160 enters the nucleus. These localization studies were complemented by a mutational analysis of the predicted NLS. Results indicate that key basic residues within the predicted NLS of Scp160 can be altered without severely affecting Scp160 function. This finding has important implications for understanding the function of Scp160, which is likely limited to the cytoplasm. Additionally, our results provide strong evidence that the presence of a predicted nuclear localization signal within the sequence of a protein should not lead to the assumption that the protein enters the nucleus in the absence of additional experimental evidence.

  17. The microRNA Expression Profile in Donation after Cardiac Death (DCD Livers and Its Ability to Identify Primary Non Function.

    Directory of Open Access Journals (Sweden)

    Shirin Elizabeth Khorsandi

    Full Text Available Donation after cardiac death (DCD livers are marginal organs for transplant and their use is associated with a higher risk of primary non function (PNF or early graft dysfunction (EGD. The aim was to determine if microRNA (miRNA was able to discriminate between DCD livers of varying clinical outcome. DCD groups were categorized as PNF retransplanted within a week (n=7, good functional outcome (n=7 peak aspartate transaminase (AST ≤ 1000 IU/L and EGD (n=9 peak AST ≥ 2500 IU/L. miRNA was extracted from archival formalin fixed post-perfusion tru-cut liver biopsies. High throughput expression analysis was performed using miRNA arrays. Bioinformatics for expression data analysis was performed and validated with real time quantitative PCR (RT-qPCR. The function of miRNA of interest was investigated using computational biology prediction algorithms. From the array analysis 16 miRNAs were identified as significantly different (p<0.05. On RT-qPCR miR-155 and miR-940 had the highest expression across all three DCD clinical groups. Only one miRNA, miR-22, was validated with marginal significance, to have differential expression between the three groups (p=0.049. From computational biology miR-22 was predicted to affect signalling pathways that impact protein turnover, metabolism and apoptosis/cell cycle. In conclusion, microRNA expression patterns have a low diagnostic potential clinically in discriminating DCD liver quality and outcome.

  18. Bombyx mori DNA/RNA non-specific nuclease: expression of isoforms in insect culture cells, subcellular localization and functional assays.

    Science.gov (United States)

    Liu, Jisheng; Swevers, Luc; Iatrou, Kostas; Huvenne, Hanneke; Smagghe, Guy

    2012-08-01

    A DNA/RNA non-specific alkaline nuclease (BmdsRNase) was isolated from the digestive juice of Bombyx mori. While originally reported to be produced by the midgut only, in this project it was found that the mRNA of this enzyme was also expressed in the epidermis, fat body, gut, thoracic muscles, Malpighian tubules, brain, and silk glands of 5th instar larvae, indicating additional functions to its reported role in nucleic acid digestion in the midgut. In order to study the functional properties of BmdsRNase, three pEA-BmdsRNase expression constructs were generated, characterized by presence or absence of a signal peptide and a propeptide, and used for expression in lepidopteran Hi5 tissue culture cells. Western blot indicated that these different forms of BmdsRNase protein were not secreted into the growth medium, while they were detected in the pellets and supernatants of Hi5 cell extracts. Nucleic acids cleavage experiments indicated that full-length BmdsRNase could digest dsRNA and that the processed form (absence of signal peptide and propeptide) of BmdsRNase could degrade both DNA and dsRNA in Hi5 cell culture. Using a reporter assay targeted by transfected homologous dsRNA, it was shown that the digestive property of the processed form could interfere with the RNAi response. Immunostaining of processed BmdsRNase protein showed asymmetric localization in the cellular cytoplasm and co-localization with Flag-tagged Dicer-2 was also observed. In conclusion, our in vitro studies indicated that intracellular protein isoforms of BmdsRNase can be functional and involved in the regulation of nucleic acid metabolism in the cytoplasm. In particular, because of its propensity to degrade dsRNA, the enzyme might be involved in the innate immune response against invading nucleic acids such as RNA viruses. Copyright © 2012 Elsevier Ltd. All rights reserved.

  19. Function and structure in phage Qbeta RNA replicase. Association of EF-Tu-Ts with the other enzyme subunits

    DEFF Research Database (Denmark)

    Blumenthal, T; Young, R A; Brown, S

    1976-01-01

    of EF-Tu-Ts from the other enzyme subunits: whereas free EF-Tu-Ts binds GDP independently of salt concentration, this binding by Qbeta replicase is sensitive to high ionic strength and remains so in the presence of Qbeta RNA. Furthermore, RNA does not allow the release of EF-Ts from EF-Tu by GTP...... for one another increases with increasing ionic strength. The enzyme is capable of initiation of RNA synthesis with synthetic templates only when in the low ionic strength conformation. Elongation of initiated polynucleotide chains is not affectedby ionic strength. Addition of Qbeta RNA to the enzyme also...... alters its quaternary structure: the EF-Tu-Ts cannot be covalently attached to the other enzyme subunits with bifunctional cross-linking reagents in the presence of RNA. This conformational change is not influenced by ionic strength. The addition of Qbeta RNA to the enzyme, does not result in the release...

  20. Recovery of balance function among individuals with total knee arthroplasty: Comparison of responsiveness among four balance tests.

    Science.gov (United States)

    Chan, Andy C M; Ouyang, Xi H; Jehu, Deborah A M; Chung, Raymond C K; Pang, Marco Y C

    2018-01-01

    Balance deficits are common after total knee arthroplasty (TKA); however the responsiveness of commonly used balance measurement tools has not been well defined. The objective of this prospective study was to compare the internal and external responsiveness of four measurement tools in assessing recovery of balance function following TKA. A total of 134 individuals with TKA (95 women; age: 66.3±6.6years) completed the Balance Evaluation Systems Test (BESTest), Mini-BESTest, Brief-BESTest, and Berg Balance Scale (BBS) at 2, 4, 8, 12, and 24 weeks post-TKA. The Functional Gait Assessment (FGA) served as the anchor measure, and was also measured across these time points. Internal responsiveness was indicated by the standardized response mean (SRM), while external responsiveness was reflected by the degree of association of the changes of balance scores with those of FGA. The SRM ranged from 0.60-1.14 for the BESTest, 0.40-0.94 for the Mini-BESTest, 0.27-0.91 for the Brief-BESTest, and 0.19-0.70 for the BBS, over time. The change in BESTest and Mini-BESTest scores predicted the change in the FGA scores across all time periods, except for the Mini-BESTest between weeks 12-24, accounting for 13-27%, and 12-24% of the variance, respectively. The Brief-BESTest scores only predicted FGA scores between the weeks 2-4 (R 2 =20%). The changes in BBS scores were not associated with the FGA. The BESTest is the most responsive in measuring recovery of balance among individuals with TKA. The Mini-BESTest is a reasonable option during time constraints. Copyright © 2017. Published by Elsevier B.V.

  1. Methods for RNA Analysis

    DEFF Research Database (Denmark)

    Olivarius, Signe

    While increasing evidence appoints diverse types of RNA as key players in the regulatory networks underlying cellular differentiation and metabolism, the potential functions of thousands of conserved RNA structures encoded in mammalian genomes remain to be determined. Since the functions of most...... RNAs rely on interactions with proteins, the establishment of protein-binding profiles is essential for the characterization of RNAs. Aiming to facilitate RNA analysis, this thesis introduces proteomics- as well as transcriptomics-based methods for the functional characterization of RNA. First, RNA......-protein pulldown combined with mass spectrometry analysis is applied for in vivo as well as in vitro identification of RNA-binding proteins, the latter succeeding in verifying known RNA-protein interactions. Secondly, acknowledging the significance of flexible promoter usage for the diversification...

  2. Knee Function and Subjective Stability Following Total Condylar Arthroplasty in Joints with Preoperative Varus or Valgus Deformity.

    Science.gov (United States)

    Kokoszka, Paweł; Markuszewski, Jacek; Łapaj, Łukasz; Kruczyński, Jacek

    2015-10-01

    Total knee arthroplasty in joints with valgus or varus deformity is technically demanding. Careful soft tissue balance as well as restitution of anatomical knee axis has a profound effect on postoperative function of the joint, however little is known about differences in subjective stability following surgery between preoperative valgus and varus knees. Studied group consisted of 60 patients who underwent total condylar knee arthroplasty with one type of implant (Stryker Triathlon). Mean follow-up was 2,9 years (1-6 years). The group included 25 patients with valgus and 35 patients with varus preoperative deformity. All patients filled Knee Injury and Osteoarthritis Outcome Score (KOOS) forms. Detailed clinical and radiological assessment was performed. Mean KOOS score was slightly higher in patients with varus deformity, as compared to cases with valgus deformity. At physical examination higher LCL deficiency rate was observed in varus knees. Subjective instability was reported by eight patients (5 valgus and 3 varus). In all cases instability coexisted with decreased MCL tightness and implant position was correct in those patients. No subjective instability was reported by patients with clinical LCL deficiency. Furthermore KOOS scores in these patients were higher (85,8) as compared to cases with decreased MCL tension (79,1). 1. In patients with proper implant alignment subjective instability is related to postoperative MCL deficiency, regardless preoperative deformity in coronal plane. 2. The post-op LCL laxity does not compromise subjective stability, nor influence subjective outcome, as demonstrated with KOOS scores.

  3. RNA-seq analysis of Quercus pubescens Leaves: de novo transcriptome assembly, annotation and functional markers development.

    Directory of Open Access Journals (Sweden)

    Sara Torre

    Full Text Available Quercus pubescens Willd., a species distributed from Spain to southwest Asia, ranks high for drought tolerance among European oaks. Q. pubescens performs a role of outstanding significance in most Mediterranean forest ecosystems, but few mechanistic studies have been conducted to explore its response to environmental constrains, due to the lack of genomic resources. In our study, we performed a deep transcriptomic sequencing in Q. pubescens leaves, including de novo assembly, functional annotation and the identification of new molecular markers. Our results are a pre-requisite for undertaking molecular functional studies, and may give support in population and association genetic studies. 254,265,700 clean reads were generated by the Illumina HiSeq 2000 platform, with an average length of 98 bp. De novo assembly, using CLC Genomics, produced 96,006 contigs, having a mean length of 618 bp. Sequence similarity analyses against seven public databases (Uniprot, NR, RefSeq and KOGs at NCBI, Pfam, InterPro and KEGG resulted in 83,065 transcripts annotated with gene descriptions, conserved protein domains, or gene ontology terms. These annotations and local BLAST allowed identify genes specifically associated with mechanisms of drought avoidance. Finally, 14,202 microsatellite markers and 18,425 single nucleotide polymorphisms (SNPs were, in silico, discovered in assembled and annotated sequences. We completed a successful global analysis of the Q. pubescens leaf transcriptome using RNA-seq. The assembled and annotated sequences together with newly discovered molecular markers provide genomic information for functional genomic studies in Q. pubescens, with special emphasis to response mechanisms to severe constrain of the Mediterranean climate. Our tools enable comparative genomics studies on other Quercus species taking advantage of large intra-specific ecophysiological differences.

  4. RNA topology

    OpenAIRE

    Frank-Kamenetskii, Maxim D.

    2013-01-01

    A new variety on non-coding RNA has been discovered by several groups: circular RNA (circRNA). This discovery raises intriguing questions about the possibility of the existence of knotted RNA molecules and the existence of a new class of enzymes changing RNA topology, RNA topoisomerases.

  5. A Genome-Wide mRNA Screen and Functional Analysis Reveal FOXO3 as a Candidate Gene for Chicken Growth

    Science.gov (United States)

    Chen, Biao; Xu, Jiguo; He, Xiaomei; Xu, Haiping; Li, Guihuan; Du, Hongli; Nie, Qinghua; Zhang, Xiquan

    2015-01-01

    Chicken growth performance provides direct economic benefits to the poultry industry. However, the underlying genetic mechanisms are unclear. The objective of this study was to identify candidate genes associated with chicken growth and investigate their potential mechanisms. We used RNA-Seq to study the breast muscle transcriptome in high and low tails of Recessive White Rock (WRRh, WRRl) and Xinghua chickens (XHh, XHl). A total of 60, 23, 153 and 359 differentially expressed genes were detected in WRRh vs. WRRl, XHh vs. XHl, WRRh vs. XHh and WRRl vs. XHl, respectively. GO, KEGG pathway and gene network analyses showed that CEBPB, FBXO32, FOXO3 and MYOD1 played key roles in growth. The functions of FBXO32 and FOXO3 were validated. FBXO32 was predominantly expressed in leg muscle, heart and breast muscle. After decreased FBXO32 expression, growth-related genes such as PDK4, IGF2R and IGF2BP3 were significantly down-regulated (P chickens with normal body weight (P chicken growth. Our observations provide new clues to understand the molecular basis of chicken growth. PMID:26366565

  6. Healing and functional outcome of a subscapularis peel repair with a stem-based repair after total shoulder arthroplasty.

    Science.gov (United States)

    Gobezie, Reuben; Denard, Patrick J; Shishani, Yousef; Romeo, Anthony A; Lederman, Evan

    2017-09-01

    The purpose of this study was to evaluate functional outcome and healing of a subscapularis peel with a stem-based repair after total shoulder arthroplasty (TSA). The hypothesis was that the repair would lead to subscapularis healing in the majority of cases. A prospective analysis was performed on a consecutive series of TSAs. Range of motion and functional outcome were assessed according to American Shoulder and Elbow Surgeons, Single Assessment Numeric Evaluation, Simple Shoulder Test, and visual analog scale scores at a minimum follow-up of 1 year. Belly-press and lift-off tests were also performed. An ultrasound evaluation assessed subscapularis healing at final follow-up. At a mean follow-up of 15 months, 60 patients (mean age, 64 years) were examined. Mean forward flexion improved from 115° to 137°. External rotation at the side improved from 27° to 52°, and internal rotation improved from L4 to L2 (P 90% postoperative healing of the subscapularis. Copyright © 2017 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Elsevier Inc. All rights reserved.

  7. Acute central nervous system (CNS) toxicity of total body irradiation (TBI) measured using neuropsychological testing of attention functions

    International Nuclear Information System (INIS)

    Wenz, Frederik; Steinvorth, Sarah; Lohr, Frank; Hacke, Werner; Wannenmacher, Michael

    1999-01-01

    Purpose: The purpose of this study was to investigate acute normal tissue damage of low irradiation doses to the healthy, adult central nervous system (CNS) using neuropsychological testing of attention functions. Methods and Materials: Neuropsychological testing (IQ, attention [modified Trail-Making Test A, Digit Symbol Test, D2 Test, Wiener Determination Machine]) was used to examine 40 patients (43 ± 10 years) before and immediately after the first fraction (1.2 Gy) of hyperfractionated total body irradiation (TBI) at the University of Heidelberg. The patients received antiemetic premedication. Test results are given as mean percentiles ± standard deviation, with 50 ± 34 being normal. Thirty-eight control patients (53 ± 15 years) were studied to quantify the influence of hospitalization, stress, and repeated testing. Results: The patients showed normal baseline test results (IQ = 101 ± 14, attention = 54 ± 28) and no decrease in test results after 1.2 Gy TBI. Attention functions improved (66 ± 25) corresponding to a practice effect of repeated testing that was seen in the control group, although alternate versions of the tests were used (IQ = 104 ± 10, attention before = 42 ± 29, attention after = 52 ± 31). Conclusion: Our data show no deterioration of neuropsychologic test results acutely after 1.2 Gy whole body exposure in adult patients without CNS disease receiving antiemetic medication

  8. Profile and functional analysis of small RNAs derived from Aspergillus fumigatus infected with double-stranded RNA mycoviruses.

    Science.gov (United States)

    Özkan, Selin; Mohorianu, Irina; Xu, Ping; Dalmay, Tamas; Coutts, Robert H A

    2017-05-30

    Mycoviruses are viruses that naturally infect and replicate in fungi. Aspergillus fumigatus, an opportunistic pathogen causing fungal lung diseases in humans and animals, was recently shown to harbour several different types of mycoviruses. A well-characterised defence against virus infection is RNA silencing. The A. fumigatus genome encodes essential components of the RNA silencing machinery, including Dicer, Argonaute and RNA-dependent RNA polymerase (RdRP) homologues. Active silencing of double-stranded (ds)RNA and the generation of small RNAs (sRNAs) has been shown for several mycoviruses and it is anticipated that a similar mechanism will be activated in A. fumigatus isolates infected with mycoviruses. To investigate the existence and nature of A. fumigatus sRNAs, sRNA-seq libraries of virus-free and virus-infected isolates were created using Scriptminer adapters and compared. Three dsRNA viruses were investigated: Aspergillus fumigatus partitivirus-1 (AfuPV-1, PV), Aspergillus fumigatus chrysovirus (AfuCV, CV) and Aspergillus fumigatus tetramycovirus-1 (AfuTmV-1, NK) which were selected because they induce phenotypic changes such as coloration and sectoring. The dsRNAs of all three viruses, which included two conventionally encapsidated ones PV and CV and one unencapsidated example NK, were silenced and yielded characteristic vsiRNAs together with co-incidental silencing of host fungal genes which shared sequence homology with the viral genomes. Virus-derived sRNAs were detected and characterised in the presence of virus infection. Differentially expressed A. fumigatus microRNA-like (miRNA-like) sRNAs and small interfering RNAs (siRNAs) were detected and validated. Host sRNA loci which were differentially expressed as a result of virus infection were also identified. To our knowledge, this is the first study reporting the sRNA profiles of A. fumigatus isolates.

  9. Raman crystallography of RNA.

    Science.gov (United States)

    Gong, Bo; Chen, Jui-Hui; Yajima, Rieko; Chen, Yuanyuan; Chase, Elaine; Chadalavada, Durga M; Golden, Barbara L; Carey, Paul R; Bevilacqua, Philip C

    2009-10-01

    Raman crystallography is the application of Raman spectroscopy to single crystals. This technique has been applied to a variety of protein molecules where it has provided unique information about biopolymer folding, substrate binding, and catalysis. Here, we describe the application of Raman crystallography to functional RNA molecules. RNA represents unique opportunities and challenges for Raman crystallography. One issue that confounds studies of RNA is its tendency to adopt multiple non-functional folds. Raman crystallography has the advantage that it isolates a single state of the RNA within the crystal and can evaluate its fold, metal ion binding properties (ligand identity, stoichiometry, and affinity), proton binding properties (identity, stoichiometry, and affinity), and catalytic potential. In particular, base-specific stretches can be identified and then associated with the binding of metal ions and protons. Because measurements are carried out in the hanging drop at ambient, rather than cryo, conditions and because RNA crystals tend to be approximately 70% solvent, RNA dynamics and conformational changes become experimentally accessible. This review focuses on experimental setup and procedures, acquisition and interpretation of Raman data, and determination of physicochemical properties of the RNA. Raman crystallographic and solution biochemical experiments on the HDV RNA enzyme are summarized and found to be in excellent agreement. Remarkably, characterization of the crystalline state has proven to help rather than hinder functional characterization of functional RNA, most likely because the tendency of RNA to fold heterogeneously is limited in a crystalline environment. Future applications of Raman crystallography to RNA are briefly discussed.

  10. Structural and functional characterization of the coxsackievirus B3 CRE(2C): role of CRE(2C) in negative- and positive-strand RNA synthesis.

    NARCIS (Netherlands)

    Ooij, M.J.M. van; Vogt, D.A.; Paul, A.; Castro, C.; Kuijpers, J.M.; Kuppeveld, F.J.M. van; Cameron, C.E.; Wimmer, E.; Andino, R.; Melchers, W.J.G.

    2006-01-01

    A stem-loop element located within the 2C-coding region of the coxsackievirus B3 (CVB3) genome has been proposed to function as a cis-acting replication element (CRE). It is shown here that disruption of this structure indeed interfered with viral RNA replication in vivo and abolished uridylylation

  11. Influence of body mass index (BMI on functional improvements at 3 years following total knee replacement: a retrospective cohort study.

    Directory of Open Access Journals (Sweden)

    Paul Baker

    Full Text Available BACKGROUND: The number of patients presenting for total knee replacement who are classified as obese is increasing. The functional benefits of performing TKR in these patients are unclear. AIM: To assess the influence pre-operative body mass index has upon knee specific function, general health status and patient satisfaction at 3 years following total knee replacement. DESIGN: Retrospective comparative cohort study using prospectively collected data from an institutional arthroplasty register. METHODS: 1367 patients were assessed using the Western Ontario and McMaster University Osteoarthritis Index (WOMAC and Medical Outcomes Trust Short Form-36 (SF-36 scores supplemented by a validated measure of satisfaction pre-operatively and subsequently at 1,2 and 3 year post-operatively. Comparisons were made by dividing the cohort into 4 groups based on body mass index (BMI 18.5-25.0 kg/m(2 (n = 253;>25.0-30.0 kg/m(2 (n = 559;>30.0-35.0 kg/m(2 (n = 373;>35.0 kg/m(2 (n = 182. RESULTS: Despite lower pre-operative, 1 and 3 year WOMAC and SF-36 scores patients with the highest BMIs >35.0 kg/m(2 experienced similar improvements to patients with a 'normal' BMI (18.5-25.0 kg/m(2 at 1 year (Difference in WOMAC improvement = 0.0 (95%CI -5.2 to 5.2, p = 1.00 and this improvement was sustained at up to 3 years (Difference in 1 year to 3 year improvement = 2.2 (95%CI: -2.1 to 6.5, p = 1.00. This effect was also observed for the SF-36 mental and physical component scores. Despite equivalent functional improvements levels of satisfaction in the >35.0 kg/m(2 group were lower than for any other BMI group (>35.0 kg/m(2 = 84.6% satisfied versus 18.5-5.0 kg/m(2 = 93.3% satisfied,p = 0.01 as was the proportion of patients who stated they would have the operation again (>35.0 kg/m(2 = 69.6% versus 18.5-25.0 kg/m(2 = 82.2%,p = 0.01. CONCLUSION: Obese and morbidly obese patients gain as much functional benefit from

  12. A noncoding RNA containing a SINE-B1 motif associates with meiotic metaphase chromatin and has an indispensable function during spermatogenesis.

    Directory of Open Access Journals (Sweden)

    Ryusuke Nakajima

    Full Text Available A search for early response genes that are activated following germ cell induction from mouse embryonic stem cells in vitro led us to the isolation of a long noncoding RNA that contains a SINE (short interspersed element-B1F motif that was named R53. In situ hybridization and northern blot analyses revealed that the R53 subfragment RNA bears a B1F motif, is processed from the primary transcript, is expressed in adult testis and is predominantly localized in meiotic metaphase chromatin during spermatogenesis. Recent studies of chromosome-associated RNAs have explored novel functions of noncoding RNAs. Specifically, chromosome-bound noncoding RNAs function not only as structural components of chromosome but also as scaffolds that recruit epigenetic modulators for transcriptional regulation, and they are dynamically rearranged during the cell cycle. However, few studies have explored meiotic chromatin; thus, R53 RNA appears to be the first long noncoding RNA to be tightly associated with the metaphase chromatin during spermatogenesis. Furthermore, R53 knockdown using a lentivirus-mediated RNAi injected into mouse testis and organ culture of the fragments revealed a remarkable reduction in postmeiotic cells and irregular up-regulation of several postmeiotic genes, which suggests the possibility that the SINE-B1-derived noncoding RNA R53 plays an indispensable role in the transcriptional regulation of key spermatogenesis genes.

  13. The Tudor domain protein Tapas, a homolog of the vertebrate Tdrd7, functions in the piRNA pathway to regulate retrotransposons in germline of Drosophila melanogaster.

    Science.gov (United States)

    Patil, Veena S; Anand, Amit; Chakrabarti, Alisha; Kai, Toshie

    2014-10-06

    Piwi-interacting RNAs (piRNAs) are a special class of small RNAs that provide defense against transposable elements in animal germline cells. In Drosophila, germline piRNAs are thought to be processed at a unique perinuclear structure, the nuage, that houses piRNA pathway proteins including the Piwi clade of Argonaute family proteins, along with several Tudor domain proteins, RNA helicases and nucleases. We previously demonstrated that Tudor domain protein Tejas (Tej), an ortholog of vertebrate Tdrd5, is an important component of the piRNA pathway. In the current study, we identified the paralog of the Drosophila tej gene, tapas (tap), which is an ortholog of vertebrate Tdrd7. Like Tej, Tap is localized at the nuage. Alone, tap loss leads to a mild increase in transposon expression and decrease in piRNAs targeting transposons expressed in the germline. The tap gene genetically interacts with other piRNA pathway genes and we also show that Tap physically interacts with piRNA pathway components, such as Piwi family proteins Aubergine and Argonaute3 and the RNA helicases Spindle-E and Vasa. Together with tej, tap is required for survival of germline cells during early stages and for polarity formation. We further observed that loss of tej and tap together results in more severe defects in the piRNA pathway in germline cells compared to single mutants: the double-mutant ovaries exhibit mis-localization of piRNA pathway components and significantly greater reduction of piRNAs against transposons predominantly expressed in germline compared to single mutants. The single or double mutants did not have any reduction in piRNAs mapping to transposons predominantly expressed in gonadal somatic cells or those derived from unidirectional clusters such as flamenco. Consistently, the loss of both tej and tap function resulted in mis-localization of Piwi in germline cells, whereas Piwi remained localized to the nucleus in somatic cells. Our observations suggest that tej and tap

  14. Small RNA-dependent expression of secondary metabolism is controlled by Krebs cycle function in Pseudomonas fluorescens.

    Science.gov (United States)

    Takeuchi, Kasumi; Kiefer, Patrick; Reimmann, Cornelia; Keel, Christoph; Dubuis, Christophe; Rolli, Joëlle; Vorholt, Julia A; Haas, Dieter

    2009-12-11

    Pseudomonas fluorescens CHA0, an antagonist of phytopathogenic fungi in the rhizosphere of crop plants, elaborates and excretes several secondary metabolites with antibiotic properties. Their synthesis depends on three small RNAs (RsmX, RsmY, and RsmZ), whose expression is positively controlled by the GacS-GacA two-component system at high cell population densities. To find regulatory links between primary and secondary metabolism in P. fluorescens and in the related species Pseudomonas aeruginosa, we searched for null mutations that affected central carbon metabolism as well as the expression of rsmY-gfp and rsmZ-gfp reporter constructs but without slowing down the growth rate in rich media. Mutation in the pycAB genes (for pyruvate carboxylase) led to down-regulation of rsmXYZ and secondary metabolism, whereas mutation in fumA (for a fumarase isoenzyme) resulted in up-regulation of the three small RNAs and secondary metabolism in the absence of detectable nutrient limitation. These effects required the GacS sensor kinase but not the accessory sensors RetS and LadS. An analysis of intracellular metabolites in P. fluorescens revealed a strong positive correlation between small RNA expression and the pools of 2-oxoglutarate, succinate, and fumarate. We conclude that Krebs cycle intermediates (already known to control GacA-dependent virulence factors in P. aeruginosa) exert a critical trigger function in secondary metabolism via the expression of GacA-dependent small RNAs.

  15. Structure–function relations in the NTPase domain of the antiviral tRNA ribotoxin Escherichia coli PrrC

    International Nuclear Information System (INIS)

    Meineke, Birthe; Shuman, Stewart

    2012-01-01

    Breakage of tRNA by Escherichia coli anticodon nuclease PrrC (EcoPrrC) underlies a host antiviral response to phage T4 infection. Expression of EcoPrrC is cytocidal in yeast, signifying that PrrC ribotoxicity crosses phylogenetic domain boundaries. EcoPrrC consists of an N-terminal NTPase module that resembles ABC transporters and a C-terminal nuclease module that is sui generis. PrrC homologs are prevalent in many other bacteria. Here we report that Haemophilus influenzae PrrC is toxic in E. coli and yeast. To illuminate structure–activity relations, we conducted a new round of mutational analysis of EcoPrrC guided by primary structure conservation among toxic PrrC homologs. We indentify 17 candidate active site residues in the NTPase module that are essential for toxicity in yeast when EcoPrrC is expressed at high gene dosage. Their functions could be educed by integrating mutational data with the atomic structure of the transition-state complex of a homologous ABC protein.

  16. MicroRNA Functions in Brite/Brown Fat — Novel Perspectives towards Anti-Obesity Strategies

    Directory of Open Access Journals (Sweden)

    Michael Karbiener

    2014-09-01

    Full Text Available Current anti-obesity strategies are aiming at restricting energy uptake, but still, obesity treatment is far from being satisfactory. The discovery of active brown adipose tissue (BAT in adult humans currently opens new avenues to combat obesity and follow-up complications as it tackles the other site of the energy balance: energy expenditure via non-shivering thermogenesis. This process of energy dissipation in the adipose tissue is tightly controlled, and the elucidation of its regulatory network is a key plank for therapeutic applications. MicroRNAs (miRNAs belong to a novel class of regulatory determinants which are small non-coding RNAs with vital roles in regulating gene expression that also play a role in many human diseases. In this review we summarize miRNAs which have been shown to govern thermogenic, i.e. brite or brown, adipocyte recruitment and physiology. Notably, most miRNAs in this context have so far been characterized solely in mice, revealing a great demand for more human studies. As in the context of other diseases, RNA-based therapeutics have meanwhile entered clinical trials, further exploring the functions of miRNAs in brown and white adipose tissues could result in novel therapeutic approaches to treat obesity and its follow-up complications.

  17. Structure-function relations in the NTPase domain of the antiviral tRNA ribotoxin Escherichia coli PrrC

    Energy Technology Data Exchange (ETDEWEB)

    Meineke, Birthe; Shuman, Stewart, E-mail: s-shuman@ski.mskcc.org

    2012-06-05

    Breakage of tRNA by Escherichia coli anticodon nuclease PrrC (EcoPrrC) underlies a host antiviral response to phage T4 infection. Expression of EcoPrrC is cytocidal in yeast, signifying that PrrC ribotoxicity crosses phylogenetic domain boundaries. EcoPrrC consists of an N-terminal NTPase module that resembles ABC transporters and a C-terminal nuclease module that is sui generis. PrrC homologs are prevalent in many other bacteria. Here we report that Haemophilus influenzae PrrC is toxic in E. coli and yeast. To illuminate structure-activity relations, we conducted a new round of mutational analysis of EcoPrrC guided by primary structure conservation among toxic PrrC homologs. We indentify 17 candidate active site residues in the NTPase module that are essential for toxicity in yeast when EcoPrrC is expressed at high gene dosage. Their functions could be educed by integrating mutational data with the atomic structure of the transition-state complex of a homologous ABC protein.

  18. Pre-operative functional mobility as an independent determinant of inpatient functional recovery after total knee arthroplasty during three periods that coincided with changes in clinical pathways.

    Science.gov (United States)

    van der Sluis, G; Goldbohm, R A; Elings, J E; Nijhuis-van der Sanden, M W; Akkermans, R P; Bimmel, R; Hoogeboom, T J; van Meeteren, N L

    2017-02-01

    To investigate whether pre-operative functional mobility is a determinant of delayed inpatient recovery of activities (IRoA) after total knee arthroplasty (TKA) in three periods that coincided with changes in the clinical pathway. All patients (n = 682, 73% women, mean age 70 years, standard deviation 9) scheduled for TKA between 2009 and 2015 were pre-operatively screened for functional mobility by the Timed-up-and-Go test (TUG) and De Morton mobility index (DEMMI). The cut-off point for delayed IRoA was set on the day that 70% of the patients were recovered, according to the Modified Iowa Levels of Assistance Scale (mILAS) (a 5-item activity scale). In a multivariable logistic regression analysis, we added either the TUG or the DEMMI to a reference model including established determinants. Both the TUG (Odds Ratio (OR) 1.10 per second, 95% confidence intervals (CI) 1.06 to 1.15) and the DEMMI (OR 0.96 per point on the 100-point scale, 95% CI 0.95 to 0.98) were statistically significant determinants of delayed IRoA in a model that also included age, BMI, ASA score and ISAR score. These associations did not depend on the time period during which the TKA took place, as assessed by tests for interaction. Functional mobility, as assessed pre-operatively by the TUG and DEMMI, is an independent and stable determinant of delayed inpatient recovery of activities after TKA. Future research, focusing on improvement of pre-operative functional mobility through tailored physiotherapy intervention, should indicate whether such intervention enhances post-operative recovery among high-risk patients. Cite this article: Bone Joint J 2017;99-B:211-17. ©2017 The British Editorial Society of Bone & Joint Surgery.

  19. Diversity of the total bacterial community associated with Ghanaian and Brazilian cocoa bean fermentation samples as revealed by a 16 S rRNA gene clone library.

    Science.gov (United States)

    Garcia-Armisen, Tamara; Papalexandratou, Zoi; Hendryckx, Hugo; Camu, Nicholas; Vrancken, Gino; De Vuyst, Luc; Cornelis, Pierre

    2010-08-01

    Cocoa bean fermentation is a spontaneous process involving a succession of microbial activities, starting with yeasts, followed by lactic acid bacteria and acetic acid bacteria. So far, all microbiological studies about cocoa bean fermentation were based on culture-dependent (isolation, cultivation, and identification), or, more recently, culture-independent (PCR-DGGE, or polymerase chain reaction denaturing gradient gel electrophoresis) methods. Using a metagenomic approach, total DNA was extracted from heap and box fermentations at different time points and from different locations (Ghana and Brazil, respectively) to generate a 16 S rDNA clone library that was sequenced. The sequencing data revealed a low bacterial diversity in the fermentation samples and were in accordance with the results obtained through culture-dependent and a second, culture-independent analysis (PCR-DGGE), suggesting that almost all bacteria involved in the fermentation process are cultivable. One exception was the identification by 16 S rDNA library sequencing of Gluconacetobacter species of acetic acid bacteria that were not detected by the two other approaches. The presence of Enterobacteriaceae related to Erwinia/Pantoea/Tatumella, as revealed by 16 S rDNA library sequencing, suggests an impact of these bacteria on fermentation.

  20. Different functions of the insect soluble and membrane-bound trehalase genes in chitin biosynthesis revealed by RNA interference.

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    Jie Chen

    Full Text Available BACKGROUND: Trehalase, an enzyme that hydrolyzes trehalose to yield two glucose molecules, plays a pivotal role in various physiological processes. In recent years, trehalase proteins have been purified from several insect species and are divided into soluble (Tre-1 and membrane-bound (Tre-2 trehalases. However, no functions of the two trehalases in chitin biosynthesis in insects have yet been reported. PRINCIPAL FINDINGS: The membrane-bound trehalase of Spodoptera exigua (SeTre-2 was characterized in our laboratory previously. In this study, we cloned the soluble trehalase gene (SeTre-1 and investigated the tissue distribution and developmental expression pattern of the two trehalase genes. SeTre-1 was expressed highly in cuticle and Malpighian tubules, while SeTre-2 was expressed in tracheae and fat body. In the midgut, the two trehalase genes were expressed in different locations. Additionally, the expression profiles of both trehalase mRNAs and their enzyme activities suggest that they may play different roles in chitin biosynthesis. The RNA interference (RNAi of either SeTre-1 or SeTre-2 was gene-specific and effective, with efficiency rates up to 83% at 72 h post injection. After RNAi of SeTre-1 and SeTre-2, significant higher mortality rates were observed during the larva-pupa stage and pupa-adult stage, and the lethal phenotypes were classified and analyzed. Additionally, the change trends of concentration of trehalose and glucose appeared reciprocally in RNAi-mutants. Moreover, knockdown of SeTre-1 gene largely inhibited the expression of chitin synthase gene A (CHSA and reduced the chitin content in the cuticle to two-thirds relative to the control insects. The chitin synthase gene B (CHSB expression, however, was inhibited more by the injection of dsRNA for SeTre-2, and the chitin content in the midgut decreased by about 25%. CONCLUSIONS: SeTre-1 plays a major role in CHSA expression and chitin synthesis in the cuticle, and SeTre-2