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Sample records for activator-dependent transcription initiation

  1. DREAM Controls the On/Off Switch of Specific Activity-Dependent Transcription Pathways

    Science.gov (United States)

    Mellström, Britt; Sahún, Ignasi; Ruiz-Nuño, Ana; Murtra, Patricia; Gomez-Villafuertes, Rosa; Savignac, Magali; Oliveros, Juan C.; Gonzalez, Paz; Kastanauskaite, Asta; Knafo, Shira; Zhuo, Min; Higuera-Matas, Alejandro; Errington, Michael L.; Maldonado, Rafael; DeFelipe, Javier; Jefferys, John G. R.; Bliss, Tim V. P.; Dierssen, Mara

    2014-01-01

    Changes in nuclear Ca2+ homeostasis activate specific gene expression programs and are central to the acquisition and storage of information in the brain. DREAM (downstream regulatory element antagonist modulator), also known as calsenilin/KChIP-3 (K+ channel interacting protein 3), is a Ca2+-binding protein that binds DNA and represses transcription in a Ca2+-dependent manner. To study the function of DREAM in the brain, we used transgenic mice expressing a Ca2+-insensitive/CREB-independent dominant active mutant DREAM (daDREAM). Using genome-wide analysis, we show that DREAM regulates the expression of specific activity-dependent transcription factors in the hippocampus, including Npas4, Nr4a1, Mef2c, JunB, and c-Fos. Furthermore, DREAM regulates its own expression, establishing an autoinhibitory feedback loop to terminate activity-dependent transcription. Ablation of DREAM does not modify activity-dependent transcription because of gene compensation by the other KChIP family members. The expression of daDREAM in the forebrain resulted in a complex phenotype characterized by loss of recurrent inhibition and enhanced long-term potentiation (LTP) in the dentate gyrus and impaired learning and memory. Our results indicate that DREAM is a major master switch transcription factor that regulates the on/off status of specific activity-dependent gene expression programs that control synaptic plasticity, learning, and memory. PMID:24366545

  2. Binding of TFIIIC to sine elements controls the relocation of activity-dependent neuronal genes to transcription factories.

    Directory of Open Access Journals (Sweden)

    Luca Crepaldi

    Full Text Available In neurons, the timely and accurate expression of genes in response to synaptic activity relies on the interplay between epigenetic modifications of histones, recruitment of regulatory proteins to chromatin and changes to nuclear structure. To identify genes and regulatory elements responsive to synaptic activation in vivo, we performed a genome-wide ChIPseq analysis of acetylated histone H3 using somatosensory cortex of mice exposed to novel enriched environmental (NEE conditions. We discovered that Short Interspersed Elements (SINEs located distal to promoters of activity-dependent genes became acetylated following exposure to NEE and were bound by the general transcription factor TFIIIC. Importantly, under depolarizing conditions, inducible genes relocated to transcription factories (TFs, and this event was controlled by TFIIIC. Silencing of the TFIIIC subunit Gtf3c5 in non-stimulated neurons induced uncontrolled relocation to TFs and transcription of activity-dependent genes. Remarkably, in cortical neurons, silencing of Gtf3c5 mimicked the effects of chronic depolarization, inducing a dramatic increase of both dendritic length and branching. These findings reveal a novel and essential regulatory function of both SINEs and TFIIIC in mediating gene relocation and transcription. They also suggest that TFIIIC may regulate the rearrangement of nuclear architecture, allowing the coordinated expression of activity-dependent neuronal genes.

  3. Binding of TFIIIC to sine elements controls the relocation of activity-dependent neuronal genes to transcription factories.

    Science.gov (United States)

    Crepaldi, Luca; Policarpi, Cristina; Coatti, Alessandro; Sherlock, William T; Jongbloets, Bart C; Down, Thomas A; Riccio, Antonella

    2013-01-01

    In neurons, the timely and accurate expression of genes in response to synaptic activity relies on the interplay between epigenetic modifications of histones, recruitment of regulatory proteins to chromatin and changes to nuclear structure. To identify genes and regulatory elements responsive to synaptic activation in vivo, we performed a genome-wide ChIPseq analysis of acetylated histone H3 using somatosensory cortex of mice exposed to novel enriched environmental (NEE) conditions. We discovered that Short Interspersed Elements (SINEs) located distal to promoters of activity-dependent genes became acetylated following exposure to NEE and were bound by the general transcription factor TFIIIC. Importantly, under depolarizing conditions, inducible genes relocated to transcription factories (TFs), and this event was controlled by TFIIIC. Silencing of the TFIIIC subunit Gtf3c5 in non-stimulated neurons induced uncontrolled relocation to TFs and transcription of activity-dependent genes. Remarkably, in cortical neurons, silencing of Gtf3c5 mimicked the effects of chronic depolarization, inducing a dramatic increase of both dendritic length and branching. These findings reveal a novel and essential regulatory function of both SINEs and TFIIIC in mediating gene relocation and transcription. They also suggest that TFIIIC may regulate the rearrangement of nuclear architecture, allowing the coordinated expression of activity-dependent neuronal genes.

  4. DNA replication initiator Cdc6 also regulates ribosomal DNA transcription initiation.

    Science.gov (United States)

    Huang, Shijiao; Xu, Xiaowei; Wang, Guopeng; Lu, Guoliang; Xie, Wenbing; Tao, Wei; Zhang, Hongyin; Jiang, Qing; Zhang, Chuanmao

    2016-04-01

    RNA-polymerase-I-dependent ribosomal DNA (rDNA) transcription is fundamental to rRNA processing, ribosome assembly and protein synthesis. However, how this process is initiated during the cell cycle is not fully understood. By performing a proteomic analysis of transcription factors that bind RNA polymerase I during rDNA transcription initiation, we identified that the DNA replication initiator Cdc6 interacts with RNA polymerase I and its co-factors, and promotes rDNA transcription in G1 phase in an ATPase-activity-dependent manner. We further showed that Cdc6 is targeted to the nucleolus during late mitosis and G1 phase in a manner that is dependent on B23 (also known as nucleophosmin, NPM1), and preferentially binds to the rDNA promoter through its ATP-binding domain. Overexpression of Cdc6 increases rDNA transcription, whereas knockdown of Cdc6 results in a decreased association of both RNA polymerase I and the RNA polymerase I transcription factor RRN3 with rDNA, and a reduction of rDNA transcription. Furthermore, depletion of Cdc6 impairs the interaction between RRN3 and RNA polymerase I. Taken together, our data demonstrate that Cdc6 also serves as a regulator of rDNA transcription initiation, and indicate a mechanism by which initiation of rDNA transcription and DNA replication can be coordinated in cells. © 2016. Published by The Company of Biologists Ltd.

  5. Structural Basis of Mitochondrial Transcription Initiation.

    Science.gov (United States)

    Hillen, Hauke S; Morozov, Yaroslav I; Sarfallah, Azadeh; Temiakov, Dmitry; Cramer, Patrick

    2017-11-16

    Transcription in human mitochondria is driven by a single-subunit, factor-dependent RNA polymerase (mtRNAP). Despite its critical role in both expression and replication of the mitochondrial genome, transcription initiation by mtRNAP remains poorly understood. Here, we report crystal structures of human mitochondrial transcription initiation complexes assembled on both light and heavy strand promoters. The structures reveal how transcription factors TFAM and TFB2M assist mtRNAP to achieve promoter-dependent initiation. TFAM tethers the N-terminal region of mtRNAP to recruit the polymerase to the promoter whereas TFB2M induces structural changes in mtRNAP to enable promoter opening and trapping of the DNA non-template strand. Structural comparisons demonstrate that the initiation mechanism in mitochondria is distinct from that in the well-studied nuclear, bacterial, or bacteriophage transcription systems but that similarities are found on the topological and conceptual level. These results provide a framework for studying the regulation of gene expression and DNA replication in mitochondria. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Transcription initiation complex structures elucidate DNA opening.

    Science.gov (United States)

    Plaschka, C; Hantsche, M; Dienemann, C; Burzinski, C; Plitzko, J; Cramer, P

    2016-05-19

    Transcription of eukaryotic protein-coding genes begins with assembly of the RNA polymerase (Pol) II initiation complex and promoter DNA opening. Here we report cryo-electron microscopy (cryo-EM) structures of yeast initiation complexes containing closed and open DNA at resolutions of 8.8 Å and 3.6 Å, respectively. DNA is positioned and retained over the Pol II cleft by a network of interactions between the TATA-box-binding protein TBP and transcription factors TFIIA, TFIIB, TFIIE, and TFIIF. DNA opening occurs around the tip of the Pol II clamp and the TFIIE 'extended winged helix' domain, and can occur in the absence of TFIIH. Loading of the DNA template strand into the active centre may be facilitated by movements of obstructing protein elements triggered by allosteric binding of the TFIIE 'E-ribbon' domain. The results suggest a unified model for transcription initiation with a key event, the trapping of open promoter DNA by extended protein-protein and protein-DNA contacts.

  7. Non-canonical transcription initiation: the expanding universe of transcription initiating substrates.

    Science.gov (United States)

    Barvík, Ivan; Rejman, Dominik; Panova, Natalya; Šanderová, Hana; Krásný, Libor

    2017-03-01

    RNA polymerase (RNAP) is the central enzyme of transcription of the genetic information from DNA into RNA. RNAP recognizes four main substrates: ATP, CTP, GTP and UTP. Experimental evidence from the past several years suggests that, besides these four NTPs, other molecules can be used to initiate transcription: (i) ribooligonucleotides (nanoRNAs) and (ii) coenzymes such as NAD+, NADH, dephospho-CoA and FAD. The presence of these molecules at the 5΄ ends of RNAs affects the properties of the RNA. Here, we discuss the expanding portfolio of molecules that can initiate transcription, their mechanism of incorporation, effects on RNA and cellular processes, and we present an outlook toward other possible initiation substrates. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  8. Non-canonical transcription initiation: the expanding universe of transcription initiating substrates

    Czech Academy of Sciences Publication Activity Database

    Barvík, I.; Rejman, Dominik; Panova, Natalya; Šanderová, Hana; Krásný, Libor

    2017-01-01

    Roč. 41, č. 2 (2017), s. 131-138 ISSN 0168-6445 R&D Projects: GA ČR GA15-05228S; GA ČR GA15-11711S Institutional support: RVO:61388963 ; RVO:61388971 Keywords : RNA polymerase * non-canonical transcription initiation * transcription initiating substrate * nicotinamide adenine dinucleotide (NAD(+)) * coenzymes * RNA stability Subject RIV: EB - Genetics ; Molecular Biology; EE - Microbiology, Virology (MBU-M) OBOR OECD: Biochemistry and molecular biology; Microbiology (MBU-M) Impact factor: 12.198, year: 2016

  9. Transcriptional response of zebrafish embryos exposed to neurotoxic compounds reveals a muscle activity dependent hspb11 expression.

    Directory of Open Access Journals (Sweden)

    Nils Klüver

    Full Text Available Acetylcholinesterase (AChE inhibitors are widely used as pesticides and drugs. Their primary effect is the overstimulation of cholinergic receptors which results in an improper muscular function. During vertebrate embryonic development nerve activity and intracellular downstream events are critical for the regulation of muscle fiber formation. Whether AChE inhibitors and related neurotoxic compounds also provoke specific changes in gene transcription patterns during vertebrate development that allow them to establish a mechanistic link useful for identification of developmental toxicity pathways has, however, yet not been investigated. Therefore we examined the transcriptomic response of a known AChE inhibitor, the organophosphate azinphos-methyl (APM, in zebrafish embryos and compared the response with two non-AChE inhibiting unspecific control compounds, 1,4-dimethoxybenzene (DMB and 2,4-dinitrophenol (DNP. A highly specific cluster of APM induced gene transcripts was identified and a subset of strongly regulated genes was analyzed in more detail. The small heat shock protein hspb11 was found to be the most sensitive induced gene in response to AChE inhibitors. Comparison of expression in wildtype, ache and sop(fixe mutant embryos revealed that hspb11 expression was dependent on the nicotinic acetylcholine receptor (nAChR activity. Furthermore, modulators of intracellular calcium levels within the whole embryo led to a transcriptional up-regulation of hspb11 which suggests that elevated intracellular calcium levels may regulate the expression of this gene. During early zebrafish development, hspb11 was specifically expressed in muscle pioneer cells and Hspb11 morpholino-knockdown resulted in effects on slow muscle myosin organization. Our findings imply that a comparative toxicogenomic approach and functional analysis can lead to the identification of molecular mechanisms and specific marker genes for potential neurotoxic compounds.

  10. CDK9-dependent RNA polymerase II pausing controls transcription initiation.

    Science.gov (United States)

    Gressel, Saskia; Schwalb, Björn; Decker, Tim Michael; Qin, Weihua; Leonhardt, Heinrich; Eick, Dirk; Cramer, Patrick

    2017-10-10

    Gene transcription can be activated by decreasing the duration of RNA polymerase II pausing in the promoter-proximal region, but how this is achieved remains unclear. Here we use a 'multi-omics' approach to demonstrate that the duration of polymerase pausing generally limits the productive frequency of transcription initiation in human cells ('pause-initiation limit'). We further engineer a human cell line to allow for specific and rapid inhibition of the P-TEFb kinase CDK9, which is implicated in polymerase pause release. CDK9 activity decreases the pause duration but also increases the productive initiation frequency. This shows that CDK9 stimulates release of paused polymerase and activates transcription by increasing the number of transcribing polymerases and thus the amount of mRNA synthesized per time. CDK9 activity is also associated with long-range chromatin interactions, suggesting that enhancers can influence the pause-initiation limit to regulate transcription.

  11. A code for transcription initiation in mammalian genomes

    DEFF Research Database (Denmark)

    Frith, Martin C.; Valen, Eivind Dale; Krogh, Anders

    2007-01-01

    that initiation events are clustered on the chromosomes at multiple scales - clusters within clusters - indicating multiple regulatory processes. Within the smallest of such clusters, which can be interpreted as core promoters, the local DNA sequence predicts the relative transcription start usage of each...... of large- and small-scale effects: the selection of transcription start sites is largely governed by the local DNA sequence, whereas the transcriptional activity of a locus is regulated at a different level; it is affected by distal features or events such as enhancers and chromatin remodeling....

  12. One nuclear calcium transient induced by a single burst of action potentials represents the minimum signal strength in activity-dependent transcription in hippocampal neurons.

    Science.gov (United States)

    Yu, Yan; Oberlaender, Kristin; Bengtson, C Peter; Bading, Hilmar

    2017-07-01

    Neurons undergo dramatic changes in their gene expression profiles in response to synaptic stimulation. The coupling of neuronal excitation to gene transcription is well studied and is mediated by signaling pathways activated by cytoplasmic and nuclear calcium transients. Despite this, the minimum synaptic activity required to induce gene expression remains unknown. To address this, we used cultured hippocampal neurons and cellular compartment analysis of temporal activity by fluorescence in situ hybridization (catFISH) that allows detection of nascent transcripts in the cell nucleus. We found that a single burst of action potentials, consisting of 24.4±5.1 action potentials during a 6.7±1.9s depolarization of 19.5±2.0mV causing a 9.3±0.9s somatic calcium transient, is sufficient to activate transcription of the immediate early gene arc (also known as Arg3.1). The total arc mRNA yield produced after a single burst-induced nuclear calcium transient was very small and, compared to unstimulated control neurons, did not lead to a significant increase in arc mRNA levels measured using quantitative reverse transcriptase PCR (qRT-PCR) of cell lysates. Significantly increased arc mRNA levels became detectable in hippocampal neurons that had undergone 5-8 consecutive burst-induced nuclear calcium transients at 0.05-0.15Hz. These results indicate that a single burst-induced nuclear calcium transient can activate gene expression and that transcription is rapidly shut off after synaptic stimulation has ceased. Copyright © 2017 Elsevier Ltd. All rights reserved.

  13. Dissecting Cell-Type Composition and Activity-Dependent Transcriptional State in Mammalian Brains by Massively Parallel Single-Nucleus RNA-Seq.

    Science.gov (United States)

    Hu, Peng; Fabyanic, Emily; Kwon, Deborah Y; Tang, Sheng; Zhou, Zhaolan; Wu, Hao

    2017-12-07

    Massively parallel single-cell RNA sequencing can precisely resolve cellular diversity in a high-throughput manner at low cost, but unbiased isolation of intact single cells from complex tissues such as adult mammalian brains is challenging. Here, we integrate sucrose-gradient-assisted purification of nuclei with droplet microfluidics to develop a highly scalable single-nucleus RNA-seq approach (sNucDrop-seq), which is free of enzymatic dissociation and nucleus sorting. By profiling ∼18,000 nuclei isolated from cortical tissues of adult mice, we demonstrate that sNucDrop-seq not only accurately reveals neuronal and non-neuronal subtype composition with high sensitivity but also enables in-depth analysis of transient transcriptional states driven by neuronal activity, at single-cell resolution, in vivo. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. 5' diversity of human hepatic PXR (NR1I2) transcripts and identification of the major transcription initiation site.

    Science.gov (United States)

    Kurose, Kouichi; Koyano, Satoru; Ikeda, Shinobu; Tohkin, Masahiro; Hasegawa, Ryuichi; Sawada, Jun-Ichi

    2005-05-01

    The human pregnane X receptor (PXR) is a crucial regulator of the genes encoding several major cytochrome P450 enzymes and transporters, such as CYP3A4 and MDR1, but its own transcriptional regulation remains unclear. To elucidate the transcriptional mechanisms of human PXR gene, we first endeavored to identify the transcription initiation site of human PXR using 5'-RACE. Five types of 5'-variable transcripts (a, b, c, d, and e) with common exon 2 sequence were found, and comparison of these sequences with the genomic sequence suggested that their 5' diversity is derived from initiation by alternative promoters and alternative splicing. None of the exons found in our study contain any new in-frame coding regions. Newly identified introns IVS-a and IVS-b were found to have CT-AC splice sites that do not follow the GT-AG rule of conventional donor and acceptor splice sites. Of the five types of 5' variable transcripts identified, RT-PCR showed that type-a was the major transcript type. Four transcription initiation sites (A-D) for type-a transcript were identified by 5'-RACE using GeneRacer RACE Ready cDNA (human liver) constructed by the oligo-capping method. Putative TATA boxes were located approximately 30 bp upstream from the transcriptional start sites of the major transcript (C) and the longest minor transcript (A) expressed in the human liver. These results indicate that the initiation of transcription of human PXR is more complex than previously reported.

  15. HIV-1 reverse transcription initiation: a potential target for novel antivirals?

    NARCIS (Netherlands)

    Abbink, Truus E. M.; Berkhout, Ben

    2008-01-01

    Reverse transcription is an essential step in the retroviral life cycle, as it converts the genomic RNA into DNA. In this review, we describe recent developments concerning the initiation step of this complex, multi-step reaction. During initiation of reverse transcription, a cellular tRNA primer is

  16. In vitro transcription in the presence of DNA oligonucleotides can generate strong anomalous initiation sites.

    Science.gov (United States)

    Chow, C W; Clark, M P; Rinaldo, J E; Chalkley, R

    1996-03-01

    In the present study, we have explored an unexpected observation in transcription initiation that is mediated by single-stranded oligonucleotides. Initially, our goal was to understand the function of different upstream regulatory elements/initiation sites in the rat xanthine dehydrogenase/oxidase (XDH/XO) promoter. We performed in vitro transcription with HeLa nuclear extracts in the presence of different double-stranded oligonucleotides against upstream elements as competitors. A new and unusual transcription initiation site was detected by primer extension. This new initiation site maps to the downstream region of the corresponding competitor. Subsequent analyses have indicated that the induction of a new transcription initiation site is anomalous which is due to the presence of a small amount of single-stranded oligonucleotide in the competitor. We found that this anomalous initiation site is insensitive to the orientation of the promoter and requires only a small amount of single-stranded oligonucleotide (< 2-fold molar excess relative to template). We surmise that a complementary interaction between the single-stranded oligonucleotide and transiently denatured promoter template may be responsible for this sequence-specific transcription initiation artifact. To study the regulation of transcription initiation by in vitro transcription approaches, we propose that one should probe the effect of removing transacting factors by adding an excess of a cognate oligonucleotide which does not bear exact sequence identity to the template.

  17. Principles for RNA metabolism and alternative transcription initiation within closely spaced promoters

    DEFF Research Database (Denmark)

    Chen, Yun; Pai, Athma A; Herudek, Jan

    2016-01-01

    Mammalian transcriptomes are complex and formed by extensive promoter activity. In addition, gene promoters are largely divergent and initiate transcription of reverse-oriented promoter upstream transcripts (PROMPTs). Although PROMPTs are commonly terminated early, influenced by polyadenylation s...... suggest that basic building blocks of divergently transcribed core promoter pairs, in combination with the wealth of TSSs in mammalian genomes, provide a framework with which evolution shapes transcriptomes.......Mammalian transcriptomes are complex and formed by extensive promoter activity. In addition, gene promoters are largely divergent and initiate transcription of reverse-oriented promoter upstream transcripts (PROMPTs). Although PROMPTs are commonly terminated early, influenced by polyadenylation...

  18. A 5' splice site enhances the recruitment of basal transcription initiation factors in vivo

    DEFF Research Database (Denmark)

    Damgaard, Christian Kroun; Kahns, Søren; Lykke-Andersen, Søren

    2008-01-01

    RNAs, harboring wild-type or various 5′ splice site mutations, we demonstrate a strong positive correlation between splicing efficiency and transcription activity. Interestingly, a 5′ splice site can stimulate transcription even in the absence of splicing. Chromatin immunoprecipitation experiments show enhanced...... a promoter-proximal 5′ splice site via its U1 snRNA interaction can feed back to stimulate transcription initiation by enhancing preinitiation complex assembly....

  19. A transcript finishing initiative for closing gaps in the human transcriptome

    DEFF Research Database (Denmark)

    Sogayar, Mari Cleide; Camargo, Anamaria A; Bettoni, Fabiana

    2004-01-01

    We report the results of a transcript finishing initiative, undertaken for the purpose of identifying and characterizing novel human transcripts, in which RT-PCR was used to bridge gaps between paired EST clusters, mapped against the genomic sequence. Each pair of EST clusters selected...

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

    NARCIS (Netherlands)

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

    2013-01-01

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

  1. Role of the σ54 Activator Interacting Domain in Bacterial Transcription Initiation

    Energy Technology Data Exchange (ETDEWEB)

    Siegel, Alexander R. [Univ. of California, Berkeley, CA (United States); Wemmer, David E. [Univ. of California, Berkeley, CA (United States)

    2016-10-11

    Bacterial sigma factors are subunits of RNA polymerase that direct the holoenzyme to specific sets of promoters in the genome and are a central element of regulating transcription. Most polymerase holoenzymes open the promoter and initiate transcription rapidly after binding. However, polymerase containing the members of the σ54 family must be acted on by a transcriptional activator before DNA opening and initiation occur. A key domain in these transcriptional activators forms a hexameric AAA + ATPase that acts through conformational changes brought on by ATP hydrolysis. Contacts between the transcriptional activator and σ54 are primarily made through an N-terminal σ54 activator interacting domain (AID). To better understand this mechanism of bacterial transcription initiation, we characterized the σ54 AID by NMR spectroscopy and other biophysical methods and show that it is an intrinsically disordered domain in σ54 alone. In this paper, we identified a minimal construct of the Aquifex aeolicus σ54 AID that consists of two predicted helices and retains native-like binding affinity for the transcriptional activator NtrC1. Using the NtrC1 ATPase domain, bound with the non-hydrolyzable ATP analog ADP-beryllium fluoride, we studied the NtrC1–σ54 AID complex using NMR spectroscopy. We show that the σ54 AID becomes structured after associating with the core loops of the transcriptional activators in their ATP state and that the primary site of the interaction is the first predicted helix. Finally, understanding this complex, formed as the first step toward initiation, will help unravel the mechanism of σ54 bacterial transcription initiation.

  2. Real-time observation of the initiation of RNA polymerase II transcription.

    Science.gov (United States)

    Fazal, Furqan M; Meng, Cong A; Murakami, Kenji; Kornberg, Roger D; Block, Steven M

    2015-09-10

    Biochemical and structural studies have shown that the initiation of RNA polymerase II transcription proceeds in the following stages: assembly of the polymerase with general transcription factors and promoter DNA in a 'closed' preinitiation complex (PIC); unwinding of about 15 base pairs of the promoter DNA to form an 'open' complex; scanning downstream to a transcription start site; synthesis of a short transcript, thought to be about 10 nucleotides long; and promoter escape. Here we have assembled a 32-protein, 1.5-megadalton PIC derived from Saccharomyces cerevisiae, and observe subsequent initiation processes in real time with optical tweezers. Contrary to expectation, scanning driven by the transcription factor IIH involved the rapid opening of an extended transcription bubble, averaging 85 base pairs, accompanied by the synthesis of a transcript up to the entire length of the extended bubble, followed by promoter escape. PICs that failed to achieve promoter escape nevertheless formed open complexes and extended bubbles, which collapsed back to closed or open complexes, resulting in repeated futile scanning.

  3. Eukaryotic Initiation Factor 4H Is under Transcriptional Control of p65/NF-κB

    Science.gov (United States)

    Fiume, Giuseppe; Rossi, Annalisa; de Laurentiis, Annamaria; Falcone, Cristina; Pisano, Antonio; Vecchio, Eleonora; Pontoriero, Marilena; Scala, Iris; Scialdone, Annarita; Masci, Francesca Fasanella; Mimmi, Selena; Palmieri, Camillo; Scala, Giuseppe; Quinto, Ileana

    2013-01-01

    Protein synthesis is mainly regulated at the initiation step, allowing the fast, reversible and spatial control of gene expression. Initiation of protein synthesis requires at least 13 translation initiation factors to assemble the 80S ribosomal initiation complex. Loss of translation control may result in cell malignant transformation. Here, we asked whether translational initiation factors could be regulated by NF-κB transcription factor, a major regulator of genes involved in cell proliferation, survival, and inflammatory response. We show that the p65 subunit of NF-κB activates the transcription of eIF4H gene, which is the regulatory subunit of eIF4A, the most relevant RNA helicase in translation initiation. The p65-dependent transcriptional activation of eIF4H increased the eIF4H protein content augmenting the rate of global protein synthesis. In this context, our results provide novel insights into protein synthesis regulation in response to NF-κB activation signalling, suggesting a transcription-translation coupled mechanism of control. PMID:23776612

  4. Factor C*, the specific initiation component of the mouse RNA polymerase I holoenzyme, is inactivated early in the transcription process.

    OpenAIRE

    Brun, R P; Ryan, K; Sollner-Webb, B

    1994-01-01

    Factor C* is the component of the RNA polymerase I holoenzyme (factor C) that allows specific transcriptional initiation on a factor D (SL1)- and UBF-activated rRNA gene promoter. The in vitro transcriptional capacity of a preincubated rDNA promoter complex becomes exhausted very rapidly upon initiation of transcription. This is due to the rapid depletion of C* activity. In contrast, C* activity is not unstable in the absence of transcription, even in the presence of nucleoside triphosphates ...

  5. Correlation of Meiotic DSB Formation and Transcription Initiation Around Fission Yeast Recombination Hotspots.

    Science.gov (United States)

    Yamada, Shintaro; Okamura, Mika; Oda, Arisa; Murakami, Hiroshi; Ohta, Kunihiro; Yamada, Takatomi

    2017-06-01

    Meiotic homologous recombination, a critical event for ensuring faithful chromosome segregation and creating genetic diversity, is initiated by programmed DNA double-strand breaks (DSBs) formed at recombination hotspots. Meiotic DSB formation is likely to be influenced by other DNA-templated processes including transcription, but how DSB formation and transcription interact with each other has not been understood well. In this study, we used fission yeast to investigate a possible interplay of these two events. A group of hotspots in fission yeast are associated with sequences similar to the cyclic AMP response element and activated by the ATF/CREB family transcription factor dimer Atf1-Pcr1. We first focused on one of those hotspots, ade6-3049 , and Atf1. Our results showed that multiple transcripts, shorter than the ade6 full-length messenger RNA, emanate from a region surrounding the ade6-3049 hotspot. Interestingly, we found that the previously known recombination-activation region of Atf1 is also a transactivation domain, whose deletion affected DSB formation and short transcript production at ade6-3049 These results point to a possibility that the two events may be related to each other at ade6-3049 In fact, comparison of published maps of meiotic transcripts and hotspots suggested that hotspots are very often located close to meiotically transcribed regions. These observations therefore propose that meiotic DSB formation in fission yeast may be connected to transcription of surrounding regions. Copyright © 2017 by the Genetics Society of America.

  6. Transcription of ribosomal RNA genes is initiated in the third cell cycle of bovine embryos

    DEFF Research Database (Denmark)

    Jakobsen, Anne Sørig; Avery, Birthe; Dieleman, Steph J.

    2006-01-01

    Transcription from the embryos own ribosomal genes is initiated in most species at the same time as the maternal-embryonic transition. Recently data have indicated that a minor activation may take place during the third embryonic cell cycle in the bovine, one cell cycle before the major activation...

  7. X-ray Crystal Structures Elucidate the Nucleotidyl Transfer Reaction of Transcript Initiation Using Two Nucleotides

    Energy Technology Data Exchange (ETDEWEB)

    M Gleghorn; E Davydova; R Basu; L Rothman-Denes; K Murakami

    2011-12-31

    We have determined the X-ray crystal structures of the pre- and postcatalytic forms of the initiation complex of bacteriophage N4 RNA polymerase that provide the complete set of atomic images depicting the process of transcript initiation by a single-subunit RNA polymerase. As observed during T7 RNA polymerase transcript elongation, substrate loading for the initiation process also drives a conformational change of the O helix, but only the correct base pairing between the +2 substrate and DNA base is able to complete the O-helix conformational transition. Substrate binding also facilitates catalytic metal binding that leads to alignment of the reactive groups of substrates for the nucleotidyl transfer reaction. Although all nucleic acid polymerases use two divalent metals for catalysis, they differ in the requirements and the timing of binding of each metal. In the case of bacteriophage RNA polymerase, we propose that catalytic metal binding is the last step before the nucleotidyl transfer reaction.

  8. Regulation, initiation, and termination of the cenA and cex transcripts of Cellulomonas fimi

    International Nuclear Information System (INIS)

    Greenberg, N.M.; Warren, R.A.J.; Kilburn, D.G.; Miller, R.C. Jr.

    1987-01-01

    The authors characterized the in vivo transcripts of two Cellulomonas fimi genes, which encodes an extracellular endo-β-1,4-glucanase. By Northern blot analysis, cenA mRNA was detected in C. fimi RNA preparations from glycerol- and carboxymethyl cellulose-grown cells but not from glucose-grown cells. In contrast, cex mRNA was detected only in the preparations from carboxymethyl cellulose-grown cells. Therefore, the transcription of these genes is subject to regulation by the carbon source provided to C. fimi. By nuclease SI protection studies with unique 5'-labeled DNA probes and C. fimi RNA isolated in vivo, 5' termini were found 51 and 62 bases before the cenA translational initiation codon and 28 bases before the cex translational initiation codon. S1 mapping with unlabeled DNA probes and C. fimi RNA which had been isolated in vivo but which had been 5' labeled in vitro with guanylyltransferase and [α- 32 P]GTP confirmed that true transcription initiation sites for cenA and cex mRNA had been identified. Comparative analysis of the DNA sequences immediately upstream of the initiation sites of the cenA and cex mRNAs revealed a 30-base-pair region where these two sequences display at least 66% homology. S1 mapping was also used to locate the 3' termini of the cenA and cex transcripts. Three 3' termini were found for cenA messages, whereas only one 3' terminus was identified for cex mRNA. The transcripts of both genes terminate in regions where their corresponding DNA sequences contain inverted repeats

  9. DNA dynamics play a role as a basal transcription factor in the positioning and regulation of gene transcription initiation

    OpenAIRE

    Alexandrov, Boian S.; Gelev, Vladimir; Yoo, Sang Wook; Alexandrov, Ludmil B.; Fukuyo, Yayoi; Bishop, Alan R.; Rasmussen, Kim ?.; Usheva, Anny

    2009-01-01

    We assess the role of DNA breathing dynamics as a determinant of promoter strength and transcription start site (TSS) location. We compare DNA Langevin dynamic profiles of representative gene promoters, calculated with the extended non-linear PBD model of DNA with experimental data on transcription factor binding and transcriptional activity. Our results demonstrate that DNA dynamic activity at the TSS can be suppressed by mutations that do not affect basal transcription factor binding–DNA co...

  10. Structures of RNA Polymerase Closed and Intermediate Complexes Reveal Mechanisms of DNA Opening and Transcription Initiation.

    Science.gov (United States)

    Glyde, Robert; Ye, Fuzhou; Darbari, Vidya Chandran; Zhang, Nan; Buck, Martin; Zhang, Xiaodong

    2017-07-06

    Gene transcription is carried out by RNA polymerases (RNAPs). For transcription to occur, the closed promoter complex (RPc), where DNA is double stranded, must isomerize into an open promoter complex (RPo), where the DNA is melted out into a transcription bubble and the single-stranded template DNA is delivered to the RNAP active site. Using a bacterial RNAP containing the alternative σ 54 factor and cryoelectron microscopy, we determined structures of RPc and the activator-bound intermediate complex en route to RPo at 3.8 and 5.8 Å. Our structures show how RNAP-σ 54 interacts with promoter DNA to initiate the DNA distortions required for transcription bubble formation, and how the activator interacts with RPc, leading to significant conformational changes in RNAP and σ 54 that promote RPo formation. We propose that DNA melting is an active process initiated in RPc and that the RNAP conformations of intermediates are significantly different from that of RPc and RPo. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  11. Structures of transcription pre-initiation complex with TFIIH and Mediator.

    Science.gov (United States)

    Schilbach, S; Hantsche, M; Tegunov, D; Dienemann, C; Wigge, C; Urlaub, H; Cramer, P

    2017-11-09

    For the initiation of transcription, RNA polymerase II (Pol II) assembles with general transcription factors on promoter DNA to form the pre-initiation complex (PIC). Here we report cryo-electron microscopy structures of the Saccharomyces cerevisiae PIC and PIC-core Mediator complex at nominal resolutions of 4.7 Å and 5.8 Å, respectively. The structures reveal transcription factor IIH (TFIIH), and suggest how the core and kinase TFIIH modules function in the opening of promoter DNA and the phosphorylation of Pol II, respectively. The TFIIH core subunit Ssl2 (a homologue of human XPB) is positioned on downstream DNA by the 'E-bridge' helix in TFIIE, consistent with TFIIE-stimulated DNA opening. The TFIIH kinase module subunit Tfb3 (MAT1 in human) anchors the kinase Kin28 (CDK7), which is mobile in the PIC but preferentially located between the Mediator hook and shoulder in the PIC-core Mediator complex. Open spaces between the Mediator head and middle modules may allow access of the kinase to its substrate, the C-terminal domain of Pol II.

  12. ITPI: Initial Transcription Process-Based Identification Method of Bioactive Components in Traditional Chinese Medicine Formula

    Directory of Open Access Journals (Sweden)

    Baixia Zhang

    2016-01-01

    Full Text Available Identification of bioactive components is an important area of research in traditional Chinese medicine (TCM formula. The reported identification methods only consider the interaction between the components and the target proteins, which is not sufficient to explain the influence of TCM on the gene expression. Here, we propose the Initial Transcription Process-based Identification (ITPI method for the discovery of bioactive components that influence transcription factors (TFs. In this method, genome-wide chip detection technology was used to identify differentially expressed genes (DEGs. The TFs of DEGs were derived from GeneCards. The components influencing the TFs were derived from STITCH. The bioactive components in the formula were identified by evaluating the molecular similarity between the components in formula and the components that influence the TF of DEGs. Using the formula of Tian-Zhu-San (TZS as an example, the reliability and limitation of ITPI were examined and 16 bioactive components that influence TFs were identified.

  13. The relationship between transcription initiation RNAs and CCCTC-binding factor (CTCF localization

    Directory of Open Access Journals (Sweden)

    Taft Ryan J

    2011-08-01

    Full Text Available Abstract Background Transcription initiation RNAs (tiRNAs are nuclear localized 18 nucleotide RNAs derived from sequences immediately downstream of RNA polymerase II (RNAPII transcription start sites. Previous reports have shown that tiRNAs are intimately correlated with gene expression, RNA polymerase II binding and behaviors, and epigenetic marks associated with transcription initiation, but not elongation. Results In the present work, we show that tiRNAs are commonly found at genomic CCCTC-binding factor (CTCF binding sites in human and mouse, and that CTCF sites that colocalize with RNAPII are highly enriched for tiRNAs. To directly investigate the relationship between tiRNAs and CTCF we examined tiRNAs originating near the intronic CTCF binding site in the human tumor suppressor gene, p21 (cyclin-dependent kinase inhibitor 1A gene, also known as CDKN1A. Inhibition of CTCF-proximal tiRNAs resulted in increased CTCF localization and increased p21 expression, while overexpression of CTCF-proximal tiRNA mimics decreased CTCF localization and p21 expression. We also found that tiRNA-regulated CTCF binding influences the levels of trimethylated H3K27 at the alternate upstream p21 promoter, and affects the levels of alternate p21 (p21alt transcripts. Extending these studies to another randomly selected locus with conserved CTCF binding we found that depletion of tiRNA alters nucleosome density proximal to sites of tiRNA biogenesis. Conclusions Taken together, these data suggest that tiRNAs modulate local epigenetic structure, which in turn regulates CTCF localization.

  14. Transcription initiation patterns indicate divergent strategies for gene regulation at the chromatin level.

    Directory of Open Access Journals (Sweden)

    Elizabeth A Rach

    2011-01-01

    Full Text Available The application of deep sequencing to map 5' capped transcripts has confirmed the existence of at least two distinct promoter classes in metazoans: "focused" promoters with transcription start sites (TSSs that occur in a narrowly defined genomic span and "dispersed" promoters with TSSs that are spread over a larger window. Previous studies have explored the presence of genomic features, such as CpG islands and sequence motifs, in these promoter classes, but virtually no studies have directly investigated the relationship with chromatin features. Here, we show that promoter classes are significantly differentiated by nucleosome organization and chromatin structure. Dispersed promoters display higher associations with well-positioned nucleosomes downstream of the TSS and a more clearly defined nucleosome free region upstream, while focused promoters have a less organized nucleosome structure, yet higher presence of RNA polymerase II. These differences extend to histone variants (H2A.Z and marks (H3K4 methylation, as well as insulator binding (such as CTCF, independent of the expression levels of affected genes. Notably, differences are conserved across mammals and flies, and they provide for a clearer separation of promoter architectures than the presence and absence of CpG islands or the occurrence of stalled RNA polymerase. Computational models support the stronger contribution of chromatin features to the definition of dispersed promoters compared to focused start sites. Our results show that promoter classes defined from 5' capped transcripts not only reflect differences in the initiation process at the core promoter but also are indicative of divergent transcriptional programs established within gene-proximal nucleosome organization.

  15. Study of the variation of the nuclear transcriptional map during de initial development of Drosophyla melanogaster embryos

    International Nuclear Information System (INIS)

    Alonso, C.E.V.

    1987-01-01

    The variation of nuclear transcriptional map during the initial development of Drosophyla melanogaster embryos were studied. Thermic treatment, chromatographic techniques and liquid scintilation in embryos inoculated with radioactive uridine were used. (L.J.C.)

  16. A structural model of the E. coli PhoB Dimer in the transcription initiation complex

    Directory of Open Access Journals (Sweden)

    Tung Chang-Shung

    2012-03-01

    Full Text Available Abstract Background There exist > 78,000 proteins and/or nucleic acids structures that were determined experimentally. Only a small portion of these structures corresponds to those of protein complexes. While homology modeling is able to exploit knowledge-based potentials of side-chain rotomers and backbone motifs to infer structures for new proteins, no such general method exists to extend our understanding of protein interaction motifs to novel protein complexes. Results We use a Motif Binding Geometries (MBG approach, to infer the structure of a protein complex from the database of complexes of homologous proteins taken from other contexts (such as the helix-turn-helix motif binding double stranded DNA, and demonstrate its utility on one of the more important regulatory complexes in biology, that of the RNA polymerase initiating transcription under conditions of phosphate starvation. The modeled PhoB/RNAP/σ-factor/DNA complex is stereo-chemically reasonable, has sufficient interfacial Solvent Excluded Surface Areas (SESAs to provide adequate binding strength, is physically meaningful for transcription regulation, and is consistent with a variety of known experimental constraints. Conclusions Based on a straightforward and easy to comprehend concept, "proteins and protein domains that fold similarly could interact similarly", a structural model of the PhoB dimer in the transcription initiation complex has been developed. This approach could be extended to enable structural modeling and prediction of other bio-molecular complexes. Just as models of individual proteins provide insight into molecular recognition, catalytic mechanism, and substrate specificity, models of protein complexes will provide understanding into the combinatorial rules of cellular regulation and signaling.

  17. Core Mediator structure at 3.4 Å extends model of transcription initiation complex.

    Science.gov (United States)

    Nozawa, Kayo; Schneider, Thomas R; Cramer, Patrick

    2017-05-11

    Mediator is a multiprotein co-activator that binds the transcription pre-initiation complex (PIC) and regulates RNA polymerase (Pol) II. The Mediator head and middle modules form the essential core Mediator (cMed), whereas the tail and kinase modules play regulatory roles. The architecture of Mediator and its position on the PIC are known, but atomic details are limited to Mediator subcomplexes. Here we report the crystal structure of the 15-subunit cMed from Schizosaccharomyces pombe at 3.4 Å resolution. The structure shows an unaltered head module, and reveals the intricate middle module, which we show is globally required for transcription. Sites of known Mediator mutations cluster at the interface between the head and middle modules, and in terminal regions of the head subunits Med6 (ref. 16) and Med17 (ref. 17) that tether the middle module. The structure led to a model for Saccharomyces cerevisiae cMed that could be combined with the 3.6 Å cryo-electron microscopy structure of the core PIC (cPIC). The resulting atomic model of the cPIC-cMed complex informs on interactions of the submodules forming the middle module, called beam, knob, plank, connector, and hook. The hook is flexibly linked to Mediator by a conserved hinge and contacts the transcription initiation factor IIH (TFIIH) kinase that phosphorylates the carboxy (C)-terminal domain (CTD) of Pol II and was recently positioned on the PIC. The hook also contains residues that crosslink to the CTD and reside in a previously described cradle. These results provide a framework for understanding Mediator function, including its role in stimulating CTD phosphorylation by TFIIH.

  18. Full-length mRNA sequencing uncovers a widespread coupling between transcription initiation and mRNA processing.

    Science.gov (United States)

    Anvar, Seyed Yahya; Allard, Guy; Tseng, Elizabeth; Sheynkman, Gloria M; de Klerk, Eleonora; Vermaat, Martijn; Yin, Raymund H; Johansson, Hans E; Ariyurek, Yavuz; den Dunnen, Johan T; Turner, Stephen W; 't Hoen, Peter A C

    2018-03-29

    The multifaceted control of gene expression requires tight coordination of regulatory mechanisms at transcriptional and post-transcriptional level. Here, we studied the interdependence of transcription initiation, splicing and polyadenylation events on single mRNA molecules by full-length mRNA sequencing. In MCF-7 breast cancer cells, we find 2700 genes with interdependent alternative transcription initiation, splicing and polyadenylation events, both in proximal and distant parts of mRNA molecules, including examples of coupling between transcription start sites and polyadenylation sites. The analysis of three human primary tissues (brain, heart and liver) reveals similar patterns of interdependency between transcription initiation and mRNA processing events. We predict thousands of novel open reading frames from full-length mRNA sequences and obtained evidence for their translation by shotgun proteomics. The mapping database rescues 358 previously unassigned peptides and improves the assignment of others. By recognizing sample-specific amino-acid changes and novel splicing patterns, full-length mRNA sequencing improves proteogenomics analysis of MCF-7 cells. Our findings demonstrate that our understanding of transcriptome complexity is far from complete and provides a basis to reveal largely unresolved mechanisms that coordinate transcription initiation and mRNA processing.

  19. Structural basis for the initiation of eukaryotic transcription-coupled DNA repair.

    Science.gov (United States)

    Xu, Jun; Lahiri, Indrajit; Wang, Wei; Wier, Adam; Cianfrocco, Michael A; Chong, Jenny; Hare, Alissa A; Dervan, Peter B; DiMaio, Frank; Leschziner, Andres E; Wang, Dong

    2017-11-30

    Eukaryotic transcription-coupled repair (TCR) is an important and well-conserved sub-pathway of nucleotide excision repair that preferentially removes DNA lesions from the template strand that block translocation of RNA polymerase II (Pol II). Cockayne syndrome group B (CSB, also known as ERCC6) protein in humans (or its yeast orthologues, Rad26 in Saccharomyces cerevisiae and Rhp26 in Schizosaccharomyces pombe) is among the first proteins to be recruited to the lesion-arrested Pol II during the initiation of eukaryotic TCR. Mutations in CSB are associated with the autosomal-recessive neurological disorder Cockayne syndrome, which is characterized by progeriod features, growth failure and photosensitivity. The molecular mechanism of eukaryotic TCR initiation remains unclear, with several long-standing unanswered questions. How cells distinguish DNA lesion-arrested Pol II from other forms of arrested Pol II, the role of CSB in TCR initiation, and how CSB interacts with the arrested Pol II complex are all unknown. The lack of structures of CSB or the Pol II-CSB complex has hindered our ability to address these questions. Here we report the structure of the S. cerevisiae Pol II-Rad26 complex solved by cryo-electron microscopy. The structure reveals that Rad26 binds to the DNA upstream of Pol II, where it markedly alters its path. Our structural and functional data suggest that the conserved Swi2/Snf2-family core ATPase domain promotes the forward movement of Pol II, and elucidate key roles for Rad26 in both TCR and transcription elongation.

  20. Frequent dual initiation of reverse transcription in murine leukemia virus-based vectors containing two primer-binding sites

    International Nuclear Information System (INIS)

    Voronin, Yegor A.; Pathak, Vinay K.

    2003-01-01

    Retroviruses package two copies of viral RNA into each virion. Although each RNA contains a primer-binding site for initiation of DNA synthesis, it is unknown whether reverse transcription is initiated on both RNAs. To determine whether a single virion is capable of initiating reverse transcription more than once, we constructed a murine leukemia virus-based vector containing a second primer-binding site (PBS) derived from spleen necrosis virus and inserted the green fluorescent protein gene (GFP) between the two PBSs. Initiation of reverse transcription at either PBS results in a provirus that expresses GFP. However, initiation at both PBSs can result in the deletion of GFP, which can be detected by flow cytometry and Southern blotting analysis. Approximately 22-29% of the proviruses formed deleted the GFP in a single replication cycle, indicating the minimum proportion of virions that initiated reverse transcription on both PBSs. These results show that a significant proportion of MLV-based vectors containing two PBSs have the capacity to initiate reverse transcription more than once

  1. Core Promoter Plasticity Between Maize Tissues and Genotypes Contrasts with Predominance of Sharp Transcription Initiation Sites.

    Science.gov (United States)

    Mejía-Guerra, María Katherine; Li, Wei; Galeano, Narmer F; Vidal, Mabel; Gray, John; Doseff, Andrea I; Grotewold, Erich

    2015-12-01

    Core promoters are crucial for gene regulation, providing blueprints for the assembly of transcriptional machinery at transcription start sites (TSSs). Empirically, TSSs define the coordinates of core promoters and other regulatory sequences. Thus, experimental TSS identification provides an essential step in the characterization of promoters and their features. Here, we describe the application of CAGE (cap analysis of gene expression) to identify genome-wide TSSs used in root and shoot tissues of two maize (Zea mays) inbred lines (B73 and Mo17). Our studies indicate that most TSS clusters are sharp in maize, similar to mice, but distinct from Arabidopsis thaliana, Drosophila melanogaster, or zebra fish, in which a majority of genes have broad-shaped TSS clusters. We established that ∼38% of maize promoters are characterized by a broader TATA-motif consensus, and this motif is significantly enriched in genes with sharp TSSs. A noteworthy plasticity in TSS usage between tissues and inbreds was uncovered, with ∼1500 genes showing significantly different dominant TSSs, sometimes affecting protein sequence by providing alternate translation initiation codons. We experimentally characterized instances in which this differential TSS utilization results in protein isoforms with additional domains or targeted to distinct subcellular compartments. These results provide important insights into TSS selection and gene expression in an agronomically important crop. © 2015 American Society of Plant Biologists. All rights reserved.

  2. Structural insights into the mycobacteria transcription initiation complex from analysis of X-ray crystal structures

    Energy Technology Data Exchange (ETDEWEB)

    Hubin, Elizabeth A.; Lilic, Mirjana; Darst, Seth A.; Campbell, Elizabeth A.

    2017-07-13

    The mycobacteria RNA polymerase (RNAP) is a target for antimicrobials against tuberculosis, motivating structure/function studies. Here we report a 3.2 Å-resolution crystal structure of a Mycobacterium smegmatis (Msm) open promoter complex (RPo), along with structural analysis of the Msm RPo and a previously reported 2.76 Å-resolution crystal structure of an Msm transcription initiation complex with a promoter DNA fragment. We observe the interaction of the Msm RNAP α-subunit C-terminal domain (αCTD) with DNA, and we provide evidence that the αCTD may play a role in Mtb transcription regulation. Our results reveal the structure of an Actinobacteria-unique insert of the RNAP β' subunit. Finally, our analysis reveals the disposition of the N-terminal segment of Msm σA, which may comprise an intrinsically disordered protein domain unique to mycobacteria. The clade-specific features of the mycobacteria RNAP provide clues to the profound instability of mycobacteria RPo compared with E. coli.

  3. ERK-dependent phosphorylation of the transcription initiation factor TIF-IA is required for RNA polymerase I transcription and cell growth

    DEFF Research Database (Denmark)

    Zhao, Jian; Yuan, Xuejun; Frödin, Morten

    2003-01-01

    -specific transcription initiation factor TIF-IA. Activation of TIF-IA and ribosomal gene transcription is sensitive to PD98059, indicating that TIF-IA is targeted by MAPK in vivo. Phosphopeptide mapping and mutational analysis reveals two serine residues (S633 and S649) that are phosphorylated by ERK and RSK kinases....... Replacement of S649 by alanine inactivates TIF-IA, inhibits pre-rRNA synthesis, and retards cell growth. The results provide a link between growth factor signaling, ribosome production, and cell growth, and may have a major impact on the mechanism of cell transformation....

  4. Promoter binding, initiation, and elongation by bacteriophage T7 RNA polymerase. A single-molecule view of the transcription cycle.

    Science.gov (United States)

    Skinner, Gary M; Baumann, Christoph G; Quinn, Diana M; Molloy, Justin E; Hoggett, James G

    2004-01-30

    A single-molecule transcription assay has been developed that allows, for the first time, the direct observation of promoter binding, initiation, and elongation by a single RNA polymerase (RNAP) molecule in real-time. To promote DNA binding and transcription initiation, a DNA molecule tethered between two optically trapped beads was held near a third immobile surface bead sparsely coated with RNAP. By driving the optical trap holding the upstream bead with a triangular oscillation while measuring the position of both trapped beads, we observed the onset of promoter binding, promoter escape (productive initiation), and processive elongation by individual RNAP molecules. After DNA template release, transcription re-initiation on the same DNA template is possible; thus, multiple enzymatic turnovers by an individual RNAP molecule can be observed. Using bacteriophage T7 RNAP, a commonly used RNAP paradigm, we observed the association and dissociation (k(off)= 2.9 s(-1)) of T7 RNAP and promoter DNA, the transition to the elongation mode (k(for) = 0.36 s(-1)), and the processive synthesis (k(pol) = 43 nt s(-1)) and release of a gene-length RNA transcript ( approximately 1200 nt). The transition from initiation to elongation is much longer than the mean lifetime of the binary T7 RNAP-promoter DNA complex (k(off) > k(for)), identifying a rate-limiting step between promoter DNA binding and promoter escape.

  5. Tuning of Recombinant Protein Expression in Escherichia coli by Manipulating Transcription, Translation Initiation Rates, and Incorporation of Noncanonical Amino Acids.

    Science.gov (United States)

    Schlesinger, Orr; Chemla, Yonatan; Heltberg, Mathias; Ozer, Eden; Marshall, Ryan; Noireaux, Vincent; Jensen, Mogens Høgh; Alfonta, Lital

    2017-06-16

    Protein synthesis in cells has been thoroughly investigated and characterized over the past 60 years. However, some fundamental issues remain unresolved, including the reasons for genetic code redundancy and codon bias. In this study, we changed the kinetics of the Eschrichia coli transcription and translation processes by mutating the promoter and ribosome binding domains and by using genetic code expansion. The results expose a counterintuitive phenomenon, whereby an increase in the initiation rates of transcription and translation lead to a decrease in protein expression. This effect can be rescued by introducing slow translating codons into the beginning of the gene, by shortening gene length or by reducing initiation rates. On the basis of the results, we developed a biophysical model, which suggests that the density of co-transcriptional-translation plays a role in bacterial protein synthesis. These findings indicate how cells use codon bias to tune translation speed and protein synthesis.

  6. Structural and functional aspects of winged-helix domains at the core of transcription initiation complexes.

    Science.gov (United States)

    Teichmann, Martin; Dumay-Odelot, Hélène; Fribourg, Sébastien

    2012-01-01

    The winged helix (WH) domain is found in core components of transcription systems in eukaryotes and prokaryotes. It represents a sub-class of the helix-turn-helix motif. The WH domain participates in establishing protein-DNA and protein-protein-interactions. Here, we discuss possible explanations for the enrichment of this motif in transcription systems.

  7. TIP48/Reptin and H2A.Z requirement for initiating chromatin remodeling in estrogen-activated transcription.

    Directory of Open Access Journals (Sweden)

    Mathieu Dalvai

    2013-04-01

    Full Text Available Histone variants, including histone H2A.Z, are incorporated into specific genomic sites and participate in transcription regulation. The role of H2A.Z at these sites remains poorly characterized. Our study investigates changes in the chromatin environment at the Cyclin D1 gene (CCND1 during transcriptional initiation in response to estradiol in estrogen receptor positive mammary tumour cells. We show that H2A.Z is present at the transcription start-site and downstream enhancer sequences of CCND1 when the gene is poorly transcribed. Stimulation of CCND1 expression required release of H2A.Z concomitantly from both these DNA elements. The AAA+ family members TIP48/reptin and the histone variant H2A.Z are required to remodel the chromatin environment at CCND1 as a prerequisite for binding of the estrogen receptor (ERα in the presence of hormone. TIP48 promotes acetylation and exchange of H2A.Z, which triggers a dissociation of the CCND1 3' enhancer from the promoter, thereby releasing a repressive intragenic loop. This release then enables the estrogen receptor to bind to the CCND1 promoter. Our findings provide new insight into the priming of chromatin required for transcription factor access to their target sequence. Dynamic release of gene loops could be a rapid means to remodel chromatin and to stimulate transcription in response to hormones.

  8. High-density transcriptional initiation signals underline genomic islands in bacteria.

    Directory of Open Access Journals (Sweden)

    Qianli Huang

    Full Text Available Genomic islands (GIs, frequently associated with the pathogenicity of bacteria and having a substantial influence on bacterial evolution, are groups of "alien" elements which probably undergo special temporal-spatial regulation in the host genome. Are there particular hallmark transcriptional signals for these "exotic" regions? We here explore the potential transcriptional signals that underline the GIs beyond the conventional views on basic sequence composition, such as codon usage and GC property bias. It showed that there is a significant enrichment of the transcription start positions (TSPs in the GI regions compared to the whole genome of Salmonella enterica and Escherichia coli. There was up to a four-fold increase for the 70% GIs, implying high-density TSPs profile can potentially differentiate the GI regions. Based on this feature, we developed a new sliding window method GIST, Genomic-island Identification by Signals of Transcription, to identify these regions. Subsequently, we compared the known GI-associated features of the GIs detected by GIST and by the existing method Islandviewer to those of the whole genome. Our method demonstrates high sensitivity in detecting GIs harboring genes with biased GI-like function, preferred subcellular localization, skewed GC property, shorter gene length and biased "non-optimal" codon usage. The special transcriptional signals discovered here may contribute to the coordinate expression regulation of foreign genes. Finally, by using GIST, we detected many interesting GIs in the 2011 German E. coli O104:H4 outbreak strain TY-2482, including the microcin H47 system and gene cluster ycgXEFZ-ymgABC that activates the production of biofilm matrix. The aforesaid findings highlight the power of GIST to predict GIs with distinct intrinsic features to the genome. The heterogeneity of cumulative TSPs profiles may not only be a better identity for "alien" regions, but also provide hints to the special

  9. Alternate capping mechanisms for transcription of spring viremia of carp virus: evidence for independent mRNA initiation.

    Science.gov (United States)

    Gupta, K C; Roy, P

    1980-01-01

    Two alternate mechanisms of mRNA capping for spring viremia of carp virus have been observed. Under normal reaction conditions, a ppG residue of the capping GTP is transferred to a pA moiety of the 5' termini of mRNA transcripts. However, in reaction conditions where GppNHp is used instead of GTP, an alternate capping mechanism occurs whereby a pG residue of the capping GTP is transferred to a ppA moiety of the transcripts. The first mechanism is identical to that described previously for vesicular stomatitis virus (G. Abraham, D. P. Rhodes, and A. K. Banerjee, Nature [London] 255:37-40, 1975; A. K. Banerjee, S. A. Moyer, and D. P. Rhodes, Virology 61:547-558, 1974), and thus appears to be a conserved function during the evolution of rhabdoviruses. The alternate mechanism of capping indicates not only that capping can take place by two procedures, but also that the substrate termini have di- or triphosphate 5' ends, indicating that they are probably independently initiated. An analog of ATP, AppNHp, has been found to completely inhibit the initiation of transcription by spring viremia of carp virus, suggesting that a cleavage between the beta and gamma phosphates of ATP is essential for the initiation of transcription. However, in the presence of GppNHp, uncapped (ppAp and pppAp), capped (GpppAp), and capped methylated (m7GpppAmpAp and GpppAmpAp) transcripts are detected. Size analyses of oligodeoxythymidylic acid-cellulose-bound transcripts resolved by formamide gel electrophoresis demonstrated that full-size mRNA transcripts are synthesized as well as larger RNA species. The presence of GppNHp and S-adenosylhomocysteine in reaction mixtures did not have any effect on the type of unmethylated transcription products. Our results favor a transcription model postulated previously (D. H. L. Bishop, in H. Fraenkel-Conrat and R. R. Wagner, ed., Comprehensive Virology, vol. 10, Plenum Press, New York, 1977; D. H. L. Bishop and A. Flamand, in D. C. Burke and W. C. Russell

  10. TFIIH with inactive XPD helicase functions in transcription initiation but is defective in DNA repair

    NARCIS (Netherlands)

    G.S. Winkler (Sebastiaan); U. Fiedler; W. Vermeulen (Wim); F. Coin (Frédéric); R.D. Wood (Richard); H.T.M. Timmers (Marc); G. Weeda (Geert); J.H.J. Hoeijmakers (Jan); S.J. Araú jo; J-M. Egly (Jean-Marc)

    2000-01-01

    textabstractTFIIH is a multisubunit protein complex involved in RNA polymerase II transcription and nucleotide excision repair, which removes a wide variety of DNA lesions including UV-induced photoproducts. Mutations in the DNA-dependent ATPase/helicase subunits of TFIIH, XPB and

  11. The Calmodulin-Binding Transcription Activator CAMTA1 Is Required for Long-Term Memory Formation in Mice

    Science.gov (United States)

    Bas-Orth, Carlos; Tan, Yan-Wei; Oliveira, Ana M. M.; Bengtson, C. Peter; Bading, Hilmar

    2016-01-01

    The formation of long-term memory requires signaling from the synapse to the nucleus to mediate neuronal activity-dependent gene transcription. Synapse-to-nucleus communication is initiated by influx of calcium ions through synaptic NMDA receptors and/or L-type voltage-gated calcium channels and involves the activation of transcription factors by…

  12. An Ancient Transcription Factor Initiates the Burst of piRNA Production During Early Meiosis in Mouse Testes

    Science.gov (United States)

    Li, Xin Zhiguo; Roy, Christian K.; Dong, Xianjun; Bolcun-Filas, Ewelina; Wang, Jie; Han, Bo W.; Xu, Jia; Moore, Melissa J.; Schimenti, John C.; Weng, Zhiping; Zamore, Phillip D.

    2013-01-01

    SUMMARY Animal germ cells produce PIWI-interacting RNAs (piRNAs), small silencing RNAs that suppress transposons and enable gamete maturation. Mammalian transposon-silencing piRNAs accumulate early in spermatogenesis, whereas pachytene piRNAs are produced later during post-natal spermatogenesis and account for >95% of all piRNAs in the adult mouse testis. Mutants defective for pachytene piRNA pathway proteins fail to produce mature sperm, but neither the piRNA precursor transcripts nor the trigger for pachytene piRNA production is known. Here, we show that the transcription factor A-MYB initiates pachytene piRNA production. A-MYB drives transcription of both pachytene piRNA precursor RNAs and the mRNAs for core piRNA biogenesis factors, including MIWI, the protein through which pachytene piRNAs function. A-MYB regulation of piRNA pathway proteins and piRNA genes creates a coherent feed-forward loop that ensures the robust accumulation of pachytene piRNAs. This regulatory circuit, which can be detected in rooster testes, likely predates the divergence of birds and mammals. PMID:23523368

  13. Molecular cloning of the human gene for von Willebrand factor and identification of the transcription initiation site

    International Nuclear Information System (INIS)

    Collins, C.J.; Underdahl, J.P.; Levene, R.B.; Ravera, C.P.; Morin, M.J.; Dombalagian, M.J.; Ricca, G.; Livingston, D.M.; Lynch, D.C.

    1987-01-01

    A series of overlapping cosmid genomic clones have been isolated that contain the entire coding unit of the human gene for van Willebrand factor (vWf), a major component of the hemostatic system. The cloned segments span ≅ 175 kilobases of human DNA sequence, and hybridization analysis suggest that the vWf coding unit is ≅150 kilobases in length. Within one of these clones, the vWF transcription initiation site has been mapped and a portion of the vWf promoter region has been sequenced, revealing a typical TATA box, a downstream CCAAT box, and a perfect downstream repeat of the 8 base pairs containing the transcription start site. Sequencing of a segment of another genomic clone has revealed the vWF translation termination codon. Where tested, comparative restriction analysis of cloned and chromosomal DNA segments strongly suggests that no major alterations occurred during cloning and that there is only one complete copy of the vWf gene in the human haploid genome. Similar analyses of DNA from vWf-producing endothelial cells and nonexpressing leukocytes suggest that vWf gene expression is not accompanied by gross genomic rearrangements. In addition, there is significant homology of C-terminal coding sequences among the vWf genes of several vertebrate species

  14. Cell Biological Mechanisms of Activity-Dependent Synapse to Nucleus Translocation of CRTC1 in Neurons

    Directory of Open Access Journals (Sweden)

    Toh Hean eCh'ng

    2015-09-01

    Full Text Available Previous studies have revealed a critical role for CREB-regulated transcriptional coactivator (CRTC1 in regulating neuronal gene expression during learning and memory. CRTC1 localizes to synapses but undergoes activity-dependent nuclear translocation to regulate the transcription of CREB target genes. Here we investigate the long-distance retrograde transport of CRTC1 in hippocampal neurons. We show that local elevations in calcium, triggered by activation of synaptic glutamate receptors and L-type voltage-gated calcium channels, initiate active, dynein-mediated retrograde transport of CRTC1 along microtubules. We identify a nuclear localization signal within CRTC1, and characterize three conserved serine residues whose dephosphorylation is required for nuclear import. Domain analysis reveals that the amino-terminal third of CRTC1 contains all of the signals required for regulated nucleocytoplasmic trafficking. We fuse this region to Dendra2 to generate a reporter construct and perform live-cell imaging coupled with local uncaging of glutamate and photoconversion to characterize the dynamics of stimulus-induced retrograde transport and nuclear accumulation.

  15. A linguistic representation of the regulation of transcription initiation. I. An ordered array of complex symbols with distinctive features.

    Science.gov (United States)

    Collado-Vides, J

    1993-01-01

    The inadequacy of context-free grammars in the description of regulatory information contained in DNA gave the formal justification for a linguistic approach to the study of gene regulation. Based on that result, we have initiated a linguistic formalization of the regulatory arrays of 107 sigma 70 E. coli promoters. The complete sequences of promoter (Pr), operator (Op) and activator binding sites (I) have previously been identified as the smallest elements, or categories, for a combinatorial analysis of the range of transcription initiation of sigma 70 promoters. These categories are conceptually equivalent to phonemes of natural language. Several features associated with these categories are required in a complete description of regulatory arrays of promoters. We have to select the best way to describe the properties that are pertinent for the description of such regulatory regions. In this paper we define distinctive features of regulatory regions based on the following criteria: identification of subclasses of substitutable elements, simplicity, selection of the most directly related information, and distinction of one array among the whole set of promoters. Alternative ways to represent distances in between regulatory sites are discussed, permitting, together with a principle of precedence, the identification of an ordered set of complex symbols as a unique representation for a promoter and its associated regulatory sites. In the accompanying paper additional distinctive features of promoters and regulatory sites are identified.

  16. Identification of EhTIF-IA: The putative E. histolytica orthologue of the human ribosomal RNA transcription initiation factor-IA.

    Science.gov (United States)

    Srivastava, Ankita; Bhattacharya, Alok; Bhattacharya, Sudha; Jhingan, Gagan Deep

    2016-03-01

    Initiation of rDNA transcription requires the assembly of a specific multi-protein complex at the rDNA promoter containing the RNA Pol I with auxiliary factors. One of these factors is known as Rrn3P in yeast and Transcription Initiation Factor IA (TIF-IA) in mammals. Rrn3p/TIF-IA serves as a bridge between RNA Pol I and the pre-initiation complex at the promoter. It is phosphorylated at multiple sites and is involved in regulation of rDNA transcription in a growth-dependent manner. In the early branching parasitic protist Entamoeba histolytica, the rRNA genes are present exclusively on circular extra chromosomal plasmids. The protein factors involved in regulation of rDNA transcription in E. histolytica are not known. We have identified the E. histolytica equivalent of TIF-1A (EhTIF-IA) by homology search within the database and was further cloned and expressed. Immuno-localization studies showed that EhTIF-IA co-localized partially with fibrillarin in the peripherally localized nucleolus. EhTIF-IA was shown to interact with the RNA Pol I-specific subunit RPA12 both in vivo and in vitro. Mass spectroscopy data identified RNA Pol I-specific subunits and other nucleolar proteins to be the interacting partners of EhTIF-IA. Our study demonstrates for the first time a conserved putative RNA Pol I transcription factor TIF-IA in E. histolytica.

  17. In Vitro Whole Genome DNA Binding Analysis of the Bacterial Replication Initiator and Transcription Factor DnaA.

    Directory of Open Access Journals (Sweden)

    Janet L Smith

    2015-05-01

    Full Text Available DnaA, the replication initiation protein in bacteria, is an AAA+ ATPase that binds and hydrolyzes ATP and exists in a heterogeneous population of ATP-DnaA and ADP-DnaA. DnaA binds cooperatively to the origin of replication and several other chromosomal regions, and functions as a transcription factor at some of these regions. We determined the binding properties of Bacillus subtilis DnaA to genomic DNA in vitro at single nucleotide resolution using in vitro DNA affinity purification and deep sequencing (IDAP-Seq. We used these data to identify 269 binding regions, refine the consensus sequence of the DnaA binding site, and compare the relative affinity of binding regions for ATP-DnaA and ADP-DnaA. Most sites had a slightly higher affinity for ATP-DnaA than ADP-DnaA, but a few had a strong preference for binding ATP-DnaA. Of the 269 sites, only the eight strongest binding ones have been observed to bind DnaA in vivo, suggesting that other cellular factors or the amount of available DnaA in vivo restricts DnaA binding to these additional sites. Conversely, we found several chromosomal regions that were bound by DnaA in vivo but not in vitro, and that the nucleoid-associated protein Rok was required for binding in vivo. Our in vitro characterization of the inherent ability of DnaA to bind the genome at single nucleotide resolution provides a backdrop for interpreting data on in vivo binding and regulation of DnaA, and is an approach that should be adaptable to many other DNA binding proteins.

  18. TAF4 nucleates a core subcomplex of TFIID and mediates activated transcription from a TATA-less promoter

    OpenAIRE

    Wright, Kevin J.; Marr, Michael T.; Tjian, Robert

    2006-01-01

    Activator-dependent recruitment of TFIID initiates formation of the transcriptional preinitiation complex. TFIID binds core promoter DNA elements and directs the assembly of other general transcription factors, leading to binding of RNA polymerase II and activation of RNA synthesis. How TATA box-binding protein (TBP) and the TBP-associated factors (TAFs) are assembled into a functional TFIID complex with promoter recognition and coactivator activities in vivo remains unknown. Here, we use RNA...

  19. HFR1 Sequesters PIF1 to Govern the Transcriptional Network Underlying Light-Initiated Seed Germination in Arabidopsis[C][W][OPEN

    Science.gov (United States)

    Shi, Hui; Zhong, Shangwei; Mo, Xiaorong; Liu, Na; Nezames, Cynthia D.; Deng, Xing Wang

    2013-01-01

    Seed germination is the first step for seed plants to initiate a new life cycle. Light plays a predominant role in promoting seed germination, where the initial phase is mediated by photoreceptor phytochrome B (phyB). Previous studies showed that PHYTOCHROME-INTERACTING FACTOR1 (PIF1) represses seed germination downstream of phyB. Here, we identify a positive regulator of phyB-dependent seed germination, LONG HYPOCOTYL IN FAR-RED1 (HFR1). HFR1 blocks PIF1 transcriptional activity by forming a heterodimer with PIF1 that prevents PIF1 from binding to DNA. Our whole-genomic analysis shows that HFR1 and PIF1 oppositely mediate the light-regulated transcriptome in imbibed seeds. Through the HFR1–PIF1 module, light regulates expression of numerous genes involved in cell wall loosening, cell division, and hormone pathways to initiate seed germination. The functionally antagonistic HFR1–PIF1 pair constructs a fail-safe mechanism for fine-tuning seed germination during low-level illumination, ensuring a rapid response to favorable environmental changes. This study identifies the HFR1–PIF1 pair as a central module directing the whole genomic transcriptional network to rapidly initiate light-induced seed germination. PMID:24179122

  20. Transcriptional Response in Mouse Thyroid Tissue after 211At Administration: Effects of Absorbed Dose, Initial Dose-Rate and Time after Administration.

    Directory of Open Access Journals (Sweden)

    Nils Rudqvist

    Full Text Available 211At-labeled radiopharmaceuticals are potentially useful for tumor therapy. However, a limitation has been the preferential accumulation of released 211At in the thyroid gland, which is a critical organ for such therapy. The aim of this study was to determine the effect of absorbed dose, dose-rate, and time after 211At exposure on genome-wide transcriptional expression in mouse thyroid gland.BALB/c mice were i.v. injected with 1.7, 7.5 or 100 kBq 211At. Animals injected with 1.7 kBq were killed after 1, 6, or 168 h with mean thyroid absorbed doses of 0.023, 0.32, and 1.8 Gy, respectively. Animals injected with 7.5 and 100 kBq were killed after 6 and 1 h, respectively; mean thyroid absorbed dose was 1.4 Gy. Total RNA was extracted from pooled thyroids and the Illumina RNA microarray platform was used to determine mRNA levels. Differentially expressed transcripts and enriched GO terms were determined with adjusted p-value 1.5, and p-value <0.05, respectively.In total, 1232 differentially expressed transcripts were detected after 211At administration, demonstrating a profound effect on gene regulation. The number of regulated transcripts increased with higher initial dose-rate/absorbed dose at 1 or 6 h. However, the number of regulated transcripts decreased with mean absorbed dose/time after 1.7 kBq 211At administration. Furthermore, similar regulation profiles were seen for groups administered 1.7 kBq. Interestingly, few previously proposed radiation responsive genes were detected in the present study. Regulation of immunological processes were prevalent at 1, 6, and 168 h after 1.7 kBq administration (0.023, 0.32, 1.8 Gy.

  1. A downstream CpG island controls transcript initiation and elongation and the methylation state of the imprinted Airn macro ncRNA promoter.

    Directory of Open Access Journals (Sweden)

    Martha V Koerner

    Full Text Available A CpG island (CGI lies at the 5' end of the Airn macro non-protein-coding (nc RNA that represses the flanking Igf2r promoter in cis on paternally inherited chromosomes. In addition to being modified on maternally inherited chromosomes by a DNA methylation imprint, the Airn CGI shows two unusual organization features: its position immediately downstream of the Airn promoter and transcription start site and a series of tandem direct repeats (TDRs occupying its second half. The physical separation of the Airn promoter from the CGI provides a model to investigate if the CGI plays distinct transcriptional and epigenetic roles. We used homologous recombination to generate embryonic stem cells carrying deletions at the endogenous locus of the entire CGI or just the TDRs. The deleted Airn alleles were analyzed by using an ES cell imprinting model that recapitulates the onset of Igf2r imprinted expression in embryonic development or by using knock-out mice. The results show that the CGI is required for efficient Airn initiation and to maintain the unmethylated state of the Airn promoter, which are both necessary for Igf2r repression on the paternal chromosome. The TDRs occupying the second half of the CGI play a minor role in Airn transcriptional elongation or processivity, but are essential for methylation on the maternal Airn promoter that is necessary for Igf2r to be expressed from this chromosome. Together the data indicate the existence of a class of regulatory CGIs in the mammalian genome that act downstream of the promoter and transcription start.

  2. [Participation of the piRNA pathway in recruiting a component of RNA polymerase I transcription initiation complex to germline cell nucleoli].

    Science.gov (United States)

    Fefelova, E A; Stolyarenko, A D; Yakushev, E Y; Gvozdev, V A; Klenov, M S

    2017-01-01

    Proteins of the Piwi family and short Piwi-interacting RNAs (piRNAs) ensure the protection of the genome from transposable elements. We have previously shown that nuclear Piwi protein tends to concentrate in the nucleoli of the cells of Drosophila melanogaster ovaries. It could be hypothesized that the function of Piwi in the nucleolus is associated with the repression of R1 and R2 retrotransposons inserted into the rDNA cluster. Here, we show that Piwi participates in recruiting Udd protein to nucleoli. Udd is a component of the conserved Selectivity Factor I-like (SL1-like) complex, which is required for transcription initiation by RNA polymerase I. We found that Udd localization depends on Piwi in germline cells, but not in somatic cells of the ovaries. In contrast, knockdowns of the SL1-like components (Udd or TAF1b) do not disrupt Piwi localization. We also observed that the absence of Udd or TAF1b in germline cells, as well as the impairment of Piwi nuclear localization lead to the accumulation of late stage egg chambers in the ovaries, which could be explained by reduced rRNA transcription. These results allow us to propose for the first time a role for Piwi in the nucleolus that is not directly associated with transposable element repression.

  3. Improved detection limit in rapid detection of human enterovirus 71 and coxsackievirus A16 by a novel reverse transcription-isothermal multiple-self-matching-initiated amplification assay.

    Science.gov (United States)

    Ding, Xiong; Nie, Kai; Shi, Lei; Zhang, Yong; Guan, Li; Zhang, Dan; Qi, Shunxiang; Ma, Xuejun

    2014-06-01

    Rapid detection of human enterovirus 71 (EV71) and coxsackievirus A16 (CVA16) is important in the early phase of hand-foot-and-mouth disease (HFMD). In this study, we developed and evaluated a novel reverse transcription-isothermal multiple-self-matching-initiated amplification (RT-IMSA) assay for the rapid detection of EV71 and CVA16 by use of reverse transcriptase, together with a strand displacement DNA polymerase. Real-time RT-IMSA assays using a turbidimeter and visual RT-IMSA assays to detect EV71 and CVA16 were established and completed in 1 h, and the reported corresponding real-time reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assays targeting the same regions of the VP1 gene were adopted as parallel tests. Through testing VP1 RNAs transcribed in vitro, the real-time RT-IMSA assays exhibited better linearity of quantification, with R(2) values of 0.952 (for EV71) and 0.967 (for CVA16), than the real-time RT-LAMP assays, which had R(2) values of 0.803 (for EV71) and 0.904 (for CVA16). Additionally, the detection limits of the real-time RT-IMSA assays (approximately 937 for EV71 and 67 for CVA16 copies/reaction) were higher than those of real-time RT-LAMP assays (approximately 3,266 for EV71 and 430 for CVA16 copies/reaction), and similar results were observed in the visual RT-IMSA assays. The new approaches also possess high specificities for the corresponding targets, with no cross-reactivity observed. In clinical assessment, compared to commercial reverse transcription-quantitative PCR (qRT-PCR) kits, the diagnostic sensitivities of the real-time RT-IMSA assays (96.4% for EV71 and 94.6% for CVA16) were higher than those of the real-time RT-LAMP assays (91.1% for EV71 and 90.8% for CVA16). The visual RT-IMSA assays also exhibited the same results. In conclusion, this proof-of-concept study suggests that the novel RT-IMSA assay is superior to the RT-LAMP assay in terms of detection limit and has the potential to rapidly detect EV71

  4. Activity-dependent modulation of neural circuit synaptic connectivity

    Directory of Open Access Journals (Sweden)

    Charles R Tessier

    2009-07-01

    Full Text Available In many nervous systems, the establishment of neural circuits is known to proceed via a two-stage process; 1 early, activity-independent wiring to produce a rough map characterized by excessive synaptic connections, and 2 subsequent, use-dependent pruning to eliminate inappropriate connections and reinforce maintained synapses. In invertebrates, however, evidence of the activity-dependent phase of synaptic refinement has been elusive, and the dogma has long been that invertebrate circuits are “hard-wired” in a purely activity-independent manner. This conclusion has been challenged recently through the use of new transgenic tools employed in the powerful Drosophila system, which have allowed unprecedented temporal control and single neuron imaging resolution. These recent studies reveal that activity-dependent mechanisms are indeed required to refine circuit maps in Drosophila during precise, restricted windows of late-phase development. Such mechanisms of circuit refinement may be key to understanding a number of human neurological diseases, including developmental disorders such as Fragile X syndrome (FXS and autism, which are hypothesized to result from defects in synaptic connectivity and activity-dependent circuit function. This review focuses on our current understanding of activity-dependent synaptic connectivity in Drosophila, primarily through analyzing the role of the fragile X mental retardation protein (FMRP in the Drosophila FXS disease model. The particular emphasis of this review is on the expanding array of new genetically-encoded tools that are allowing cellular events and molecular players to be dissected with ever greater precision and detail.

  5. Spectrometric study of the folding process of i-motif-forming DNA sequences upstream of the c-kit transcription initiation site

    International Nuclear Information System (INIS)

    Bucek, Pavel; Gargallo, Raimundo; Kudrev, Andrei

    2010-01-01

    The c-kit oncogene shows a cytosine-rich DNA region upstream of the transcription initiation site which forms an i-motif structure at slightly acidic pH values (Bucek et al. ). In the present study, the pH-induced formation of i-motif - forming sequences 5'-CCC CTC CCT CGC GCC CGC CCG-3' (ckitC1, native), 5'-CCC TTC CCT TGT GCC CGC CCG-3' (ckitC2) and 5'-CCCTT CCC TTTTT CCC T CCC T-3' (ckitC3) was studied by spectroscopic techniques, such as UV molecular absorption and circular dichroism (CD), in tandem with two multivariate data analysis methods, the hard modelling-based matrix method and the soft modelling-based MCR-ALS approach. Use of the hard chemical modelling enabled us to propose the equilibrium model, which describes spectral changes as functions of solution acidity. Additionally, the intrinsic protonation constant, K in , and the cooperativity parameters, ω c , and ω a , were calculated from the fitting procedure of the coupled CD and molecular absorption spectra. In the case of ckitC2 and ckitC3, the hard model correctly reproduced the spectral variations observed experimentally. The results indicated that folding was accompanied by a cooperative process, i.e. the enhancement of protonated structure stability upon protonation. In contrast, unfolding was accompanied by an anticooperative process. Finally, folding of the native sequence, ckitC1, seemed to follow a more complex mechanism.

  6. Control of neuropeptide expression by parallel activity-dependent pathways in caenorhabditis elegans

    DEFF Research Database (Denmark)

    Rojo Romanos, Teresa; Petersen, Jakob Gramstrup; Pocock, Roger

    2017-01-01

    Monitoring of neuronal activity within circuits facilitates integrated responses and rapid changes in behavior. We have identified a system in Caenorhabditis elegans where neuropeptide expression is dependent on the ability of the BAG neurons to sense carbon dioxide. In C. Elegans, CO 2 sensing...... is predominantly coordinated by the BAG-expressed receptor-type guanylate cyclase GCY-9. GCY-9 binding to CO 2 causes accumulation of cyclic GMP and opening of the cGMP-gated TAX-2/TAX-4 cation channels; provoking an integrated downstream cascade that enables C. Elegans to avoid high CO 2. Here we show that c...... that expression of flp-19::GFP is controlled in parallel to GCY-9 by the activity-dependent transcription factor CREB (CRH-1) and the cAMP-dependent protein kinase (KIN-2) signaling pathway. We therefore show that two parallel pathways regulate neuropeptide gene expression in the BAG sensory neurons: the ability...

  7. A novel embryological theory of autism causation involving endogenous biochemicals capable of initiating cellular gene transcription: a possible link between twelve autism risk factors and the autism 'epidemic'.

    Science.gov (United States)

    King, Chiara R

    2011-05-01

    Human alpha-fetoprotein is a pregnancy-associated protein with an undetermined physiological role. As human alpha-fetoprotein binds retinoids and inhibits estrogen-dependent cancer cell proliferation, and because retinoic acid (a retinol metabolite) and estradiol (an estrogen) can both initiate cellular gene transcription, it is hypothesized here that alpha-fetoprotein functions during critical gestational periods to prevent retinoic acid and maternal estradiol from inappropriately stimulating gene expression in developing brain regions which are sensitive to these chemicals. Prenatal/maternal factors linked to increased autism risk include valproic acid, thalidomide, alcohol, rubella, cytomegalovirus, depression, schizophrenia, obsessive-compulsive disorder, autoimmune disease, stress, allergic reaction, and hypothyroidism. It will be shown how each of these risk factors may initiate expression of genes which are sensitive to retinoic acid and/or estradiol - whether by direct promotion or by reducing production of alpha-fetoprotein. It is thus hypothesized here that autism is not a genetic disorder, but is rather an epigenetic disruption in brain development caused by gestational exposure to chemicals and/or conditions which either inhibit alpha-fetoprotein production or directly promote retinoic acid-sensitive or estradiol-sensitive gene expression. This causation model leads to potential chemical explanations for autistic brain morphology, the distinct symptomatology of Asperger's syndrome, and the differences between high-functioning and low-functioning autism with regard to mental retardation, physical malformation, and sex ratio. It will be discussed how folic acid may cause autism under the retinoic acid/estradiol model, and the history of prenatal folic acid supplementation will be shown to coincide with the history of what is popularly known as the autism epidemic. It is thus hypothesized here that prenatal folic acid supplementation has contributed to the

  8. The Role of CREB, SRF, and MEF2 in Activity-Dependent Neuronal Plasticity in the Visual Cortex.

    Science.gov (United States)

    Pulimood, Nisha S; Rodrigues, Wandilson Dos Santos; Atkinson, Devon A; Mooney, Sandra M; Medina, Alexandre E

    2017-07-12

    The transcription factors CREB (cAMP response element binding factor), SRF (serum response factor), and MEF2 (myocyte enhancer factor 2) play critical roles in the mechanisms underlying neuronal plasticity. However, the role of the activation of these transcription factors in the different components of plasticity in vivo is not well known. In this study, we tested the role of CREB, SRF, and MEF2 in ocular dominance plasticity (ODP), a paradigm of activity-dependent neuronal plasticity in the visual cortex. These three proteins bind to the synaptic activity response element (SARE), an enhancer sequence found upstream of many plasticity-related genes (Kawashima et al., 2009; Rodríguez-Tornos et al., 2013), and can act cooperatively to express Arc , a gene required for ODP (McCurry et al., 2010). We used viral-mediated gene transfer to block the transcription function of CREB, SRF, and MEF2 in the visual cortex, and measured visually evoked potentials in awake male and female mice before and after a 7 d monocular deprivation, which allowed us to examine both the depression component (Dc-ODP) and potentiation component (Pc-ODP) of plasticity independently. We found that CREB, SRF, and MEF2 are all required for ODP, but have differential effects on Dc-ODP and Pc-ODP. CREB is necessary for both Dc-ODP and Pc-ODP, whereas SRF and MEF2 are only needed for Dc-ODP. This finding supports previous reports implicating SRF and MEF2 in long-term depression (required for Dc-ODP), and CREB in long-term potentiation (required for Pc-ODP). SIGNIFICANCE STATEMENT Activity-dependent neuronal plasticity is the cellular basis for learning and memory, and it is crucial for the refinement of neuronal circuits during development. Identifying the mechanisms of activity-dependent neuronal plasticity is crucial to finding therapeutic interventions in the myriad of disorders where it is disrupted, such as Fragile X syndrome, Rett syndrome, epilepsy, major depressive disorder, and autism

  9. From retinal waves to activity-dependent retinogeniculate map development.

    Science.gov (United States)

    Markowitz, Jeffrey; Cao, Yongqiang; Grossberg, Stephen

    2012-01-01

    A neural model is described of how spontaneous retinal waves are formed in infant mammals, and how these waves organize activity-dependent development of a topographic map in the lateral geniculate nucleus, with connections from each eye segregated into separate anatomical layers. The model simulates the spontaneous behavior of starburst amacrine cells and retinal ganglion cells during the production of retinal waves during the first few weeks of mammalian postnatal development. It proposes how excitatory and inhibitory mechanisms within individual cells, such as Ca(2+)-activated K(+) channels, and cAMP currents and signaling cascades, can modulate the spatiotemporal dynamics of waves, notably by controlling the after-hyperpolarization currents of starburst amacrine cells. Given the critical role of the geniculate map in the development of visual cortex, these results provide a foundation for analyzing the temporal dynamics whereby the visual cortex itself develops.

  10. From retinal waves to activity-dependent retinogeniculate map development.

    Directory of Open Access Journals (Sweden)

    Jeffrey Markowitz

    Full Text Available A neural model is described of how spontaneous retinal waves are formed in infant mammals, and how these waves organize activity-dependent development of a topographic map in the lateral geniculate nucleus, with connections from each eye segregated into separate anatomical layers. The model simulates the spontaneous behavior of starburst amacrine cells and retinal ganglion cells during the production of retinal waves during the first few weeks of mammalian postnatal development. It proposes how excitatory and inhibitory mechanisms within individual cells, such as Ca(2+-activated K(+ channels, and cAMP currents and signaling cascades, can modulate the spatiotemporal dynamics of waves, notably by controlling the after-hyperpolarization currents of starburst amacrine cells. Given the critical role of the geniculate map in the development of visual cortex, these results provide a foundation for analyzing the temporal dynamics whereby the visual cortex itself develops.

  11. Polycistronic transcription of fused cassettes and identification of translation initiation signals in an unusual gene cassette array from Pseudomonas aeruginosa [version 3; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Érica L. Fonseca

    2015-11-01

    Full Text Available The gene cassettes found in class 1 integrons are generally promoterless units composed by an open reading frame (ORF, a short 5’ untranslated region (UTR and a 3’ recombination site (attC. Fused gene cassettes are generated by partial or total loss of the attC from the first cassette in an array, creating, in some cases, a fusion with the ORF from the next cassette. These structures are rare and little is known about their mechanisms of mobilization and expression. The aim of this study was to evaluate the dynamic of mobilization and transcription of the gcu14-blaGES-1/aacA4 gene cassette array, which harbours a fused gene cassette represented by blaGES-1/aacA4. The cassette array was analyzed by Northern blot and real-time reverse transcription-polymerase chain reaction (RT-PCR in order to assess the transcription mechanism of blaGES-1/aacA4 fused cassette. Also, inverse polymerase chain reactions (PCR were performed to detect the free circular forms of gcu14, blaGES-1 and aacA4. The Northern blot and real time RT-PCR revealed a polycistronic transcription, in which the fused cassette blaGES-1/aacA4 is transcribed as a unique gene, while gcu14 (with a canonical attC recombination site has a monocistronic transcription. The gcu14 cassette, closer to the weak configuration of cassette promoter (PcW, had a higher transcription level than blaGES-1/aacA4, indicating that the cassette position affects the transcript amounts. The presence of ORF-11 at attI1, immediately preceding gcu14, and of a Shine-Dalgarno sequence upstream blaGES-1/aacA4 composes a scenario for the occurrence of array translation. Inverse PCR generated amplicons corresponding to gcu14, gcu14-aacA4 and gcu14-blaGES-1/aacA4 free circular forms, but not to blaGES-1 and aacA4 alone, indicating that the GES-1 truncated attC is not substrate of integrase activity and that these genes are mobilized together as a unique cassette. This study was original in showing the transcription

  12. C-terminal BRE overexpression in 11q23-rearranged and t(8;16) acute myeloid leukemia is caused by intragenic transcription initiation.

    Science.gov (United States)

    Marneth, A E; Prange, K H M; Al Hinai, A S A; Bergevoet, S M; Tesi, N; Janssen-Megens, E M; Kim, B; Sharifi, N; Yaspo, M L; Kuster, J; Sanders, M A; Stoetman, E C G; Knijnenburg, J; Arentsen-Peters, T C J M; Zwaan, C M; Stunnenberg, H G; van den Heuvel-Eibrink, M M; Haferlach, T; Fornerod, M; Jansen, J H; Valk, P J M; van der Reijden, B A; Martens, J H A

    2018-03-01

    Overexpression of the BRE (brain and reproductive organ-expressed) gene defines a distinct pediatric and adult acute myeloid leukemia (AML) subgroup. Here we identify a promoter enriched for active chromatin marks in BRE intron 4 causing strong biallelic expression of a previously unknown C-terminal BRE transcript. This transcript starts with BRE intron 4 sequences spliced to exon 5 and downstream sequences, and if translated might code for an N terminally truncated BRE protein. Remarkably, the new BRE transcript was highly expressed in over 50% of 11q23/KMT2A (lysine methyl transferase 2A)-rearranged and t(8;16)/KAT6A-CREBBP cases, while it was virtually absent from other AML subsets and normal tissues. In gene reporter assays, the leukemia-specific fusion protein KMT2A-MLLT3 transactivated the intragenic BRE promoter. Further epigenome analyses revealed 97 additional intragenic promoter marks frequently bound by KMT2A in AML with C-terminal BRE expression. The corresponding genes may be part of a context-dependent KMT2A-MLLT3-driven oncogenic program, because they were higher expressed in this AML subtype compared with other groups. C-terminal BRE might be an important contributor to this program because in a case with relapsed AML, we observed an ins(11;2) fusing CHORDC1 to BRE at the region where intragenic transcription starts in KMT2A-rearranged and KAT6A-CREBBP AML.

  13. Cocaine promotes both initiation and elongation phase of HIV-1 transcription by activating NF-κB and MSK1 and inducing selective epigenetic modifications at HIV-1 LTR

    International Nuclear Information System (INIS)

    Sahu, Geetaram; Farley, Kalamo; El-Hage, Nazira; Aiamkitsumrit, Benjamas; Fassnacht, Ryan; Kashanchi, Fatah; Ochem, Alex; Simon, Gary L.; Karn, Jonathan; Hauser, Kurt F.; Tyagi, Mudit

    2015-01-01

    Cocaine accelerates human immunodeficiency virus (HIV-1) replication by altering specific cell-signaling and epigenetic pathways. We have elucidated the underlying molecular mechanisms through which cocaine exerts its effect in myeloid cells, a major target of HIV-1 in central nervous system (CNS). We demonstrate that cocaine treatment promotes HIV-1 gene expression by activating both nuclear factor-kappa B (NF-ĸB) and mitogen- and stress-activated kinase 1 (MSK1). MSK1 subsequently catalyzes the phosphorylation of histone H3 at serine 10, and p65 subunit of NF-ĸB at 276th serine residue. These modifications enhance the interaction of NF-ĸB with P300 and promote the recruitment of the positive transcription elongation factor b (P-TEFb) to the HIV-1 LTR, supporting the development of an open/relaxed chromatin configuration, and facilitating the initiation and elongation phases of HIV-1 transcription. Results are also confirmed in primary monocyte derived macrophages (MDM). Overall, our study provides detailed insights into cocaine-driven HIV-1 transcription and replication. - Highlights: • Cocaine induces the initiation phase of HIV transcription by activating NF-ĸB. • Cocaine induced NF-ĸB phosphorylation promotes its interaction with P300. • Cocaine enhances the elongation phase of HIV transcription by stimulating MSK1. • Cocaine activated MSK1 catalyzes the phosphorylation of histone H3 at its Ser10. • Cocaine induced H3S10 phosphorylation facilitates the recruitment of P-TEFb at LTR

  14. Cocaine promotes both initiation and elongation phase of HIV-1 transcription by activating NF-κB and MSK1 and inducing selective epigenetic modifications at HIV-1 LTR

    Energy Technology Data Exchange (ETDEWEB)

    Sahu, Geetaram; Farley, Kalamo [Division of Infectious Diseases, Department of Medicine, George Washington University, Washington, DC (United States); El-Hage, Nazira [Virginia Commonwealth University, Richmond, VA (United States); Aiamkitsumrit, Benjamas; Fassnacht, Ryan [Division of Infectious Diseases, Department of Medicine, George Washington University, Washington, DC (United States); Kashanchi, Fatah [George Mason University, Manassas, VA (United States); Ochem, Alex [ICGEB, Wernher and Beit Building, Anzio Road, Observatory, 7925 Cape Town (South Africa); Simon, Gary L. [Division of Infectious Diseases, Department of Medicine, George Washington University, Washington, DC (United States); Karn, Jonathan [Case Western Reserve University, Cleveland, OH (United States); Hauser, Kurt F. [Virginia Commonwealth University, Richmond, VA (United States); Tyagi, Mudit, E-mail: tmudit@email.gwu.edu [Division of Infectious Diseases, Department of Medicine, George Washington University, Washington, DC (United States); Department of Microbiology, Immunology and Tropical Medicine, George Washington University, Washington, DC 20037 (United States)

    2015-09-15

    Cocaine accelerates human immunodeficiency virus (HIV-1) replication by altering specific cell-signaling and epigenetic pathways. We have elucidated the underlying molecular mechanisms through which cocaine exerts its effect in myeloid cells, a major target of HIV-1 in central nervous system (CNS). We demonstrate that cocaine treatment promotes HIV-1 gene expression by activating both nuclear factor-kappa B (NF-ĸB) and mitogen- and stress-activated kinase 1 (MSK1). MSK1 subsequently catalyzes the phosphorylation of histone H3 at serine 10, and p65 subunit of NF-ĸB at 276th serine residue. These modifications enhance the interaction of NF-ĸB with P300 and promote the recruitment of the positive transcription elongation factor b (P-TEFb) to the HIV-1 LTR, supporting the development of an open/relaxed chromatin configuration, and facilitating the initiation and elongation phases of HIV-1 transcription. Results are also confirmed in primary monocyte derived macrophages (MDM). Overall, our study provides detailed insights into cocaine-driven HIV-1 transcription and replication. - Highlights: • Cocaine induces the initiation phase of HIV transcription by activating NF-ĸB. • Cocaine induced NF-ĸB phosphorylation promotes its interaction with P300. • Cocaine enhances the elongation phase of HIV transcription by stimulating MSK1. • Cocaine activated MSK1 catalyzes the phosphorylation of histone H3 at its Ser10. • Cocaine induced H3S10 phosphorylation facilitates the recruitment of P-TEFb at LTR.

  15. Shaping inhibition: activity dependent structural plasticity of GABAergic synapses

    Directory of Open Access Journals (Sweden)

    Carmen E Flores

    2014-10-01

    Full Text Available Inhibitory transmission through the neurotransmitter Ɣ-aminobutyric acid (GABA shapes network activity in the mammalian cerebral cortex by filtering synaptic incoming information and dictating the activity of principal cells. The incredibly diverse population of cortical neurons that use GABA as neurotransmitter shows an equally diverse range of mechanisms that regulate changes in the strength of GABAergic synaptic transmission and allow them to dynamically follow and command the activity of neuronal ensembles. Similarly to glutamatergic synaptic transmission, activity-dependent functional changes in inhibitory neurotransmission are accompanied by alterations in GABAergic synapse structure that range from morphological reorganization of postsynaptic density to de novo formation and elimination of inhibitory contacts. Here we review several aspects of structural plasticity of inhibitory synapses, including its induction by different forms of neuronal activity, behavioral and sensory experience and the molecular mechanisms and signaling pathways involved. We discuss the functional consequences of GABAergic synapse structural plasticity for information processing and memory formation in view of the heterogenous nature of the structural plasticity phenomena affecting inhibitory synapses impinging on somatic and dendritic compartments of cortical and hippocampal neurons.

  16. Cocaine promotes both initiation and elongation phase of HIV-1 transcription by activating NF-κB and MSK1 and inducing selective epigenetic modifications at HIV-1 LTR

    Science.gov (United States)

    Sahu, Geetaram; Farley, Kalamo; El-Hage, Nazira; Aiamkitsumrit, Benjamas; Fassnacht, Ryan; Kashanchi, Fatah; Ochem, Alex; Simon, Gary L.; Karn, Jonathan; Hauser, Kurt F.; Tyagi, Mudit

    2015-01-01

    Cocaine accelerates human immunodeficiency virus (HIV-1) replication by altering specific cell-signaling and epigenetic pathways. We have elucidated the underlying molecular mechanisms through which cocaine exerts its effect in myeloid cells, a major target of HIV-1 in central nervous system (CNS). We demonstrate that cocaine treatment promotes HIV-1 gene expression by activating both nuclear factor-kappa B (NF-κB) and mitogen- and stress-activated kinase 1 (MSK1). MSK1 subsequently catalyzes the phosphorylation of histone H3 at serine 10, and p65 subunit of NF-κB at 276th serine residue. These modifications enhance the interaction of NF-κB with P300 and promote the recruitment of the positive transcription elongation factor b (P-TEFb) to the HIV-1 LTR, supporting the development of an open/relaxed chromatin configuration, and facilitating the initiation and elongation phases of HIV-1 transcription. Results are also confirmed in primary monocyte derived macrophages (MDM). Overall, our study provides detailed insights into cocaine-driven HIV-1 transcription and replication. PMID:25980739

  17. Basal transcription machinery

    Indian Academy of Sciences (India)

    2007-03-29

    Mar 29, 2007 ... The holoenzyme of prokaryotic RNA polymerase consists of the core enzyme, made of two , , ' and subunits, which lacks promoter selectivity and a sigma () subunit which enables the core enzyme to initiate transcription in a promoter dependent fashion. A stress sigma factor s, in prokaryotes ...

  18. Activity-dependent astrocyte swelling is mediated by pH-regulating mechanisms.

    Science.gov (United States)

    Larsen, Brian Roland; MacAulay, Nanna

    2017-10-01

    During neuronal activity in the mammalian brain, the K + released into the synaptic space is initially buffered by the astrocytic compartment. In parallel, the extracellular space (ECS) shrinks, presumably due to astrocytic cell swelling. With the Na + /K + /2Cl - cotransporter and the Kir4.1/AQP4 complex not required for the astrocytic cell swelling in the hippocampus, the molecular mechanisms underlying the activity-dependent ECS shrinkage have remained unresolved. To identify these molecular mechanisms, we employed ion-sensitive microelectrodes to measure changes in ECS, [K + ] o and [H + ] o /pH o during electrical stimulation of rat hippocampal slices. Transporters and receptors responding directly to the K + and glutamate released into the extracellular space (the K + /Cl - cotransporter, KCC, glutamate transporters and G protein-coupled receptors) did not modulate the extracellular space dynamics. The HCO3--transporting mechanism, which in astrocytes mainly constitutes the electrogenic Na + / HCO3- cotransporter 1 (NBCe1), is activated by the K + -mediated depolarization of the astrocytic membrane. Inhibition of this transporter reduced the ECS shrinkage by ∼25% without affecting the K + transients, pointing to NBCe1 as a key contributor to the stimulus-induced astrocytic cell swelling. Inhibition of the monocarboxylate cotransporters (MCT), like-wise, reduced the ECS shrinkage by ∼25% without compromising the K + transients. Isosmotic reduction of extracellular Cl - revealed a requirement for this ion in parts of the ECS shrinkage. Taken together, the stimulus-evoked astrocytic cell swelling does not appear to occur as a direct effect of the K + clearance, as earlier proposed, but partly via the pH-regulating transport mechanisms activated by the K + -induced astrocytic depolarization and the activity-dependent metabolism. © 2017 Wiley Periodicals, Inc.

  19. Neurotrophin release by neurotrophins: Implications for activity-dependent neuronal plasticity

    OpenAIRE

    Canossa, Marco; Griesbeck, Oliver; Berninger, Benedikt; Campana, Gabriele; Kolbeck, Roland; Thoenen, Hans

    1997-01-01

    Neurotrophins, secreted in an activity-dependent manner, are thought to be involved in the activity-dependent refinement of synaptic connections. Here we demonstrate that in hippocampal neurons and the rat pheochromocytoma cell line PC12 application of exogenous neurotrophins induces secretion of neurotrophins, an effect that is mediated by the activation of tyrosine kinase neurotrophin receptors (Trks). Like activity-dependent secretion of neurotrophins, neurotrophin-induced neurotrophin sec...

  20. HIV-1 transcripts use IRES-initiation under conditions where Cap-dependent translation is restricted by poliovirus 2A protease.

    Directory of Open Access Journals (Sweden)

    Raquel Amorim

    Full Text Available The 30 different species of mRNAs synthesized during the HIV-1 replication cycle are all capped and polyadenilated. Internal ribosome entry sites have been recognized in the 5' untranslated region of some mRNA species of HIV-1, which would contribute to an alternative mechanism of initiation of mRNA translation. However, the Cap-dependent translation is assumed to be the main mechanism driving the initiation of HIV-1 protein synthesis. In this work, we describe a cell system in which lower to higher levels of transient expression of the poliovirus 2A protease strongly inhibited cellular Cap-dependent translation with no toxic effect to the cells during a 72-hour time frame. In this system, the synthesis of HIV-1 proteins was inhibited in a temporal dose-dependent way. Higher levels of 2A protease expression severely inhibited HIV-1 protein synthesis during the first 24 hours of infection consequently inhibiting viral production and infectivity. Intermediate to lower levels of 2A Protease expression caused the inhibition of viral protein synthesis only during the first 48 hours of viral replication. After this period both protein synthesis and viral release were recovered to the control levels. However, the infectivity of viral progeny was still partially inhibited. These results indicate that two mechanisms of mRNA translation initiation contribute to the synthesis of HIV-1 proteins; during the first 24-48 hours of viral replication HIV-1 protein synthesis is strongly dependent on Cap-initiation, while at later time points IRES-driven translation initiation is sufficient to produce high amounts of viral particles.

  1. Deciphering Transcriptional Regulation

    DEFF Research Database (Denmark)

    Valen, Eivind

    The myriad of cells in the human body are all made from the same blueprint: the human genome. At the heart of this diversity lies the concept of gene regulation, the process in which it is decided which genes are used where and when. Genes do not function as on/off buttons, but more like a volume...... mostly near the start of the gene known as the promoter. This region contains patterns scattered in the DNA that the TFs can recognize and bind to. Such binding can prompt the assembly of the pre-initiation complex which ultimately leads to transcription of the gene. In order to achieve the regulation...... on what characterizes a hippocampus promoter. Pairing CAGE with TF binding site prediction we identi¿ed a likely key regulator of hippocampus. Finally, we developed a method for CAGE exploration. While the DeepCAGE library characterized a full 1.4 million transcription initiation events it did not capture...

  2. Production of the 2400 kb Duchenne muscular dystrophy (DMD) gene transcript; transcription time and cotranscriptional splicing

    Energy Technology Data Exchange (ETDEWEB)

    Tennyson, C.N.; Worton, R.G. [Univ. of Toronto and the Hospital for Sick Children, Ontario (Canada)

    1994-09-01

    The largest known gene in any organism is the human DMD gene which has 79 exons that span 2400 kb. The extreme nature of the DMD gene raises questions concerning the time required for transcription and whether splicing begins before transcription is complete. DMD gene transcription is induced as cultured human myoblasts differentiate to form multinucleated myotubes, providing a system for studying the kinetics of transcription and splicing. Using quantitative RT-PCR, transcript accumulation was monitored from four different regions within the gene following induction of expression. By comparing the accumulation of transcripts from the 5{prime} and 3{prime} ends of the gene we have shown that approximately 12 hours are required to transcribe 1770 kb of the gene, extrapolating to a time of 16 hours for the transcription unit expressed in muscle. Comparison of accumulation profiles for spliced and total transcript demonstrated that transcripts are spliced at the 5{prime} end before transcription is complete, providing strong evidence for cotranscriptional splicing of DMD gene transcripts. Finally, the rate of transcript accumulation was reduced at the 3{prime} end of the gene relative to the 5{prime} end, perhaps due to premature termination of transcription complexes as they traverse this enormous transcription unit. The lag between transcription initiation and the appearance of complete transcripts could be important in limiting transcript production in dividing cells and to the timing of mRNA appearance in differentiating muscle.

  3. Activation-Dependent Rapid Postsynaptic Clustering of Glycine Receptors in Mature Spinal Cord Neurons

    Science.gov (United States)

    Eto, Kei; Murakoshi, Hideji; Watanabe, Miho; Hirata, Hiromi; Moorhouse, Andrew J.; Ishibashi, Hitoshi

    2017-01-01

    Abstract Inhibitory synapses are established during development but continue to be generated and modulated in strength in the mature nervous system. In the spinal cord and brainstem, presynaptically released inhibitory neurotransmitter dominantly switches from GABA to glycine during normal development in vivo. While presynaptic mechanisms of the shift of inhibitory neurotransmission are well investigated, the contribution of postsynaptic neurotransmitter receptors to this shift is not fully elucidated. Synaptic clustering of glycine receptors (GlyRs) is regulated by activation-dependent depolarization in early development. However, GlyR activation induces hyperpolarization after the first postnatal week, and little is known whether and how presynaptically released glycine regulates postsynaptic receptors in a depolarization-independent manner in mature developmental stage. Here we developed spinal cord neuronal culture of rodents using chronic strychnine application to investigate whether initial activation of GlyRs in mature stage could change postsynaptic localization of GlyRs. Immunocytochemical analyses demonstrate that chronic blockade of GlyR activation until mature developmental stage resulted in smaller clusters of postsynaptic GlyRs that could be enlarged upon receptor activation for 1 h in the mature stage. Furthermore, live cell-imaging techniques show that GlyR activation decreases its lateral diffusion at synapses, and this phenomenon is dependent on PKC, but neither Ca2+ nor CaMKII activity. These results suggest that the GlyR activation can regulate receptor diffusion and cluster size at inhibitory synapses in mature stage, providing not only new insights into the postsynaptic mechanism of shifting inhibitory neurotransmission but also the inhibitory synaptic plasticity in mature nervous system. PMID:28197549

  4. Neuromodulation of activity-dependent synaptic enhancement at crayfish neuromuscular junction.

    Science.gov (United States)

    Qian, S M; Delaney, K R

    1997-10-17

    decay of ADSE by 5-HT was not accompanied by significant changes in the initial amplitude of activity-dependent components of enhancement 1.5 s after the train. Measurements of presynaptic [Ca2+] indicated that the time course of Ca2+ removal from the presynaptic terminals after trains was not altered by 5-HT. Changes in presynaptic action potential shape, resting membrane potential or postsynaptic impedance after trains cannot account for slower recovery of ADSE. Axonal injection of EDTA slows the removal of residual Ca2+ and the decay of synaptic augmentation after trains of action potentials (K.R. Delaney, D.W. Tank, A quantitative measure of the dependence of short-term synaptic enhancement on presynaptic residual calcium, J. Neurosci. 14 (1994) 5885-5902), but has little or no effect on the 5-HT-induced persistence of ADSE. This also suggests that the time course of ADSE in the presence of 5-HT is not determined primarily by residual Ca2+ removal kinetics. The slowing of ADSE recovery after trains by 5-HT reverses with washing in 5-HT-free saline along with the 5-HT-mediated enhancement of release.

  5. Temporal phases of activity-dependent plasticity and memory are mediated by compartmentalized routing of MAPK signaling in aplysia sensory neurons.

    Science.gov (United States)

    Shobe, Justin L; Zhao, Yali; Stough, Shara; Ye, Xiaojing; Hsuan, Vickie; Martin, Kelsey C; Carew, Thomas J

    2009-01-15

    An activity-dependent form of intermediate memory (AD-ITM) for sensitization is induced in Aplysia by a single tail shock that gives rise to plastic changes (AD-ITF) in tail sensory neurons (SNs) via the interaction of action potential firing in the SN coupled with the release of serotonin in the CNS. Activity-dependent long-term facilitation (AD-LTF, lasting >24hr) requires protein synthesis dependent persistent mitogen-activated protein kinase (MAPK) activation and translocation to the SN nucleus. We now show that the induction of the earlier temporal phase (AD-ITM and AD-ITF), which is translation and transcription independent, requires the activation of a compartmentally distinct novel signaling cascade that links second messengers, MAPK and PKC into a unified pathway within tail SNs. Since both AD-ITM and AD-LTM require MAPK activity, these collective findings suggest that presynaptic SNs route the flow of molecular information to distinct subcellular compartments during the induction of activity-dependent long-lasting memories.

  6. pH-Modulated Watson-Crick duplex-quadruplex equilibria of guanine-rich and cytosine-rich DNA sequences 140 base pairs upstream of the c-kit transcription initiation site.

    Science.gov (United States)

    Bucek, Pavel; Jaumot, Joaquim; Aviñó, Anna; Eritja, Ramon; Gargallo, Raimundo

    2009-11-23

    Guanine-rich regions of DNA are sequences capable of forming G-quadruplex structures. The formation of a G-quadruplex structure in a region 140 base pairs (bp) upstream of the c-kit transcription initiation site was recently proposed (Fernando et al., Biochemistry, 2006, 45, 7854). In the present study, the acid-base equilibria and the thermally induced unfolding of the structures formed by a guanine-rich region and by its complementary cytosine-rich strand in c-kit were studied by means of circular dichroism and molecular absorption spectroscopies. In addition, competition between the Watson-Crick duplex and the isolated structures was studied as a function of pH value and temperature. Multivariate data analysis methods based on both hard and soft modeling were used to allow accurate quantification of the various acid-base species present in the mixtures. Results showed that the G-quadruplex and i-motif coexist with the Watson-Crick duplex over the pH range from 3.0 to 6.5, approximately, under the experimental conditions tested in this study. At pH 7.0, the duplex is practically the only species present.

  7. Interaction between FMDV Lpro and transcription factor ADNP is required for viral replication

    Science.gov (United States)

    The foot-and-mouth disease virus (FMDV) leader protease (Lpro) inhibits host translation and transcription affecting the expression of several factors involved in innate immunity. In this study, we have identified the host transcription factor ADNP (activity dependent neuroprotective protein) as an ...

  8. The down-stream effects of mannan-induced lectin complement pathway activation depend quantitatively on alternative pathway amplification

    DEFF Research Database (Denmark)

    Harboe, Morten; Garred, Peter; Karlstrøm, Ellen

    2009-01-01

    Complement activation plays an important role in human pathophysiology. The effect of classical pathway activation is largely dependent on alternative pathway (AP) amplification, whereas the role of AP for the down-stream effect of mannan-induced lectin pathway (LP) activation is poorly understood...... that AP amplification is quantitatively responsible for the final effect of initial specific LP activation. TCC generation on the solid phase was distinctly but less inhibited by anti-fD. C2 bypass of the LP pathway could be demonstrated, and AP amplification was also essential during C2 bypass in LP...... as shown by complete inhibition of TCC generation in C2-deficient serum by anti-fD and anti-properdin antibodies. In conclusion, the down-stream effect of LP activation depends strongly on AP amplification in normal human serum and in the C2 bypass pathway....

  9. Novel Functions for TAF7, a Regulator of TAF1-independent Transcription

    OpenAIRE

    Devaiah, Ballachanda N.; Lu, Hanxin; Gegonne, Anne; Sercan, Zeynep; Zhang, Hongen; Clifford, Robert J.; Lee, Maxwell P.; Singer, Dinah S.

    2010-01-01

    The transcription factor TFIID components TAF7 and TAF1 regulate eukaryotic transcription initiation. TAF7 regulates transcription initiation of TAF1-dependent genes by binding to the acetyltransferase (AT) domain of TAF1 and inhibiting the enzymatic activity that is essential for transcription. TAF7 is released from the TAF1-TFIID complex upon completion of preinitiation complex assembly, allowing transcription to initiate. However, not all transcription is TAF1-dependent, and the role of TA...

  10. Activity-Dependent NPAS4 Expression and the Regulation of Gene Programs Underlying Plasticity in the Central Nervous System

    Directory of Open Access Journals (Sweden)

    José Fernando Maya-Vetencourt

    2013-01-01

    Full Text Available The capability of the brain to change functionally in response to sensory experience is most active during early stages of development but it decreases later in life when major alterations of neuronal network structures no longer take place in response to experience. This view has been recently challenged by experimental strategies based on the enhancement of environmental stimulation levels, genetic manipulations, and pharmacological treatments, which all have demonstrated that the adult brain retains a degree of plasticity that allows for a rewiring of neuronal circuitries over the entire life course. A hot spot in the field of neuronal plasticity centres on gene programs that underlie plastic phenomena in adulthood. Here, I discuss the role of the recently discovered neuronal-specific and activity-dependent transcription factor NPAS4 as a critical mediator of plasticity in the nervous system. A better understanding of how modifications in the connectivity of neuronal networks occur may shed light on the treatment of pathological conditions such as brain damage or disease in adult life, some of which were once considered untreatable.

  11. Structural insights into transcription complexes

    NARCIS (Netherlands)

    Berger, I.; Blanco, A.G.; Boelens, R.; Cavarelli, J.; Coll, M.; Folkers, G.E.; Nie, Y.; Pogenberg, V.; Schultz, P.; Wilmanns, M.; Moras, D.; Poterszman, A.

    2011-01-01

    Control of transcription allows the regulation of cell activity in response to external stimuli and research in the field has greatly benefited from efforts in structural biology. In this review, based on specific examples from the European SPINE2-COMPLEXES initiative, we illustrate the impact of

  12. Activity-dependent neurorehabilitation beyond physical trainings: "mental exercise" through mirror neuron activation

    OpenAIRE

    Yuan, Ti-Fei; Chen, Wei; Shan, Chunlei; Rocha, Nuno; Arias-Carrión, Oscar; Paes, Flávia; de Sa, Alberto Souza; Machado, Sergio

    2015-01-01

    The activity dependent brain repair mechanism has been widely adopted in many types of neurorehabilitation. The activity leads to target specific and non-specific beneficial effects in different brain regions, such as the releasing of neurotrophic factors, modulation of the cytokines and generation of new neurons in adult hood. However physical exercise program clinically are limited to some of the patients with preserved motor functions; while many patients suffered from paralysis cannot mak...

  13. The transcriptional landscape

    DEFF Research Database (Denmark)

    Nielsen, Henrik

    2011-01-01

    The application of new and less biased methods to study the transcriptional output from genomes, such as tiling arrays and deep sequencing, has revealed that most of the genome is transcribed and that there is substantial overlap of transcripts derived from the two strands of DNA. In protein coding...... regions, the map of transcripts is very complex due to small transcripts from the flanking ends of the transcription unit, the use of multiple start and stop sites for the main transcript, production of multiple functional RNA molecules from the same primary transcript, and RNA molecules made...... by independent transcription from within the unit. In genomic regions separating those that encode proteins or highly abundant RNA molecules with known function, transcripts are generally of low abundance and short-lived. In most of these cases, it is unclear to what extent a function is related to transcription...

  14. Nucleic Acid Analogue Induced Transcription of Double Stranded DNA

    DEFF Research Database (Denmark)

    1998-01-01

    RNA is transcribed from a double stranded DNA template by forming a complex by hybridizing to the template at a desired transcription initiation site one or more oligonucleic acid analogues of the PNA type capable of forming a transcription initiation site with the DNA and exposing the complex...... to the action of a DNA dependant RNA polymerase in the presence of nucleoside triphosphates. Equal length transcripts may be obtained by placing a block to transcription downstream from the initiation site or by cutting the template at such a selected location. The initiation site is formed by displacement...... of one strand of the DNA locally by the PNA hybridization....

  15. Early survival factor deprivation in the olfactory epithelium enhances activity-dependent survival

    Directory of Open Access Journals (Sweden)

    Adrien eFrançois

    2013-12-01

    Full Text Available The neuronal olfactory epithelium undergoes permanent renewal because of environmental aggression. This renewal is partly regulated by factors modulating the level of neuronal apoptosis. Among them, we had previously characterized endothelin as neuroprotective. In this study, we explored the effect of cell survival factor deprivation in the olfactory epithelium by intranasal delivery of endothelin receptors antagonists to rat pups. This treatment induced an overall increase of apoptosis in the olfactory epithelium. The responses to odorants recorded by electroolfactogram were decreased in treated animal, a result consistent with a loss of olfactory sensory neurons (OSNs. However, the treated animal performed better in an olfactory orientation test based on maternal odor compared to non-treated littermates. This improved performance could be due to activity-dependent neuronal survival of OSNs in the context of increased apoptosis level. In order to demonstrate it, we odorized pups with octanal, a known ligand for the rI7 olfactory receptor (Olr226. We quantified the number of OSN expressing rI7 by RT-qPCR and whole mount in situ hybridization. While this number was reduced by the survival factor removal treatment, this reduction was abolished by the presence of its ligand. This improved survival was optimal for low concentration of odorant and was specific for rI7-expressing OSNs. Meanwhile, the number of rI7-expressing OSNs was not affected by the odorization in non-treated littermates; showing that the activity-dependant survival of OSNs did not affect the OSN population during the 10 days of odorization in control conditions. Overall, our study shows that when apoptosis is promoted in the olfactory mucosa, the activity-dependent neuronal plasticity allows faster tuning of the olfactory sensory neuron population towards detection of environmental odorants.

  16. Novel marker for the onset of frontotemporal dementia: early increase in activity-dependent neuroprotective protein (ADNP in the face of Tau mutation.

    Directory of Open Access Journals (Sweden)

    Yulie Schirer

    Full Text Available Tauopathy, a major pathology in Alzheimer's disease, is also found in ~50% of frontotemporal dementias (FTDs. Tau transcript, a product of a single gene, undergoes alternative splicing to yield 6 protein species, each with either 3 or 4 microtubule binding repeat domains (tau 3R or 4R, associated with dynamic and stable microtubules, respectively. While the healthy human brain shows a 1/1 ratio of tau 3R/4R, this ratio may be dramatically changed in the FTD brain. We have previously discovered that activity-dependent neuroprotective protein (ADNP is essential for brain formation in the mouse, with ADNP+/- mice exhibiting tauopathy, age-driven neurodegeneration and behavioral deficits. Here, in transgenic mice overexpressing a mutated tau 4R species, in the cerebral cortex but not in the cerebellum, we showed significantly increased ADNP expression (~3-fold transcripts in the cerebral cortex of young transgenic mice (~disease onset, but not in the cerebellum, as compared to control littermates. The transgene-age-related increased ADNP expression paralleled augmented dynamic tau 3R transcript level compared to control littermates. Blocking mutated tau 4R transgene expression resulted in normalization of ADNP and tau 3R expression. ADNP was previously shown to be a member of the SWItch/Sucrose NonFermentable (SWI/SNF chromatin remodeling complex. Here, Brahma (Brm, a component of the SWI/SNF complex regulating alternative splicing, showed a similar developmental expression pattern to ADNP. Immunoprecipitations further suggested Brm-ADNP interaction coupled to ADNP - polypyrimidine tract-binding protein (PTB-associated splicing factor (PSF-binding, with PSF being a direct regulator of tau transcript splicing. It should be noted that although we have shown a correlation between levels of ADNP and tau isoform expression three months of age, we are not presenting evidence of a direct link between the two. Future research into ADNP/tau relations is

  17. Memory and pattern storage in neural networks with activity dependent synapses

    Science.gov (United States)

    Mejias, J. F.; Torres, J. J.

    2009-01-01

    We present recently obtained results on the influence of the interplay between several activity dependent synaptic mechanisms, such as short-term depression and facilitation, on the maximum memory storage capacity in an attractor neural network [1]. In contrast with the case of synaptic depression, which drastically reduces the capacity of the network to store and retrieve activity patterns [2], synaptic facilitation is able to enhance the memory capacity in different situations. In particular, we find that a convenient balance between depression and facilitation can enhance the memory capacity, reaching maximal values similar to those obtained with static synapses, that is, without activity-dependent processes. We also argue, employing simple arguments, that this level of balance is compatible with experimental data recorded from some cortical areas, where depression and facilitation may play an important role for both memory-oriented tasks and information processing. We conclude that depressing synapses with a certain level of facilitation allow to recover the good retrieval properties of networks with static synapses while maintaining the nonlinear properties of dynamic synapses, convenient for information processing and coding.

  18. Activity-Dependent IGF-1 Exocytosis is Controlled by the Ca2+-Sensor Synaptotagmin-10

    Science.gov (United States)

    Cao, Peng; Maximov, Anton; Südhof, Thomas C.

    2011-01-01

    Synaptotagmins Syt1, Syt2, Syt7, and Syt9 act as Ca2+-sensors for synaptic and neuroendocrine exocytosis, but the function of other synaptotagmins remains unknown. Here, we show that olfactory bulb neurons secrete IGF-1 by an activity-dependent pathway of exocytosis, and that Syt10 functions as the Ca2+-sensor that triggers IGF-1 exocytosis in these neurons. Deletion of Syt10 impaired activity-dependent IGF-1 secretion in olfactory bulb neurons, resulting in smaller neurons and an overall decrease in synapse numbers. Exogenous IGF-1 completely reversed the Syt10 knockout phenotype. Syt10 co-localized with IGF-1 in somatodendritic vesicles of olfactory bulb neurons, and Ca2+-binding to Syt10 caused these vesicles to undergo exocytosis, thereby secreting IGF-1. Thus, Syt10 controls a previously unrecognized pathway of Ca2+-dependent exocytosis that is spatially and temporally distinct from Ca2+-dependent synaptic vesicle exocytosis controlled by Syt1 in the same neurons, and two different synaptotagmins regulate distinct Ca2+-dependent membrane fusion reactions during exocytosis in the same neuron. PMID:21496647

  19. Seasonal variation and solar activity dependence of the quiet-time ionospheric trough

    Science.gov (United States)

    Ishida, T.; Ogawa, Y.; Kadokura, A.; Hiraki, Y.; Häggström, I.

    2014-08-01

    We have conducted a statistical analysis of the ionospheric F region trough, focusing on its seasonal variation and solar activity dependence under geomagnetically quiet and moderate conditions, using plasma parameter data obtained via Common Program 3 observations performed by the European Incoherent Scatter (EISCAT) radar between 1982 and 2011. We have confirmed that there is a major difference in frictional heating between the high- and low-latitude sides of the EISCAT field of view (FOV) at ~73°0'N-60°5'N (geomagnetic latitude) at an altitude of 325 km, which is associated with trough formation. Our statistical results show that the high-latitude and midlatitude troughs occur on the high- and low-latitude sides of the FOV, respectively. Seasonal variations indicate that dissociative recombination accompanied by frictional heating is a main cause of trough formation in sunlit regions. During summer, therefore, the occurrence rate is maintained at 80-90% in the postmidnight high-latitude region owing to frictional heating by eastward return flow. Solar activity dependence on trough formation indicates that field-aligned currents modulate the occurrence rate of the trough during the winter and equinox seasons. In addition, the trough becomes deeper via dissociative recombination caused by an increased ion temperature with F10.7, at least in the equinox and summer seasons but not in winter.

  20. Transcription regulation by the Mediator complex.

    Science.gov (United States)

    Soutourina, Julie

    2018-04-01

    Alterations in the regulation of gene expression are frequently associated with developmental diseases or cancer. Transcription activation is a key phenomenon in the regulation of gene expression. In all eukaryotes, mediator of RNA polymerase II transcription (Mediator), a large complex with modular organization, is generally required for transcription by RNA polymerase II, and it regulates various steps of this process. The main function of Mediator is to transduce signals from the transcription activators bound to enhancer regions to the transcription machinery, which is assembled at promoters as the preinitiation complex (PIC) to control transcription initiation. Recent functional studies of Mediator with the use of structural biology approaches and functional genomics have revealed new insights into Mediator activity and its regulation during transcription initiation, including how Mediator is recruited to transcription regulatory regions and how it interacts and cooperates with PIC components to assist in PIC assembly. Novel roles of Mediator in the control of gene expression have also been revealed by showing its connection to the nuclear pore and linking Mediator to the regulation of gene positioning in the nuclear space. Clear links between Mediator subunits and disease have also encouraged studies to explore targeting of this complex as a potential therapeutic approach in cancer and fungal infections.

  1. Eukaryotic translation initiator protein 1A isoform, CCS-3, enhances the transcriptional repression of p21CIP1 by proto-oncogene FBI-1 (Pokemon/ZBTB7A).

    Science.gov (United States)

    Choi, Won-Il; Kim, Youngsoo; Kim, Yuri; Yu, Mi-young; Park, Jungeun; Lee, Choong-Eun; Jeon, Bu-Nam; Koh, Dong-In; Hur, Man-Wook

    2009-01-01

    FBI-1, a member of the POK (POZ and Kruppel) family of transcription factors, plays a role in differentiation, oncogenesis, and adipogenesis. eEF1A is a eukaryotic translation elongation factor involved in several cellular processes including embryogenesis, oncogenic transformation, cell proliferation, and cytoskeletal organization. CCS-3, a potential cervical cancer suppressor, is an isoform of eEF1A. We found that eEF1A forms a complex with FBI-1 by co-immunoprecipitation, SDS-PAGE, and MALDI-TOF Mass analysis of the immunoprecipitate. GST fusion protein pull-downs showed that FBI-1 directly interacts with eEF1A and CCS-3 via the zinc finger and POZ-domain of FBI-1. FBI-1 co-localizes with either eEF1A or CCS-3 at the nuclear periplasm. CCS-3 enhances transcriptional repression of the p21CIP1 gene (hereafter referred to as p21) by FBI-1. The POZ-domain of FBI-1 interacts with the co-repressors, SMRT and BCoR. We found that CCS-3 also interacts with the co-repressors independently. The molecular interaction between the co-repressors and CCS-3 at the POZ-domain of FBI-1 appears to enhance FBI-1 mediated transcriptional repression. Our data suggest that CCS-3 may be important in cell differentiation, tumorigenesis, and oncogenesis by interacting with the proto-oncogene FBI-1 and transcriptional co-repressors. Copyright 2009 S. Karger AG, Basel.

  2. Characteristics of seasonal variation and solar activity dependence of the geomagnetic solar quiet daily variation

    Science.gov (United States)

    Shinbori, A.; Koyama, Y.; Nose, M.; Hori, T.

    2017-12-01

    Characteristics of seasonal variation and solar activity dependence of the X- and Y-components of the geomagnetic solar quiet (Sq) daily variation at Memanbetsu in mid-latitudes and Guam near the equator have been investigated using long-term geomagnetic field data with 1-h time resolution from 1957 to 2016. In this analysis, we defined the quiet day when the maximum value of the Kp index is less than 3 for that day. In this analysis, we used the monthly average of the adjusted daily F10.7 corresponding to geomagnetically quiet days. For identification of the monthly mean Sq variation in the X and Y components (Sq-X and Sq-Y), we first determined the baseline of the X and Y components from the average value from 22 to 2 h (LT: local time) for each quiet day. Next, we calculated a deviation from the baseline of the X- and Y-components of the geomagnetic field for each quiet day, and computed the monthly mean value of the deviation for each local time. As a result, Sq-X and Sq-Y shows a clear seasonal variation and solar activity dependence. The amplitude of seasonal variation increases significantly during high solar activities, and is proportional to the solar F10.7 index. The pattern of the seasonal variation is quite different between Sq-X and Sq-Y. The result of the correlation analysis between the solar F10.7 index and Sq-X and Sq-Y shows almost the linear relationship, but the slope and intercept of the linear fitted line varies as function of local time and month. This implies that the sensitivity of Sq-X and Sq-Y to the solar activity is different for different local times and seasons. The local time dependence of the offset value of Sq-Y at Guam and its seasonal variation suggest a magnetic field produced by inter-hemispheric field-aligned currents (FACs). From the sign of the offset value of Sq-Y, it is infer that the inter-hemispheric FACs flow from the summer to winter hemispheres in the dawn and dusk sectors and from the winter to summer hemispheres in

  3. Regenerative Rehabilitation: Combining Stem Cell Therapies and Activity-Dependent Stimulation.

    Science.gov (United States)

    Moritz, Chet T; Ambrosio, Fabrisia

    2017-07-01

    The number of clinical trials in regenerative medicine is burgeoning, and stem cell/tissue engineering technologies hold the possibility of becoming the standard of care for a multitude of diseases and injuries. Advances in regenerative biology reveal novel molecular and cellular targets, with potential to optimize tissue healing and functional recovery, thereby refining rehabilitation clinical practice. The purpose of this review is to (1) highlight the potential for synergy between the fields of regenerative medicine and rehabilitation, a convergence of disciplines known as regenerative rehabilitation; (2) provide translational examples of regenerative rehabilitation within the context of neuromuscular injuries and diseases; and (3) offer recommendations for ways to leverage activity dependence via combined therapy and technology, with the goal of enhancing long-term recovery. The potential clinical benefits of regenerative rehabilitation will likely become a critical aspect in the standard of care for many neurological and musculoskeletal disorders.

  4. A unified architecture of transcriptional regulatory elements

    DEFF Research Database (Denmark)

    Andersson, Robin; Sandelin, Albin Gustav; Danko, Charles G.

    2015-01-01

    Gene expression is precisely controlled in time and space through the integration of signals that act at gene promoters and gene-distal enhancers. Classically, promoters and enhancers are considered separate classes of regulatory elements, often distinguished by histone modifications. However...... and enhancers are considered a single class of functional element, with a unified architecture for transcription initiation. The context of interacting regulatory elements and the surrounding sequences determine local transcriptional output as well as the enhancer and promoter activities of individual elements....

  5. CHD chromatin remodelers and the transcription cycle

    Science.gov (United States)

    Murawska, Magdalena

    2011-01-01

    It is well established that ATP-dependent chromatin remodelers modulate DNA access of transcription factors and RNA polymerases by “opening” or “closing” chromatin structure. However, this view is far too simplistic. Recent findings have demonstrated that these enzymes not only set the stage for the transcription machinery to act but also are actively involved at every step of the transcription process. As a consequence, they affect initiation, elongation, termination and RNA processing. In this review we will use the CHD family as a paradigm to illustrate the progress that has been made in revealing these new concepts. PMID:22223048

  6. Transcriptional regulation by competing transcription factor modules.

    Directory of Open Access Journals (Sweden)

    Rutger Hermsen

    2006-12-01

    Full Text Available Gene regulatory networks lie at the heart of cellular computation. In these networks, intracellular and extracellular signals are integrated by transcription factors, which control the expression of transcription units by binding to cis-regulatory regions on the DNA. The designs of both eukaryotic and prokaryotic cis-regulatory regions are usually highly complex. They frequently consist of both repetitive and overlapping transcription factor binding sites. To unravel the design principles of these promoter architectures, we have designed in silico prokaryotic transcriptional logic gates with predefined input-output relations using an evolutionary algorithm. The resulting cis-regulatory designs are often composed of modules that consist of tandem arrays of binding sites to which the transcription factors bind cooperatively. Moreover, these modules often overlap with each other, leading to competition between them. Our analysis thus identifies a new signal integration motif that is based upon the interplay between intramodular cooperativity and intermodular competition. We show that this signal integration mechanism drastically enhances the capacity of cis-regulatory domains to integrate signals. Our results provide a possible explanation for the complexity of promoter architectures and could be used for the rational design of synthetic gene circuits.

  7. WRKY transcription factors

    Science.gov (United States)

    Bakshi, Madhunita; Oelmüller, Ralf

    2014-01-01

    WRKY transcription factors are one of the largest families of transcriptional regulators found exclusively in plants. They have diverse biological functions in plant disease resistance, abiotic stress responses, nutrient deprivation, senescence, seed and trichome development, embryogenesis, as well as additional developmental and hormone-controlled processes. WRKYs can act as transcriptional activators or repressors, in various homo- and heterodimer combinations. Here we review recent progress on the function of WRKY transcription factors in Arabidopsis and other plant species such as rice, potato, and parsley, with a special focus on abiotic, developmental, and hormone-regulated processes. PMID:24492469

  8. MPTP-meditated hippocampal dopamine deprivation modulates synaptic transmission and activity-dependent synaptic plasticity

    International Nuclear Information System (INIS)

    Zhu Guoqi; Chen Ying; Huang Yuying; Li Qinglin; Behnisch, Thomas

    2011-01-01

    Parkinson's disease (PD)-like symptoms including learning deficits are inducible by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Therefore, it is possible that MPTP may disturb hippocampal memory processing by modulation of dopamine (DA)- and activity-dependent synaptic plasticity. We demonstrate here that intraperitoneal (i.p.) MPTP injection reduces the number of tyrosine hydroxylase (TH)-positive neurons in the substantia nigra (SN) within 7 days. Subsequently, the TH expression level in SN and hippocampus and the amount of DA and its metabolite DOPAC in striatum and hippocampus decrease. DA depletion does not alter basal synaptic transmission and changes pair-pulse facilitation (PPF) of field excitatory postsynaptic potentials (fEPSPs) only at the 30 ms inter-pulse interval. In addition, the induction of long-term potentiation (LTP) is impaired whereas the duration of long-term depression (LTD) becomes prolonged. Since both LTP and LTD depend critically on activation of NMDA and DA receptors, we also tested the effect of DA depletion on NMDA receptor-mediated synaptic transmission. Seven days after MPTP injection, the NMDA receptor-mediated fEPSPs are decreased by about 23%. Blocking the NMDA receptor-mediated fEPSP does not mimic the MPTP-LTP. Only co-application of D1/D5 and NMDA receptor antagonists during tetanization resembled the time course of fEPSP potentiation as observed 7 days after i.p. MPTP injection. Together, our data demonstrate that MPTP-induced degeneration of DA neurons and the subsequent hippocampal DA depletion alter NMDA receptor-mediated synaptic transmission and activity-dependent synaptic plasticity. - Highlights: → I.p. MPTP-injection mediates death of dopaminergic neurons. → I.p. MPTP-injection depletes DA and DOPAC in striatum and hippocampus. → I.p. MPTP-injection does not alter basal synaptic transmission. → Reduction of LTP and enhancement of LTD after i.p. MPTP-injection. → Attenuation of NMDA-receptors mediated

  9. Activity-dependent control of NMDA receptor subunit composition at hippocampal mossy fibre synapses.

    Science.gov (United States)

    Carta, Mario; Srikumar, Bettadapura N; Gorlewicz, Adam; Rebola, Nelson; Mulle, Christophe

    2018-02-15

    CA3 pyramidal cells display input-specific differences in the subunit composition of synaptic NMDA receptors (NMDARs). Although at low density, GluN2B contributes significantly to NMDAR-mediated EPSCs at mossy fibre synapses. Long-term potentiation (LTP) of NMDARs triggers a modification in the subunit composition of synaptic NMDARs by insertion of GluN2B. GluN2B subunits are essential for the expression of LTP of NMDARs at mossy fibre synapses. Single neurons express NMDA receptors (NMDARs) with distinct subunit composition and biophysical properties that can be segregated in an input-specific manner. The dynamic control of the heterogeneous distribution of synaptic NMDARs is crucial to control input-dependent synaptic integration and plasticity. In hippocampal CA3 pyramidal cells from mice of both sexes, we found that mossy fibre (MF) synapses display a markedly lower proportion of GluN2B-containing NMDARs than associative/commissural synapses. The mechanism involved in such heterogeneous distribution of GluN2B subunits is not known. Here we show that long-term potentiation (LTP) of NMDARs, which is selectively expressed at MF-CA3 pyramidal cell synapses, triggers a modification in the subunit composition of synaptic NMDARs by insertion of GluN2B. This activity-dependent recruitment of GluN2B at mature MF-CA3 pyramidal cell synapses contrasts with the removal of GluN2B subunits at other glutamatergic synapses during development and in response to activity. Furthermore, although expressed at low levels, GluN2B is necessary for the expression of LTP of NMDARs at MF-CA3 pyramidal cell synapses. Altogether, we reveal a previously unknown activity-dependent regulation and function of GluN2B subunits that may contribute to the heterogeneous plasticity induction rules in CA3 pyramidal cells. © 2017 Centre Nationnal de la Recherche Scientifique. The Journal of Physiology © 2017 The Physiological Society.

  10. Cdk7 Is Required for Activity-Dependent Neuronal Gene Expression, Long-Lasting Synaptic Plasticity and Long-Term Memory

    Directory of Open Access Journals (Sweden)

    Guiqin He

    2017-11-01

    Full Text Available In the brain, de novo gene expression driven by learning-associated neuronal activities is critical for the formation of long-term memories. However, the signaling machinery mediating neuronal activity-induced gene expression, especially the rapid transcription of immediate-early genes (IEGs remains unclear. Cyclin-dependent kinases (Cdks are a family of serine/threonine kinases that have been firmly established as key regulators of transcription processes underling coordinated cell cycle entry and sequential progression in nearly all types of proliferative cells. Cdk7 is a subunit of transcriptional initiation factor II-H (TFIIH and the only known Cdk-activating kinase (CAK in metazoans. Recent studies using a novel Cdk7 specific covalent inhibitor, THZ1, revealed important roles of Cdk7 in transcription regulation in cancer cells. However, whether Cdk7 plays a role in the regulation of transcription in neurons remains unknown. In this study, we present evidence demonstrating that, in post-mitotic neurons, Cdk7 activity is positively correlated with neuronal activities in cultured primary neurons, acute hippocampal slices and in the brain. Cdk7 inhibition by THZ1 significantly suppressed mRNA levels of IEGs, selectively impaired long-lasting synaptic plasticity induced by 4 trains of high frequency stimulation (HFS and prevented the formation of long-term memories.

  11. The contribution of ketone bodies to basal and activity-dependent neuronal oxidation in vivo.

    Science.gov (United States)

    Chowdhury, Golam M I; Jiang, Lihong; Rothman, Douglas L; Behar, Kevin L

    2014-07-01

    The capacity of ketone bodies to replace glucose in support of neuronal function is unresolved. Here, we determined the contributions of glucose and ketone bodies to neocortical oxidative metabolism over a large range of brain activity in rats fasted 36 hours and infused intravenously with [2,4-(13)C₂]-D-β-hydroxybutyrate (BHB). Three animal groups and conditions were studied: awake ex vivo, pentobarbital-induced isoelectricity ex vivo, and halothane-anesthetized in vivo, the latter data reanalyzed from a recent study. Rates of neuronal acetyl-CoA oxidation from ketone bodies (V(acCoA-kbN)) and pyruvate (V(pdhN)), and the glutamate-glutamine cycle (V(cyc)) were determined by metabolic modeling of (13)C label trapped in major brain amino acid pools. V(acCoA-kbN) increased gradually with increasing activity, as compared with the steeper change in tricarboxylic acid (TCA) cycle rate (V(tcaN)), supporting a decreasing percentage of neuronal ketone oxidation: ∼100% (isoelectricity), 56% (halothane anesthesia), 36% (awake) with the BHB plasma levels achieved in our experiments (6 to 13 mM). In awake animals ketone oxidation reached saturation for blood levels >17 mM, accounting for 62% of neuronal substrate oxidation, the remainder (38%) provided by glucose. We conclude that ketone bodies present at sufficient concentration to saturate metabolism provides full support of basal (housekeeping) energy needs and up to approximately half of the activity-dependent oxidative needs of neurons.

  12. Adenosine: an activity-dependent axonal signal regulating MAP kinase and proliferation in developing Schwann cells.

    Science.gov (United States)

    Stevens, Beth; Ishibashi, Tomoko; Chen, Jiang-Fan; Fields, R Douglas

    2004-02-01

    Nonsynaptic release of ATP from electrically stimulated dorsal root gangion (DRG) axons inhibits Schwann cell (SC) proliferation and arrests SC development at the premyelinating stage, but the specific types of purinergic receptor(s) and intracellular signaling pathways involved in this form of neuron-glia communication are not known. Recent research shows that adenosine is a neuron-glial transmitter between axons and myelinating glia of the CNS. The present study investigates the possibility that adenosine might have a similar function in communicating between axons and premyelinating SCs. Using a combination of pharmacological and molecular approaches, we found that mouse SCs in culture express functional adenosine receptors and ATP receptors, a far more complex array of purinergic receptors than thought previously. Adenosine, but not ATP, activates ERK/MAPK through stimulation of cAMP-linked A2(A) adenosine receptors. Both ATP and adenosine inhibit proliferation of SCs induced by platelet-derived growth factor (PDGF), via mechanisms that are partly independent. In contrast to ATP, adenosine failed to inhibit the differentiation of SCs to the O4+ stage. This indicates that, in addition to ATP, adenosine is an activity-dependent signaling molecule between axons and premyelinating Schwann cells, but that electrical activity, acting through adenosine, has opposite effects on the differentiation of myelinating glia in the PNS and CNS.

  13. Synaptic Mechanisms of Activity-Dependent Remodeling in Visual Cortex during Monocular Deprivation

    Directory of Open Access Journals (Sweden)

    Cynthia D. Rittenhouse

    2009-01-01

    Full Text Available It has long been appreciated that in the visual cortex, particularly within a postnatal critical period for experience-dependent plasticity, the closure of one eye results in a shift in the responsiveness of cortical cells toward the experienced eye. While the functional aspects of this ocular dominance shift have been studied for many decades, their cortical substrates and synaptic mechanisms remain elusive. Nonetheless, it is becoming increasingly clear that ocular dominance plasticity is a complex phenomenon that appears to have an early and a late component. Early during monocular deprivation, deprived eye cortical synapses depress, while later during the deprivation open eye synapses potentiate. Here we review current literature on the cortical mechanisms of activity-dependent plasticity in the visual system during the critical period. These studies shed light on the role of activity in shaping neuronal structure and function in general and can lead to insights regarding how learning is acquired and maintained at the neuronal level during normal and pathological brain development.

  14. Genetically encoded proton sensors reveal activity-dependent pH changes in neurons

    Directory of Open Access Journals (Sweden)

    Joseph Valentino Raimondo

    2012-05-01

    Full Text Available The regulation of hydrogen ion concentration (pH is fundamental to cell viability, metabolism and enzymatic function. Within the nervous system, the control of pH is also involved in diverse and dynamic processes including development, synaptic transmission and the control of network excitability. As pH affects neuronal activity, and can also itself be altered by neuronal activity, the existence of tools to accurately measure hydrogen ion fluctuations is important for understanding the role pH plays under physiological and pathological conditions. Outside of their use as a marker of synaptic release, genetically encoded pH sensors have not been utilised to study hydrogen ion fluxes associated with network activity. By combining whole-cell patch clamp with simultaneous two-photon or confocal imaging, we quantified the amplitude and time course of neuronal, intracellular, acidic transients evoked by epileptiform activity in two separate in vitro models of temporal lobe epilepsy. In doing so, we demonstrate the suitability of three genetically encoded pH sensors: deGFP4, E2GFP and Cl-sensor for investigating activity-dependent pH changes at the level of single neurons.

  15. Genetically encoded proton sensors reveal activity-dependent pH changes in neurons.

    Science.gov (United States)

    Raimondo, Joseph V; Irkle, Agnese; Wefelmeyer, Winnie; Newey, Sarah E; Akerman, Colin J

    2012-01-01

    The regulation of hydrogen ion concentration (pH) is fundamental to cell viability, metabolism, and enzymatic function. Within the nervous system, the control of pH is also involved in diverse and dynamic processes including development, synaptic transmission, and the control of network excitability. As pH affects neuronal activity, and can also itself be altered by neuronal activity, the existence of tools to accurately measure hydrogen ion fluctuations is important for understanding the role pH plays under physiological and pathological conditions. Outside of their use as a marker of synaptic release, genetically encoded pH sensors have not been utilized to study hydrogen ion fluxes associated with network activity. By combining whole-cell patch clamp with simultaneous two-photon or confocal imaging, we quantified the amplitude and time course of neuronal, intracellular, acidic transients evoked by epileptiform activity in two separate in vitro models of temporal lobe epilepsy. In doing so, we demonstrate the suitability of three genetically encoded pH sensors: deGFP4, E(2)GFP, and Cl-sensor for investigating activity-dependent pH changes at the level of single neurons.

  16. Activity-Dependent Plasticity of Spike Pauses in Cerebellar Purkinje Cells

    Directory of Open Access Journals (Sweden)

    Giorgio Grasselli

    2016-03-01

    Full Text Available The plasticity of intrinsic excitability has been described in several types of neurons, but the significance of non-synaptic mechanisms in brain plasticity and learning remains elusive. Cerebellar Purkinje cells are inhibitory neurons that spontaneously fire action potentials at high frequencies and regulate activity in their target cells in the cerebellar nuclei by generating a characteristic spike burst-pause sequence upon synaptic activation. Using patch-clamp recordings from mouse Purkinje cells, we find that depolarization-triggered intrinsic plasticity enhances spike firing and shortens the duration of spike pauses. Pause plasticity is absent from mice lacking SK2-type potassium channels (SK2−/− mice and in occlusion experiments using the SK channel blocker apamin, while apamin wash-in mimics pause reduction. Our findings demonstrate that spike pauses can be regulated through an activity-dependent, exclusively non-synaptic, SK2 channel-dependent mechanism and suggest that pause plasticity—by altering the Purkinje cell output—may be crucial to cerebellar information storage and learning.

  17. The Development and Activity-Dependent Expression of Aggrecan in the Cat Visual Cortex

    Science.gov (United States)

    Sengpiel, F.; Beaver, C. J.; Crocker-Buque, A.; Kelly, G. M.; Matthews, R. T.; Mitchell, D. E.

    2013-01-01

    The Cat-301 monoclonal antibody identifies aggrecan, a chondroitin sulfate proteoglycan in the cat visual cortex and dorsal lateral geniculate nucleus (dLGN). During development, aggrecan expression increases in the dLGN with a time course that matches the decline in plasticity. Moreover, examination of tissue from selectively visually deprived cats shows that expression is activity dependent, suggesting a role for aggrecan in the termination of the sensitive period. Here, we demonstrate for the first time that the onset of aggrecan expression in area 17 also correlates with the decline in experience-dependent plasticity in visual cortex and that this expression is experience dependent. Dark rearing until 15 weeks of age dramatically reduced the density of aggrecan-positive neurons in the extragranular layers, but not in layer IV. This effect was reversible as dark-reared animals that were subsequently exposed to light showed normal numbers of Cat-301-positive cells. The reduction in aggrecan following certain early deprivation regimens is the first biochemical correlate of the functional changes to the γ-aminobutyric acidergic system that have been reported following early deprivation in cats. PMID:22368089

  18. Quantitative monitoring of activity-dependent bulk endocytosis of synaptic vesicle membrane by fluorescent dextran imaging

    Science.gov (United States)

    Clayton, Emma Louise; Cousin, Michael Alan

    2012-01-01

    Activity-dependent bulk endocytosis (ADBE) is the dominant synaptic vesicle (SV) retrieval mode in central nerve terminals during periods of intense neuronal activity. Despite this fact there are very few real time assays that report the activity of this critical SV retrieval mode. In this paper we report a simple and quantitative assay of ADBE using uptake of large flourescent dextrans as fluid phase markers. We show that almost all dextran uptake occurs in nerve terminals, using co-localisation with the fluorescent probe FM1-43. We also demonstrate that accumulated dextran cannot be unloaded by neuronal stimulation, indicating its specific loading into bulk endosomes and not SVs. Quantification of dextran uptake was achieved by using thresholding analysis to count the number of loaded nerve terminals, since monitoring the average fluorescence intensity of these nerve terminals did not accurately report the extent of ADBE. Using this analysis we showed that dextran uptake occurs very soon after stimulation and that it does not persist when stimulation terminates. Thus we have devised a simple and quantitative method to monitor ADBE in living neurones, which will be ideal for real time screening of small molecule inhibitors of this key SV retrieval mode. PMID:19766140

  19. Activity-dependent trafficking of lysosomes in dendrites and dendritic spines.

    Science.gov (United States)

    Goo, Marisa S; Sancho, Laura; Slepak, Natalia; Boassa, Daniela; Deerinck, Thomas J; Ellisman, Mark H; Bloodgood, Brenda L; Patrick, Gentry N

    2017-08-07

    In neurons, lysosomes, which degrade membrane and cytoplasmic components, are thought to primarily reside in somatic and axonal compartments, but there is little understanding of their distribution and function in dendrites. Here, we used conventional and two-photon imaging and electron microscopy to show that lysosomes traffic bidirectionally in dendrites and are present in dendritic spines. We find that lysosome inhibition alters their mobility and also decreases dendritic spine number. Furthermore, perturbing microtubule and actin cytoskeletal dynamics has an inverse relationship on the distribution and motility of lysosomes in dendrites. We also find trafficking of lysosomes is correlated with synaptic α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid-type glutamate receptors. Strikingly, lysosomes traffic to dendritic spines in an activity-dependent manner and can be recruited to individual spines in response to local activation. These data indicate the position of lysosomes is regulated by synaptic activity and thus plays an instructive role in the turnover of synaptic membrane proteins. © 2017 Goo et al.

  20. Activity-Dependent Exocytosis of Lysosomes Regulates the Structural Plasticity of Dendritic Spines.

    Science.gov (United States)

    Padamsey, Zahid; McGuinness, Lindsay; Bardo, Scott J; Reinhart, Marcia; Tong, Rudi; Hedegaard, Anne; Hart, Michael L; Emptage, Nigel J

    2017-01-04

    Lysosomes have traditionally been viewed as degradative organelles, although a growing body of evidence suggests that they can function as Ca 2+ stores. Here we examined the function of these stores in hippocampal pyramidal neurons. We found that back-propagating action potentials (bpAPs) could elicit Ca 2+ release from lysosomes in the dendrites. This Ca 2+ release triggered the fusion of lysosomes with the plasma membrane, resulting in the release of Cathepsin B. Cathepsin B increased the activity of matrix metalloproteinase 9 (MMP-9), an enzyme involved in extracellular matrix (ECM) remodelling and synaptic plasticity. Inhibition of either lysosomal Ca 2+ signaling or Cathepsin B release prevented the maintenance of dendritic spine growth induced by Hebbian activity. This impairment could be rescued by exogenous application of active MMP-9. Our findings suggest that activity-dependent exocytosis of Cathepsin B from lysosomes regulates the long-term structural plasticity of dendritic spines by triggering MMP-9 activation and ECM remodelling. Crown Copyright © 2017. Published by Elsevier Inc. All rights reserved.

  1. Long lasting protein synthesis- and activity-dependent spine shrinkage and elimination after synaptic depression.

    Directory of Open Access Journals (Sweden)

    Yazmín Ramiro-Cortés

    Full Text Available Neuronal circuits modify their response to synaptic inputs in an experience-dependent fashion. Increases in synaptic weights are accompanied by structural modifications, and activity dependent, long lasting growth of dendritic spines requires new protein synthesis. When multiple spines are potentiated within a dendritic domain, they show dynamic structural plasticity changes, indicating that spines can undergo bidirectional physical modifications. However, it is unclear whether protein synthesis dependent synaptic depression leads to long lasting structural changes. Here, we investigate the structural correlates of protein synthesis dependent long-term depression (LTD mediated by metabotropic glutamate receptors (mGluRs through two-photon imaging of dendritic spines on hippocampal pyramidal neurons. We find that induction of mGluR-LTD leads to robust and long lasting spine shrinkage and elimination that lasts for up to 24 hours. These effects depend on signaling through group I mGluRs, require protein synthesis, and activity. These data reveal a mechanism for long lasting remodeling of synaptic inputs, and offer potential insights into mental retardation.

  2. Activity-Dependent Neurorehabilitation Beyond Physical Trainings: "Mental Exercise" Through Mirror Neuron Activation.

    Science.gov (United States)

    Yuan, Ti-Fei; Chen, Wei; Shan, Chunlei; Rocha, Nuno; Arias-Carrión, Oscar; Paes, Flávia; de Sá, Alberto Souza; Machado, Sergio

    2015-01-01

    The activity dependent brain repair mechanism has been widely adopted in many types of neurorehabilitation. The activity leads to target specific and non-specific beneficial effects in different brain regions, such as the releasing of neurotrophic factors, modulation of the cytokines and generation of new neurons in adult hood. However physical exercise program clinically are limited to some of the patients with preserved motor functions; while many patients suffered from paralysis cannot make such efforts. Here the authors proposed the employment of mirror neurons system in promoting brain rehabilitation by "observation based stimulation". Mirror neuron system has been considered as an important basis for action understanding and learning by mimicking others. During the action observation, mirror neuron system mediated the direct activation of the same group of motor neurons that are responsible for the observed action. The effect is clear, direct, specific and evolutionarily conserved. Moreover, recent evidences hinted for the beneficial effects on stroke patients after mirror neuron system activation therapy. Finally some music-relevant therapies were proposed to be related with mirror neuron system.

  3. TAF(II)250: a transcription toolbox.

    Science.gov (United States)

    Wassarman, D A; Sauer, F

    2001-08-01

    Activation of RNA-polymerase-II-dependent transcription involves conversion of signals provided by gene-specific activator proteins into the synthesis of messenger RNA. This conversion requires dynamic structural changes in chromatin and assembly of general transcription factors (GTFs) and RNA polymerase II at core promoter sequence elements surrounding the transcription start site of genes. One hallmark of transcriptional activation is the interaction of DNA-bound activators with coactivators such as the TATA-box binding protein (TBP)-associated factors (TAF(II)s) within the GTF TFIID. TAF(II)250 possesses a variety of activities that are likely to contribute to the initial steps of RNA polymerase II transcription. TAF(II)250 is a scaffold for assembly of other TAF(II)s and TBP into TFIID, TAF(II)250 binds activators to recruit TFIID to particular promoters, TAF(II)250 regulates binding of TBP to DNA, TAF(II)250 binds core promoter initiator elements, TAF(II)250 binds acetylated lysine residues in core histones, and TAF(II)250 possesses protein kinase, ubiquitin-activating/conjugating and acetylase activities that modify histones and GTFs. We speculate that these activities achieve two goals--(1) they aid in positioning and stabilizing TFIID at particular promoters, and (2) they alter chromatin structure at the promoter to allow assembly of GTFs--and we propose a model for how TAF(II)250 converts activation signals into active transcription.

  4. Downstream Antisense Transcription Predicts Genomic Features That Define the Specific Chromatin Environment at Mammalian Promoters.

    Directory of Open Access Journals (Sweden)

    Christopher A Lavender

    2016-08-01

    Full Text Available Antisense transcription is a prevalent feature at mammalian promoters. Previous studies have primarily focused on antisense transcription initiating upstream of genes. Here, we characterize promoter-proximal antisense transcription downstream of gene transcription starts sites in human breast cancer cells, investigating the genomic context of downstream antisense transcription. We find extensive correlations between antisense transcription and features associated with the chromatin environment at gene promoters. Antisense transcription downstream of promoters is widespread, with antisense transcription initiation observed within 2 kb of 28% of gene transcription start sites. Antisense transcription initiates between nucleosomes regularly positioned downstream of these promoters. The nucleosomes between gene and downstream antisense transcription start sites carry histone modifications associated with active promoters, such as H3K4me3 and H3K27ac. This region is bound by chromatin remodeling and histone modifying complexes including SWI/SNF subunits and HDACs, suggesting that antisense transcription or resulting RNA transcripts contribute to the creation and maintenance of a promoter-associated chromatin environment. Downstream antisense transcription overlays additional regulatory features, such as transcription factor binding, DNA accessibility, and the downstream edge of promoter-associated CpG islands. These features suggest an important role for antisense transcription in the regulation of gene expression and the maintenance of a promoter-associated chromatin environment.

  5. Protein tyrosine phosphatase-PEST (PTP-PEST) regulates mast cell-activating signals in PTP activity-dependent and -independent manners.

    Science.gov (United States)

    Motohashi, Satoru; Koizumi, Karen; Honda, Reika; Maruyama, Atsuko; Palmer, Helen E F; Mashima, Keisuke

    2014-01-01

    Aggregation of the high-affinity IgE receptor (FcεRI) in mast cells leads to degranulation and production of numerous cytokines and lipid mediators that promote allergic inflammation. Tyrosine phosphorylation of proteins in response to FcεRI aggregation has been implicated in mast cell activation. Here, we determined the role of PTP-PEST (encoded by PTPN12) in the regulation of mast cell activation using the RBL-2H3 rat basophilic leukemia cell line as a model. PTP-PEST expression was significantly induced upon FcεRI-crosslinking, and aggregation of FcεRI induced the phosphorylation of PTP-PEST at Ser39, thus resulting in the suppression of PTP activity. By overexpressing a phosphatase-dead mutant (PTP-PEST CS) and a constitutively active mutant (PTP-PEST SA) in RBL-2H3 cells, we showed that PTP-PEST decreased degranulation and enhanced IL-4 and IL-13 transcription in FcεRI-crosslinked RBL-2H3 cells, but PTP activity of PTP-PEST was not necessary for this regulation. However, FcεRI-induced TNF-α transcription was increased by the overexpression of PTP-PEST SA and suppressed by the overexpression of PTP-PEST CS. Taken together, these results suggest that PTP-PEST is involved in the regulation of FcεRI-mediated mast cell activation through at least two different processes represented by PTP activity-dependent and -independent pathways. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. Enhancer SINEs Link Pol III to Pol II Transcription in Neurons

    Directory of Open Access Journals (Sweden)

    Cristina Policarpi

    2017-12-01

    Full Text Available Summary: Spatiotemporal regulation of gene expression depends on the cooperation of multiple mechanisms, including the functional interaction of promoters with distally located enhancers. Here, we show that, in cortical neurons, a subset of short interspersed nuclear elements (SINEs located in the proximity of activity-regulated genes bears features of enhancers. Enhancer SINEs (eSINEs recruit the Pol III cofactor complex TFIIIC in a stimulus-dependent manner and are transcribed by Pol III in response to neuronal depolarization. Characterization of an eSINE located in proximity to the Fos gene (FosRSINE1 indicated that the FosRSINE1-encoded transcript interacts with Pol II at the Fos promoter and mediates Fos relocation to Pol II factories, providing an unprecedented molecular link between Pol III and Pol II transcription. Strikingly, knockdown of the FosRSINE1 transcript induces defects of both cortical radial migration in vivo and activity-dependent dendritogenesis in vitro, demonstrating that FosRSINE1 acts as a strong enhancer of Fos expression in diverse physiological contexts. : Spatiotemporal regulation of gene expression requires the interaction between promoters and distally located enhancers. Policarpi et al. identify a subset of SINEs that functions as enhancers for activity-dependent neuronal genes. The enhancer SINE FosRSINE1 regulates Fos transcription and is necessary for both activity-dependent dendritogenesis and proper brain development. Keywords: neuroscience, epigenetics, transcription, enhancers, SINEs, neuronal activity, neuronal development

  7. Activity-dependent increases in local oxygen consumption correlate with postsynaptic currents in the mouse cerebellum in vivo

    DEFF Research Database (Denmark)

    Mathiesen, Claus; Caesar, Kirsten; Thomsen, Kirsten Engelund

    2011-01-01

    Evoked neural activity correlates strongly with rises in cerebral metabolic rate of oxygen (CMRO(2)) and cerebral blood flow (CBF). Activity-dependent rises in CMRO(2) fluctuate with ATP turnover due to ion pumping. In vitro studies suggest that increases in cytosolic Ca(2+) stimulate oxidative m...

  8. Activity-dependent increases in local oxygen consumption correlate with post-synaptic currents in the mouse cerebellum in vivo

    DEFF Research Database (Denmark)

    Mathiesen, Claus; Caesar, Kirsten; Thomsen, Kirsten Joan

    2011-01-01

    Evoked neural activity correlates strongly with rises in cerebral metabolic rate of oxygen (CMRO2) and cerebral blood flow. Activity-dependent rises in CMRO2 fluctuate with ATP turnover due to ion pumping. In vitro studies suggest that increases in cytosolic Ca2+ stimulate oxidative metabolism vi...

  9. Copine-I: Modulator of NF-kappa B Transcription and Prostate Cancer Survival

    National Research Council Canada - National Science Library

    Mayo, Marty W; Creutz, Carl

    2008-01-01

    The purpose of our studies is to elucidate how Copine-I antagonizes NF-.B transcription. Nuclear factor-.B (NF-.B) is a dynamic transcription factor that regulates important biological processes involved in cancer initiation and progression...

  10. Linking Core Promoter Classes to Circadian Transcription.

    Directory of Open Access Journals (Sweden)

    Pål O Westermark

    2016-08-01

    Full Text Available Circadian rhythms in transcription are generated by rhythmic abundances and DNA binding activities of transcription factors. Propagation of rhythms to transcriptional initiation involves the core promoter, its chromatin state, and the basal transcription machinery. Here, I characterize core promoters and chromatin states of genes transcribed in a circadian manner in mouse liver and in Drosophila. It is shown that the core promoter is a critical determinant of circadian mRNA expression in both species. A distinct core promoter class, strong circadian promoters (SCPs, is identified in mouse liver but not Drosophila. SCPs are defined by specific core promoter features, and are shown to drive circadian transcriptional activities with both high averages and high amplitudes. Data analysis and mathematical modeling further provided evidence for rhythmic regulation of both polymerase II recruitment and pause release at SCPs. The analysis provides a comprehensive and systematic view of core promoters and their link to circadian mRNA expression in mouse and Drosophila, and thus reveals a crucial role for the core promoter in regulated, dynamic transcription.

  11. Intrinsic terminators in Mycoplasma hyopneumoniae transcription.

    Science.gov (United States)

    Fritsch, Tiago Ebert; Siqueira, Franciele Maboni; Schrank, Irene Silveira

    2015-04-08

    Mycoplasma hyopneumoniae, an important pathogen of swine, exhibits a low guanine and cytosine (GC) content genome. M. hyopneumoniae genome is organised in long transcriptional units and promoter sequences have been mapped upstream of all transcription units. These analysis provided insights into the gene organisation and transcription initiation at the genome scale. However, the presence of transcriptional terminator sequences in the M. hyopneumoniae genome is poorly understood. In silico analyses demonstrated the presence of putative terminators in 82% of the 33 monocistronic units (mCs) and in 74% of the 116 polycistronic units (pCs) considering different classes of terminators. The functional activity of 23 intrinsic terminators was confirmed by RT-PCR and qPCR. Analysis of all terminators found by three software algorithms, combined with experimental results, allowed us to propose a pattern of RNA hairpin formation during the termination process and to predict the location of terminators in the M. hyopneumoniae genome sequence. The stem-loop structures of intrinsic terminators of mycoplasma diverge from the pattern of terminators found in other bacteria due the low content of guanine and cytosine. In M. hyopneumoniae, transcription can end after a transcriptional unit and before its terminator sequence and can also continue past the terminator sequence with RNA polymerases gradually releasing the RNA.

  12. Activation-dependent mitochondrial translocation of Foxp3 in human hepatocytes

    International Nuclear Information System (INIS)

    Rojas, Joselyn; Teran-Angel, Guillermo; Barbosa, Luisa; Peterson, Darrell L.; Berrueta, Lisbeth; Salmen, Siham

    2016-01-01

    Foxp3 is considered to be the master regulator for the development and function of regulatory T cells (Treg). Recently Foxp3, has been detected in extra lymphoid tissue, and in hepatocytes and has been associated with hepatocellular carcinoma (HCC), although its role has not been defined. Since it is expected that there is a relationship between protein localization, activity and cellular function, the aim of this study was to explore the subcellular localization of Foxp3 in resting and stimulated human hepatocytes. Foxp3 expression was measured by flow cytometry, subcellular fractioning, and immunofluorescence, and this data was used to track the shuttling of Foxp3 in different subcellular compartments in hepatocytes (HepG2 cell line), stimulated by using the PKC activators (PMA), core and preS1/2 antigen from hepatitis B virus (HBV). Our data shows that besides the nuclear location, mitochondrial translocation was detected after stimulation with PMA and at to a lesser extent, with preS1/2. In addition, Foxp3 is localizes at outer mitochondrial membrane. These results suggest a non-canonical role of Foxp3 in the mitochondrial compartment in human hepatocytes, and opens a new field about their role in liver damages during HBV infection. - Highlights: • The expression and subcellular distribution of Foxp3, is modulated by PMA and preS1/2. • PMA and preS1/2 increase Foxp3 expression on HepG2. • PMA and preS1/2 induce foxp3 enrichment at mitochondrial, microsomal and nuclear compartments. • Results suggest a non-canonical function of Foxp3 or a mitochondrial transcriptional activity.

  13. Activation-dependent mitochondrial translocation of Foxp3 in human hepatocytes

    Energy Technology Data Exchange (ETDEWEB)

    Rojas, Joselyn; Teran-Angel, Guillermo; Barbosa, Luisa [Instituto de Inmunología Clínica, Facultad de Medicina, Universidad de Los Andes, Merida (Venezuela, Bolivarian Republic of); Peterson, Darrell L. [Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, Richmond, VA (United States); Berrueta, Lisbeth, E-mail: lberruet@ula.ve [Instituto de Inmunología Clínica, Facultad de Medicina, Universidad de Los Andes, Merida (Venezuela, Bolivarian Republic of); Division of Preventive Medicine, Brigham and Women' s Hospital, Harvard Medical School, Boston, MA (United States); Salmen, Siham, E-mail: sihamsa@ula.ve [Instituto de Inmunología Clínica, Facultad de Medicina, Universidad de Los Andes, Merida (Venezuela, Bolivarian Republic of)

    2016-05-01

    Foxp3 is considered to be the master regulator for the development and function of regulatory T cells (Treg). Recently Foxp3, has been detected in extra lymphoid tissue, and in hepatocytes and has been associated with hepatocellular carcinoma (HCC), although its role has not been defined. Since it is expected that there is a relationship between protein localization, activity and cellular function, the aim of this study was to explore the subcellular localization of Foxp3 in resting and stimulated human hepatocytes. Foxp3 expression was measured by flow cytometry, subcellular fractioning, and immunofluorescence, and this data was used to track the shuttling of Foxp3 in different subcellular compartments in hepatocytes (HepG2 cell line), stimulated by using the PKC activators (PMA), core and preS1/2 antigen from hepatitis B virus (HBV). Our data shows that besides the nuclear location, mitochondrial translocation was detected after stimulation with PMA and at to a lesser extent, with preS1/2. In addition, Foxp3 is localizes at outer mitochondrial membrane. These results suggest a non-canonical role of Foxp3 in the mitochondrial compartment in human hepatocytes, and opens a new field about their role in liver damages during HBV infection. - Highlights: • The expression and subcellular distribution of Foxp3, is modulated by PMA and preS1/2. • PMA and preS1/2 increase Foxp3 expression on HepG2. • PMA and preS1/2 induce foxp3 enrichment at mitochondrial, microsomal and nuclear compartments. • Results suggest a non-canonical function of Foxp3 or a mitochondrial transcriptional activity.

  14. The Transcription Bubble of the RNA Polymerase-Promoter Open Complex Exhibits Conformational Heterogeneity and Millisecond-Scale Dynamics : Implications for Transcription Start-Site Selection

    NARCIS (Netherlands)

    Robb, Nicole C.; Cordes, Thorben; Hwang, Ling Chin; Gryte, Kristofer; Duchi, Diego; Craggs, Timothy D.; Santoso, Yusdi; Weiss, Shimon; Ebright, Richard H.; Kapanidis, Achillefs N.

    2013-01-01

    Bacterial transcription is initiated after RNA polymerase (RNAP) binds to promoter DNA, melts similar to 14 bp around the transcription start site and forms a single-stranded "transcription bubble" within a catalytically active RNAP-DNA open complex (RPo). There is significant flexibility in the

  15. Fragile X Mental Retardation Protein Regulates Activity-Dependent Membrane Trafficking and Trans-Synaptic Signaling Mediating Synaptic Remodeling

    Science.gov (United States)

    Sears, James C.; Broadie, Kendal

    2018-01-01

    Fragile X syndrome (FXS) is the leading monogenic cause of autism and intellectual disability. The disease arises through loss of fragile X mental retardation protein (FMRP), which normally exhibits peak expression levels in early-use critical periods, and is required for activity-dependent synaptic remodeling during this transient developmental window. FMRP canonically binds mRNA to repress protein translation, with targets that regulate cytoskeleton dynamics, membrane trafficking, and trans-synaptic signaling. We focus here on recent advances emerging in these three areas from the Drosophila disease model. In the well-characterized central brain mushroom body (MB) olfactory learning/memory circuit, FMRP is required for activity-dependent synaptic remodeling of projection neurons innervating the MB calyx, with function tightly restricted to an early-use critical period. FMRP loss is phenocopied by conditional removal of FMRP only during this critical period, and rescued by FMRP conditional expression only during this critical period. Consistent with FXS hyperexcitation, FMRP loss defects are phenocopied by heightened sensory experience and targeted optogenetic hyperexcitation during this critical period. FMRP binds mRNA encoding Drosophila ESCRTIII core component Shrub (human CHMP4 homolog) to restrict Shrub translation in an activity-dependent mechanism only during this same critical period. Shrub mediates endosomal membrane trafficking, and perturbing Shrub expression is known to interfere with neuronal process pruning. Consistently, FMRP loss and Shrub overexpression targeted to projection neurons similarly causes endosomal membrane trafficking defects within synaptic boutons, and genetic reduction of Shrub strikingly rescues Drosophila FXS model defects. In parallel work on the well-characterized giant fiber (GF) circuit, FMRP limits iontophoretic dye loading into central interneurons, demonstrating an FMRP role controlling core neuronal properties through the

  16. Harnessing transcription for bioproduction in cyanobacteria

    DEFF Research Database (Denmark)

    Stensjö, Karin; Vavitsas, Konstantinos; Tyystjärvi, Taina

    2018-01-01

    Sustainable production of biofuels and other valuable compounds is one of our future challenges. One tempting possibility is to use photosynthetic cyanobacteria as production factories. Currently, tools for genetic engineering of cyanobacteria are yet not good enough to exploit the full potential...... of cyanobacteria. A wide variety of expression systems will be required to adjust both the expression of heterologous enzyme(s) and metabolic routes to the best possible balance, allowing the optimal production of a particular substance. In bacteria, transcription, especially the initiation of transcription, has...

  17. Calcineurin /NFAT activation-dependence of leptin synthesis and vascular growth in response to mechanical stretch

    Directory of Open Access Journals (Sweden)

    Nadia Soudani

    2016-09-01

    Full Text Available Background and Aims- Hypertension and obesity are important risk factors of cardiovascular disease. They are both associated with high leptin levels and have been shown to promote vascular hypertrophy, through the RhoA/ROCK and ERK1/2 phosphorylation. Calcineurin/NFAT activation also induces vascular hypertrophy by upregulating various genes. This study aimed to decipher whether a crosstalk exists between the RhoA/ROCK pathway, Ca+2/calcineurin/NFAT pathway, and ERK1/2 phosphorylation in the process of mechanical stretch-induced vascular smooth muscle cell (VSMC hypertrophy and leptin synthesis. Methods and Results- Rat portal vein (RPV organ culture was used to investigate the effect of mechanical stretch and exogenous leptin (3.1 nM on VSMC hypertrophy and leptin synthesis. Results showed that stretching the RPV significantly upregulated leptin secretion, mRNA and protein expression, which were inhibited by the calcium channel blocker nifedipine (10 μM, the selective calcineurin inhibitor FK506 (1 nM and the ERK1/2 inhibitor PD98059 (1 μM. The transcription inhibitor actinomycin D (0.1M and the translation inhibitor cycloheximide (1 mM significantly decreased stretch-induced leptin protein expression. Mechanical stretch or leptin caused an increase in wet weight changes and protein synthesis, considered as hypertrophic markers, while they were inhibited by FK506 (0.1 nM; 1 nM. In addition, stretch or exogenous leptin significantly increased calcineurin activity and MCIP1 expression whereas leptin induced NFAT nuclear translocation in VSMCs. Moreover, in response to stretch or exogenous leptin, the Rho inhibitor C3 exoenzyme (30 ng/mL, the ROCK inhibitor Y-27632 (10 μM, and the actin depolymerization agents Latrunculin B (50 nM and cytochalasin D (1 μM reduced calcineurin activation and NFAT nuclear translocation. ERK1/2 phosphorylation was inhibited by FK506 and C3. Conclusions- Mechanical stretch-induced VSMC hypertrophy and leptin

  18. The Transcription Factor Encyclopedia

    DEFF Research Database (Denmark)

    Yusuf, Dimas; Butland, Stefanie L; Swanson, Magdalena I

    2012-01-01

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

  19. Machine Dictation and Transcription.

    Science.gov (United States)

    Harvey, Evelyn; And Others

    This instructional package contains both an instructor's manual and a student's manual for a course in machine dictation and transcription. The instructor's manual contains an overview with tips on teaching the course, letters for dictation, and a key to the letters. The student's manual contains an overview of the course and of the skills needed…

  20. Transcriptional Regulation in Haematopoiesis:

    DEFF Research Database (Denmark)

    Lauridsen, Felicia K B

    with the capacity to both self-renew and differentiate. This thesis is built upon two studies, which investigate two different aspects of the haematopoietic system; heterogeneity within the HSC compartment (presented in manuscript I), and the interplay between transcription factors controlling granulocyte/ monocyte...

  1. Alterations in knee kinematics after partial medial meniscectomy are activity dependent.

    Science.gov (United States)

    Edd, Shannon N; Netravali, Nathan A; Favre, Julien; Giori, Nicholas J; Andriacchi, Thomas P

    2015-06-01

    Alterations in knee kinematics after partial meniscectomy have been linked to the increased risk of osteoarthritis in this population. Understanding differences in kinematics during static versus dynamic activities of increased demand can provide important information regarding the possible underlying mechanisms of these alterations. Differences in the following 2 kinematics measures will increase with activity demand: (1) the offset toward external tibial rotation for the meniscectomized limb compared with the contralateral limb during stance and (2) the difference in knee flexion angle at initial foot contact between the meniscectomized and contralateral limbs. Controlled laboratory study. This study compared side-to-side differences in knee flexion and rotation angles during static and dynamic activities. Thirteen patients (2 female) were tested in a motion capture laboratory at 6 ± 2 months after unilateral, arthroscopic, partial medial meniscectomy during a static reference pose and during 3 dynamic activities: walking, stair ascent, and stair descent. The meniscectomized limb demonstrated more external tibial rotation compared with the contralateral limb during dynamic activities, and there was a trend that this offset increased with activity demand (repeated-measures analysis of variance [ANOVA] for activity, P = .07; mean limb difference: static pose, -0.1° ± 3.3°, P = .5; walking, 1.2° ± 3.8°, P = .1; stair ascent, 2.0° ± 3.2°, P = .02; stair descent, 3.0° ± 3.5°, P = .005). Similarly, the meniscectomized knee was more flexed at initial contact than the contralateral limb during dynamic activities (repeated-measures ANOVA for activity P = .006; mean limb difference: reference pose, 1.0° ± 2.5°, P = .09; walking, 2.0° ± 3.9°, P = .05; stair ascent, 5.9° ± 5.3°, P = .009; stair descent, 3.5° ± 4.0°, P = .004). These results suggest both a structural element and a potential muscular element for the differences in kinematics after

  2. Requirements for DNA strand transfer during reverse transcription in mutant HIV-1 virions

    NARCIS (Netherlands)

    Berkhout, B.; van Wamel, J.; Klaver, B.

    1995-01-01

    Retroviruses convert their RNA genome into a DNA form by means of reverse transcription. According to the current model of reverse transcription, two strand transfer reactions are needed to synthesize a full-length DNA genome. Because reverse transcription is initiated close to the 5' end of the RNA

  3. DNA Topoisomerases in Transcription

    DEFF Research Database (Denmark)

    Rødgaard, Morten Terpager

    2015-01-01

    This Ph.D. thesis summarizes the main results of my studies on the interplay between DNA topoisomerases and transcription. The work was performed from 2011 to 2015 at Aarhus University in the Laboratory of Genome Research, and was supervised by associate professor Anni H. Andersen. Most of the ex......This Ph.D. thesis summarizes the main results of my studies on the interplay between DNA topoisomerases and transcription. The work was performed from 2011 to 2015 at Aarhus University in the Laboratory of Genome Research, and was supervised by associate professor Anni H. Andersen. Most...... topoisomerase-DNA cleavage complex. The second study is an investigation of how topoisomerases influence gene regulation by keeping the genome in an optimal topological state....

  4. Recovery of rhythmic activity in a central pattern generator: analysis of the role of neuromodulator and activity-dependent mechanisms.

    Science.gov (United States)

    Zhang, Yili; Golowasch, Jorge

    2011-11-01

    The pyloric network of decapods crustaceans can undergo dramatic rhythmic activity changes. Under normal conditions the network generates low frequency rhythmic activity that depends obligatorily on the presence of neuromodulatory input from the central nervous system. When this input is removed (decentralization) the rhythmic activity ceases. In the continued absence of this input, periodic activity resumes after a few hours in the form of episodic bursting across the entire network that later turns into stable rhythmic activity that is nearly indistinguishable from control (recovery). It has been proposed that an activity-dependent modification of ionic conductance levels in the pyloric pacemaker neuron drives the process of recovery of activity. Previous modeling attempts have captured some aspects of the temporal changes observed experimentally, but key features could not be reproduced. Here we examined a model in which slow activity-dependent regulation of ionic conductances and slower neuromodulator-dependent regulation of intracellular Ca(2+) concentration reproduce all the temporal features of this recovery. Key aspects of these two regulatory mechanisms are their independence and their different kinetics. We also examined the role of variability (noise) in the activity-dependent regulation pathway and observe that it can help to reduce unrealistic constraints that were otherwise required on the neuromodulator-dependent pathway. We conclude that small variations in intracellular Ca(2+) concentration, a Ca(2+) uptake regulation mechanism that is directly targeted by neuromodulator-activated signaling pathways, and variability in the Ca(2+) concentration sensing signaling pathway can account for the observed changes in neuronal activity. Our conclusions are all amenable to experimental analysis.

  5. Transcriptional networks controlling adipocyte differentiation

    DEFF Research Database (Denmark)

    Siersbæk, R; Mandrup, Susanne

    2011-01-01

    " of the transcription factor networks operating at specific time points during adipogenesis. Using such global "snapshots," we have demonstrated that dramatic remodeling of the chromatin template occurs within the first few hours following adipogenic stimulation and that many of the early transcription factors bind...... in a cooperative fashion to transcription factor hotspots. Such hotspots are likely to represent key chromatin nodes, where many adipogenic signaling pathways converge to drive the adipogenic transcriptional reprogramming....

  6. Synaptic synthesis, dephosphorylation, and degradation: a novel paradigm for an activity-dependent neuronal control of CDKL5.

    Science.gov (United States)

    La Montanara, Paolo; Rusconi, Laura; Locarno, Albina; Forti, Lia; Barbiero, Isabella; Tramarin, Marco; Chandola, Chetan; Kilstrup-Nielsen, Charlotte; Landsberger, Nicoletta

    2015-02-13

    Mutations in the X-linked CDKL5 (cyclin-dependent kinase-like 5) gene have been associated with several forms of neurodevelopmental disorders, including atypical Rett syndrome, autism spectrum disorders, and early infantile epileptic encephalopathy. Accordingly, loss of CDKL5 in mice results in autistic-like features and impaired neuronal communication. Although the biological functions of CDKL5 remain largely unknown, recent pieces of evidence suggest that CDKL5 is involved in neuronal plasticity. Herein, we show that, at all stages of development, neuronal depolarization induces a rapid increase in CDKL5 levels, mostly mediated by extrasomatic synthesis. In young neurons, this induction is prolonged, whereas in more mature neurons, NMDA receptor stimulation induces a protein phosphatase 1-dependent dephosphorylation of CDKL5 that is mandatory for its proteasome-dependent degradation. As a corollary, neuronal activity leads to a prolonged induction of CDKL5 levels in immature neurons but to a short lasting increase of the kinase in mature neurons. Recent results demonstrate that many genes associated with autism spectrum disorders are crucial components of the activity-dependent signaling networks regulating the composition, shape, and strength of the synapse. Thus, we speculate that CDKL5 deficiency disrupts activity-dependent signaling and the consequent synapse development, maturation, and refinement. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  7. Polyphenol Compound as a Transcription Factor Inhibitor

    Directory of Open Access Journals (Sweden)

    Seyeon Park

    2015-10-01

    Full Text Available A target-based approach has been used to develop novel drugs in many therapeutic fields. In the final stage of intracellular signaling, transcription factor–DNA interactions are central to most biological processes and therefore represent a large and important class of targets for human therapeutics. Thus, we focused on the idea that the disruption of protein dimers and cognate DNA complexes could impair the transcriptional activation and cell transformation regulated by these proteins. Historically, natural products have been regarded as providing the primary leading compounds capable of modulating protein–protein or protein-DNA interactions. Although their mechanism of action is not fully defined, polyphenols including flavonoids were found to act mostly as site-directed small molecule inhibitors on signaling. There are many reports in the literature of screening initiatives suggesting improved drugs that can modulate the transcription factor interactions responsible for disease. In this review, we focus on polyphenol compound inhibitors against dimeric forms of transcription factor components of intracellular signaling pathways (for instance, c-jun/c-fos (Activator Protein-1; AP-1, c-myc/max, Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB and β-catenin/T cell factor (Tcf.

  8. Polyphenol Compound as a Transcription Factor Inhibitor.

    Science.gov (United States)

    Park, Seyeon

    2015-10-30

    A target-based approach has been used to develop novel drugs in many therapeutic fields. In the final stage of intracellular signaling, transcription factor-DNA interactions are central to most biological processes and therefore represent a large and important class of targets for human therapeutics. Thus, we focused on the idea that the disruption of protein dimers and cognate DNA complexes could impair the transcriptional activation and cell transformation regulated by these proteins. Historically, natural products have been regarded as providing the primary leading compounds capable of modulating protein-protein or protein-DNA interactions. Although their mechanism of action is not fully defined, polyphenols including flavonoids were found to act mostly as site-directed small molecule inhibitors on signaling. There are many reports in the literature of screening initiatives suggesting improved drugs that can modulate the transcription factor interactions responsible for disease. In this review, we focus on polyphenol compound inhibitors against dimeric forms of transcription factor components of intracellular signaling pathways (for instance, c-jun/c-fos (Activator Protein-1; AP-1), c-myc/max, Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and β-catenin/T cell factor (Tcf)).

  9. Transcriptional networks in epithelial-mesenchymal transition.

    Directory of Open Access Journals (Sweden)

    Christo Venkov

    Full Text Available Epithelial-mesenchymal transition (EMT changes polarized epithelial cells into migratory phenotypes associated with loss of cell-cell adhesion molecules and cytoskeletal rearrangements. This form of plasticity is seen in mesodermal development, fibroblast formation, and cancer metastasis.Here we identify prominent transcriptional networks active during three time points of this transitional process, as epithelial cells become fibroblasts. DNA microarray in cultured epithelia undergoing EMT, validated in vivo, were used to detect various patterns of gene expression. In particular, the promoter sequences of differentially expressed genes and their transcription factors were analyzed to identify potential binding sites and partners. The four most frequent cis-regulatory elements (CREs in up-regulated genes were SRY, FTS-1, Evi-1, and GC-Box, and RNA inhibition of the four transcription factors, Atf2, Klf10, Sox11, and SP1, most frequently binding these CREs, establish their importance in the initiation and propagation of EMT. Oligonucleotides that block the most frequent CREs restrain EMT at early and intermediate stages through apoptosis of the cells.Our results identify new transcriptional interactions with high frequency CREs that modulate the stability of cellular plasticity, and may serve as targets for modulating these transitional states in fibroblasts.

  10. Transcript structure and domain display: a customizable transcript visualization tool.

    Science.gov (United States)

    Watanabe, Kenneth A; Ma, Kaiwang; Homayouni, Arielle; Rushton, Paul J; Shen, Qingxi J

    2016-07-01

    Transcript Structure and Domain Display (TSDD) is a publicly available, web-based program that provides publication quality images of transcript structures and domains. TSDD is capable of producing transcript structures from GFF/GFF3 and BED files. Alternatively, the GFF files of several model organisms have been pre-loaded so that users only needs to enter the locus IDs of the transcripts to be displayed. Visualization of transcripts provides many benefits to researchers, ranging from evolutionary analysis of DNA-binding domains to predictive function modeling. TSDD is freely available for non-commercial users at http://shenlab.sols.unlv.edu/shenlab/software/TSD/transcript_display.html : jeffery.shen@unlv.nevada.edu. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  11. The Mediator complex and transcription regulation

    Science.gov (United States)

    Poss, Zachary C.; Ebmeier, Christopher C.

    2013-01-01

    The Mediator complex is a multi-subunit assembly that appears to be required for regulating expression of most RNA polymerase II (pol II) transcripts, which include protein-coding and most non-coding RNA genes. Mediator and pol II function within the pre-initiation complex (PIC), which consists of Mediator, pol II, TFIIA, TFIIB, TFIID, TFIIE, TFIIF and TFIIH and is approximately 4.0 MDa in size. Mediator serves as a central scaffold within the PIC and helps regulate pol II activity in ways that remain poorly understood. Mediator is also generally targeted by sequence-specific, DNA-binding transcription factors (TFs) that work to control gene expression programs in response to developmental or environmental cues. At a basic level, Mediator functions by relaying signals from TFs directly to the pol II enzyme, thereby facilitating TF-dependent regulation of gene expression. Thus, Mediator is essential for converting biological inputs (communicated by TFs) to physiological responses (via changes in gene expression). In this review, we summarize an expansive body of research on the Mediator complex, with an emphasis on yeast and mammalian complexes. We focus on the basics that underlie Mediator function, such as its structure and subunit composition, and describe its broad regulatory influence on gene expression, ranging from chromatin architecture to transcription initiation and elongation, to mRNA processing. We also describe factors that influence Mediator structure and activity, including TFs, non-coding RNAs and the CDK8 module. PMID:24088064

  12. Manufacturing Initiative

    Data.gov (United States)

    National Aeronautics and Space Administration — The Advanced Manufacturing Technologies (AMT) Project supports multiple activities within the Administration's National Manufacturing Initiative. A key component of...

  13. Modelling reveals kinetic advantages of co-transcriptional splicing.

    Directory of Open Access Journals (Sweden)

    Stuart Aitken

    2011-10-01

    Full Text Available Messenger RNA splicing is an essential and complex process for the removal of intron sequences. Whereas the composition of the splicing machinery is mostly known, the kinetics of splicing, the catalytic activity of splicing factors and the interdependency of transcription, splicing and mRNA 3' end formation are less well understood. We propose a stochastic model of splicing kinetics that explains data obtained from high-resolution kinetic analyses of transcription, splicing and 3' end formation during induction of an intron-containing reporter gene in budding yeast. Modelling reveals co-transcriptional splicing to be the most probable and most efficient splicing pathway for the reporter transcripts, due in part to a positive feedback mechanism for co-transcriptional second step splicing. Model comparison is used to assess the alternative representations of reactions. Modelling also indicates the functional coupling of transcription and splicing, because both the rate of initiation of transcription and the probability that step one of splicing occurs co-transcriptionally are reduced, when the second step of splicing is abolished in a mutant reporter.

  14. Modelling reveals kinetic advantages of co-transcriptional splicing.

    Science.gov (United States)

    Aitken, Stuart; Alexander, Ross D; Beggs, Jean D

    2011-10-01

    Messenger RNA splicing is an essential and complex process for the removal of intron sequences. Whereas the composition of the splicing machinery is mostly known, the kinetics of splicing, the catalytic activity of splicing factors and the interdependency of transcription, splicing and mRNA 3' end formation are less well understood. We propose a stochastic model of splicing kinetics that explains data obtained from high-resolution kinetic analyses of transcription, splicing and 3' end formation during induction of an intron-containing reporter gene in budding yeast. Modelling reveals co-transcriptional splicing to be the most probable and most efficient splicing pathway for the reporter transcripts, due in part to a positive feedback mechanism for co-transcriptional second step splicing. Model comparison is used to assess the alternative representations of reactions. Modelling also indicates the functional coupling of transcription and splicing, because both the rate of initiation of transcription and the probability that step one of splicing occurs co-transcriptionally are reduced, when the second step of splicing is abolished in a mutant reporter.

  15. Nucleocytoplasmic shuttling of transcription factors

    DEFF Research Database (Denmark)

    Cartwright, P; Helin, K

    2000-01-01

    To elicit the transcriptional response following intra- or extracellular stimuli, the signals need to be transmitted to their site of action within the nucleus. The nucleocytoplasmic shuttling of transcription factors is a mechanism mediating this process. The activation and inactivation...... of the transcriptional response is essential for cells to progress through the cell cycle in a normal manner. The involvement of cytoplasmic and nuclear accessory molecules, and the general nuclear membrane transport components, are essential for this process. Although nuclear import and export for different...... transcription factor families are regulated by similar mechanisms, there are several differences that allow for the specific activation of each transcription factor. This review discusses the general import and export pathways found to be common amongst many different transcription factors, and highlights...

  16. Transcriptional Silencing of Retroviral Vectors

    DEFF Research Database (Denmark)

    Lund, Anders Henrik; Duch, M.; Pedersen, F.S.

    1996-01-01

    . Extinction of long-term vector expression has been observed after implantation of transduced hematopoietic cells as well as fibroblasts, myoblasts and hepatocytes. Here we review the influence of vector structure, integration site and cell type on transcriptional silencing. While down-regulation of proviral...... transcription is known from a number of cellular and animal models, major insight has been gained from studies in the germ line and embryonal cells of the mouse. Key elements for the transfer and expression of retroviral vectors, such as the viral transcriptional enhancer and the binding site for the t......RNA primer for reverse transcription may have a major influence on transcriptional silencing. Alterations of these elements of the vector backbone as well as the use of internal promoter elements from housekeeping genes may contribute to reduce transcriptional silencing. The use of cell culture and animal...

  17. DNA topology and transcription

    Science.gov (United States)

    Kouzine, Fedor; Levens, David; Baranello, Laura

    2014-01-01

    Chromatin is a complex assembly that compacts DNA inside the nucleus while providing the necessary level of accessibility to regulatory factors conscripted by cellular signaling systems. In this superstructure, DNA is the subject of mechanical forces applied by variety of molecular motors. Rather than being a rigid stick, DNA possesses dynamic structural variability that could be harnessed during critical steps of genome functioning. The strong relationship between DNA structure and key genomic processes necessitates the study of physical constrains acting on the double helix. Here we provide insight into the source, dynamics, and biology of DNA topological domains in the eukaryotic cells and summarize their possible involvement in gene transcription. We emphasize recent studies that might inspire and impact future experiments on the involvement of DNA topology in cellular functions. PMID:24755522

  18. Eukaryotic transcription factors

    DEFF Research Database (Denmark)

    Staby, Lasse; O'Shea, Charlotte; Willemoës, Martin

    2017-01-01

    Gene-specific transcription factors (TFs) are key regulatory components of signaling pathways, controlling, for example, cell growth, development, and stress responses. Their biological functions are determined by their molecular structures, as exemplified by their structured DNA-binding domains...... regions with function-related, short sequence motifs and molecular recognition features with structural propensities. This review focuses on molecular aspects of TFs, which represent paradigms of ID-related features. Through specific examples, we review how the ID-associated flexibility of TFs enables....... It is furthermore emphasized how classic biochemical concepts like allostery, conformational selection, induced fit, and feedback regulation are undergoing a revival with the appreciation of ID. The review also describes the most recent advances based on computational simulations of ID-based interaction mechanisms...

  19. Proteins mediating DNA loops effectively block transcription.

    Science.gov (United States)

    Vörös, Zsuzsanna; Yan, Yan; Kovari, Daniel T; Finzi, Laura; Dunlap, David

    2017-07-01

    Loops are ubiquitous topological elements formed when proteins simultaneously bind to two noncontiguous DNA sites. While a loop-mediating protein may regulate initiation at a promoter, the presence of the protein at the other site may be an obstacle for RNA polymerases (RNAP) transcribing a different gene. To test whether a DNA loop alters the extent to which a protein blocks transcription, the lac repressor (LacI) was used. The outcome of in vitro transcription along templates containing two LacI operators separated by 400 bp in the presence of LacI concentrations that produced both looped and unlooped molecules was visualized with scanning force microscopy (SFM). An analysis of transcription elongation complexes, moving for 60 s at an average of 10 nt/s on unlooped DNA templates, revealed that they more often surpassed LacI bound to the lower affinity O2 operator than to the highest affinity Os operator. However, this difference was abrogated in looped DNA molecules where LacI became a strong roadblock independently of the affinity of the operator. Recordings of transcription elongation complexes, using magnetic tweezers, confirmed that they halted for several minutes upon encountering a LacI bound to a single operator. The average pause lifetime is compatible with RNAP waiting for LacI dissociation, however, the LacI open conformation visualized in the SFM images also suggests that LacI could straddle RNAP to let it pass. Independently of the mechanism by which RNAP bypasses the LacI roadblock, the data indicate that an obstacle with looped topology more effectively interferes with transcription. © 2017 The Authors Protein Science published by Wiley Periodicals, Inc. on behalf of The Protein Society.

  20. A critical role for topoisomerase IIb and DNA double strand breaks in transcription.

    Science.gov (United States)

    Calderwood, Stuart K

    2016-05-26

    Recent studies have indicated a novel role for topoisomerase IIb in transcription. Transcription of heat shock genes, serum-induced immediate early genes and nuclear receptor-activated genes, each required DNA double strands generated by topoisomerase IIb. Such strand breaks seemed both necessary and sufficient for transcriptional activation. In addition, such transcription was associated with initiation of the DNA damage response pathways, including the activation of the enzymes: ataxia-telangiectasia mutated (ATM), DNA-dependent protein kinase and poly (ADP ribose) polymerase 1. DNA damage response signaling was involved both in transcription and in repair of DNA breaks generated by topoisomerase IIb.

  1. Unilateral initiatives

    International Nuclear Information System (INIS)

    Anon.

    1988-01-01

    This paper reports on arms control which is generally thought of in terms of formal negotiations with an opponent, with the resulting agreements embodied in a treaty. This is not surprising, since arms control discussions between opponents are both important and politically visible. There are, however, strong reasons for countries to consider and frequently take unilateral initiatives. To do so is entirely consistent with the established major precepts of arms control which state that arms control is designed to reduce the risk of war, the costs of preparing for war, and the death and destruction if war should come. Unilateral initiatives on what weapons are purchased, which ones are eliminated and how forces are deployed can all relate to these objectives. There are two main categories of motives for unilateral initiatives in arms control. In one category, internal national objectives are the dominant, often sole, driving force; the initiative is undertaken for our own good

  2. Ports Initiative

    Science.gov (United States)

    EPA's Ports Initiative works in collaboration with the port industry, communities, and government to improve environmental performance and increase economic prosperity. This effort helps people near ports breath cleaner air and live better lives.

  3. Epigenetics regulates transcription and pathogenesis in the parasite Trichomonas vaginalis.

    Science.gov (United States)

    Pachano, Tomas; Nievas, Yesica R; Lizarraga, Ayelen; Johnson, Patricia J; Strobl-Mazzulla, Pablo H; de Miguel, Natalia

    2017-06-01

    Trichomonas vaginalis is a common sexually transmitted parasite that colonizes the human urogenital tract. Infections range from asymptomatic to highly inflammatory, depending on the host and the parasite strain. Different T. vaginalis strains vary greatly in their adherence and cytolytic capacities. These phenotypic differences might be attributed to differentially expressed genes as a consequence of extra-genetic variation, such as epigenetic modifications. In this study, we explored the role of histone acetylation in regulating gene transcription and pathogenesis in T. vaginalis. Here, we show that histone 3 lysine acetylation (H3KAc) is enriched in nucleosomes positioned around the transcription start site of active genes (BAP1 and BAP2) in a highly adherent parasite strain; compared with the low acetylation abundance in contrast to that observed in a less-adherent strain that expresses these genes at low levels. Additionally, exposition of less-adherent strain with a specific histone deacetylases inhibitor, trichostatin A, upregulated the transcription of BAP1 and BAP2 genes in concomitance with an increase in H3KAc abundance and chromatin accessibility around their transcription start sites. Moreover, we demonstrated that the binding of initiator binding protein, the transcription factor responsible for the initiation of transcription of ~75% of known T. vaginalis genes, depends on the histone acetylation state around the metazoan-like initiator to which initiator binding protein binds. Finally, we found that trichostatin A treatment increased parasite aggregation and adherence to host cells. Our data demonstrated for the first time that H3KAc is a permissive histone modification that functions to mediate both transcription and pathogenesis of the parasite T. vaginalis. © 2017 John Wiley & Sons Ltd.

  4. Genomic and chromatin signals underlying transcription start-site selection

    DEFF Research Database (Denmark)

    Valen, Eivind; Sandelin, Albin Gustav

    2011-01-01

    A central question in cellular biology is how the cell regulates transcription and discerns when and where to initiate it. Locating transcription start sites (TSSs), the signals that specify them, and ultimately elucidating the mechanisms of regulated initiation has therefore been a recurrent theme....... In recent years substantial progress has been made towards this goal, spurred by the possibility of applying genome-wide, sequencing-based analysis. We now have a large collection of high-resolution datasets identifying locations of TSSs, protein-DNA interactions, and chromatin features over whole genomes...

  5. TcoF-DB: dragon database for human transcription co-factors and transcription factor interacting proteins

    KAUST Repository

    Schaefer, Ulf

    2010-10-21

    The initiation and regulation of transcription in eukaryotes is complex and involves a large number of transcription factors (TFs), which are known to bind to the regulatory regions of eukaryotic DNA. Apart from TF-DNA binding, protein-protein interaction involving TFs is an essential component of the machinery facilitating transcriptional regulation. Proteins that interact with TFs in the context of transcription regulation but do not bind to the DNA themselves, we consider transcription co-factors (TcoFs). The influence of TcoFs on transcriptional regulation and initiation, although indirect, has been shown to be significant with the functionality of TFs strongly influenced by the presence of TcoFs. While the role of TFs and their interaction with regulatory DNA regions has been well-studied, the association between TFs and TcoFs has so far been given less attention. Here, we present a resource that is comprised of a collection of human TFs and the TcoFs with which they interact. Other proteins that have a proven interaction with a TF, but are not considered TcoFs are also included. Our database contains 157 high-confidence TcoFs and additionally 379 hypothetical TcoFs. These have been identified and classified according to the type of available evidence for their involvement in transcriptional regulation and their presence in the cell nucleus. We have divided TcoFs into four groups, one of which contains high-confidence TcoFs and three others contain TcoFs which are hypothetical to different extents. We have developed the Dragon Database for Human Transcription Co-Factors and Transcription Factor Interacting Proteins (TcoF-DB). A web-based interface for this resource can be freely accessed at http://cbrc.kaust.edu.sa/tcof/ and http://apps.sanbi.ac.za/tcof/. © The Author(s) 2010.

  6. TcoF-DB: dragon database for human transcription co-factors and transcription factor interacting proteins

    KAUST Repository

    Schaefer, Ulf; Schmeier, Sebastian; Bajic, Vladimir B.

    2010-01-01

    The initiation and regulation of transcription in eukaryotes is complex and involves a large number of transcription factors (TFs), which are known to bind to the regulatory regions of eukaryotic DNA. Apart from TF-DNA binding, protein-protein interaction involving TFs is an essential component of the machinery facilitating transcriptional regulation. Proteins that interact with TFs in the context of transcription regulation but do not bind to the DNA themselves, we consider transcription co-factors (TcoFs). The influence of TcoFs on transcriptional regulation and initiation, although indirect, has been shown to be significant with the functionality of TFs strongly influenced by the presence of TcoFs. While the role of TFs and their interaction with regulatory DNA regions has been well-studied, the association between TFs and TcoFs has so far been given less attention. Here, we present a resource that is comprised of a collection of human TFs and the TcoFs with which they interact. Other proteins that have a proven interaction with a TF, but are not considered TcoFs are also included. Our database contains 157 high-confidence TcoFs and additionally 379 hypothetical TcoFs. These have been identified and classified according to the type of available evidence for their involvement in transcriptional regulation and their presence in the cell nucleus. We have divided TcoFs into four groups, one of which contains high-confidence TcoFs and three others contain TcoFs which are hypothetical to different extents. We have developed the Dragon Database for Human Transcription Co-Factors and Transcription Factor Interacting Proteins (TcoF-DB). A web-based interface for this resource can be freely accessed at http://cbrc.kaust.edu.sa/tcof/ and http://apps.sanbi.ac.za/tcof/. © The Author(s) 2010.

  7. Activity-dependent intracellular Ca2+ transients in unmyelinated nerve fibres of the isolated adult rat vagus nerve.

    Science.gov (United States)

    Wächtler, J; Mayer, C; Grafe, P

    1998-04-01

    Confocal laser scanning microscopy was used to follow changes in the free intracellular calcium concentration ([Ca2+]i) in nerve fibres and adjacent Schwann cells in isolated rat vagus nerves. [Ca2+]i was monitored by the Ca2+-sensitive fluorescent dyes Calcium Green-1 and Fura Red. Intracellular Ca2+ transients were observed during repetitive (1-50 Hz) supramaximal electrical stimulation or by bath application of ATP. Trains of action potentials were more effective at elongated, fibre-like structures of the vagus nerves, whereas ATP-induced Ca2+ transients were found predominantly in regions of Schwann cell bodies. Activity-induced Ca2+ signals were unaffected by pharmacological manipulation of intracellular Ca2+ stores, during long-lasting application of purinergic receptor agonists, or by substitution of extracellular Na+ with Li+. However, they were abolished in the presence of Ca2+-free bathing solution or after the blocking of Ca2+ channels with Cd2+. Ca2+ transients were also observed during Ca2+ action potentials. Such "Ca2+ spikes" were elicited by electrical stimulation in the presence of a combination of tetrodotoxin and K+ channel blockers. These data suggest that voltage-dependent Ca2+ channels, activated during short trains of Na+ action potentials, produce an increase in intra-axonal [Ca2+] of rat vagus nerves. We did not find evidence for activity-dependent Ca2+ transients in the Schwann cells surrounding the unmyelinated axons.

  8. Short-term memory of motor network performance via activity-dependent potentiation of Na+/K+ pump function.

    Science.gov (United States)

    Zhang, Hong-Yan; Sillar, Keith T

    2012-03-20

    Brain networks memorize previous performance to adjust their output in light of past experience. These activity-dependent modifications generally result from changes in synaptic strengths or ionic conductances, and ion pumps have only rarely been demonstrated to play a dynamic role. Locomotor behavior is produced by central pattern generator (CPG) networks and modified by sensory and descending signals to allow for changes in movement frequency, intensity, and duration, but whether or how the CPG networks recall recent activity is largely unknown. In Xenopus frog tadpoles, swim bout duration correlates linearly with interswim interval, suggesting that the locomotor network retains a short-term memory of previous output. We discovered an ultraslow, minute-long afterhyperpolarization (usAHP) in network neurons following locomotor episodes. The usAHP is mediated by an activity- and sodium spike-dependent enhancement of electrogenic Na(+)/K(+) pump function. By integrating spike frequency over time and linking the membrane potential of spinal neurons to network performance, the usAHP plays a dynamic role in short-term motor memory. Because Na(+)/K(+) pumps are ubiquitously expressed in neurons of all animals and because sodium spikes inevitably accompany network activity, the usAHP may represent a phylogenetically conserved but largely overlooked mechanism for short-term memory of neural network function. Copyright © 2012 Elsevier Ltd. All rights reserved.

  9. Effects of kainic acid lesions in lateral geniculate nucleus: activity dependence of retrograde axonal transport of fluorescent dyes.

    Science.gov (United States)

    Woodward, W R; Coull, B M

    1988-06-28

    Kainic acid lesions in the dorsal lateral geniculate nucleus of rats block the retrograde axonal transport of fluorescent dyes in corticogeniculate neurons without affecting the retrograde transport of D-aspartate or the orthograde transport of radiolabelled proteins in these neurons. This blocking of dye transport does not appear to be a consequence of kainic acid-induced damage to axon terminals in the geniculate since retinal ganglion cells are still able to transport dyes retrograde. A more likely explanation for these results is that fluorescent dye transport requires electrical activity in neurons, and elimination of the geniculate afferents to visual cortex reduces impulse traffic in cortical output fibers to a level below that required to support detectable dye transport. This interpretation is supported by the observation that kainic acid lesions also reduce retrograde transport of dyes in cortical neurons which project to the superior colliculus. Electrical stimulation in the subcortical white matter restores the transport of dye compounds in corticogeniculate neurons: evidence consistent with an activity-dependent mechanism of retrograde transport for these substances. These results provide evidence that axon terminals of retinal ganglion cells and corticogeniculate neurons survive in kainate-lesioned geniculates and are capable of normal neuronal function.

  10. Activity-dependent branching ratios in stocks, solar x-ray flux, and the Bak-Tang-Wiesenfeld sandpile model

    Science.gov (United States)

    Martin, Elliot; Shreim, Amer; Paczuski, Maya

    2010-01-01

    We define an activity-dependent branching ratio that allows comparison of different time series Xt . The branching ratio bx is defined as bx=E[ξx/x] . The random variable ξx is the value of the next signal given that the previous one is equal to x , so ξx={Xt+1∣Xt=x} . If bx>1 , the process is on average supercritical when the signal is equal to x , while if bxmarket hypothesis.” For stock volumes, solar x-ray flux intensities, and the Bak-Tang-Wiesenfeld (BTW) sandpile model, bx is supercritical for small values of activity and subcritical for the largest ones, indicating a tendency to return to a typical value. For stock volumes this tendency has an approximate power-law behavior. For solar x-ray flux and the BTW model, there is a broad regime of activity where bx≃1 , which we interpret as an indicator of critical behavior. This is true despite different underlying probability distributions for Xt and for ξx . For the BTW model the distribution of ξx is Gaussian, for x sufficiently larger than 1, and its variance grows linearly with x . Hence, the activity in the BTW model obeys a central limit theorem when sampling over past histories. The broad region of activity where bx is close to one disappears once bulk dissipation is introduced in the BTW model—supporting our hypothesis that it is an indicator of criticality.

  11. First Exon Length Controls Active Chromatin Signatures and Transcription

    Directory of Open Access Journals (Sweden)

    Nicole I. Bieberstein

    2012-07-01

    Full Text Available Here, we explore the role of splicing in transcription, employing both genome-wide analysis of human ChIP-seq data and experimental manipulation of exon-intron organization in transgenic cell lines. We show that the activating histone modifications H3K4me3 and H3K9ac map specifically to first exon-intron boundaries. This is surprising, because these marks help recruit general transcription factors (GTFs to promoters. In genes with long first exons, promoter-proximal levels of H3K4me3 and H3K9ac are greatly reduced; consequently, GTFs and RNA polymerase II are low at transcription start sites (TSSs and exhibit a second, promoter-distal peak from which transcription also initiates. In contrast, short first exons lead to increased H3K4me3 and H3K9ac at promoters, higher expression levels, accuracy in TSS usage, and a lower frequency of antisense transcription. Therefore, first exon length is predictive for gene activity. Finally, splicing inhibition and intron deletion reduce H3K4me3 levels and transcriptional output. Thus, gene architecture and splicing determines transcription quantity and quality as well as chromatin signatures.

  12. Prevalence of transcription promoters within archaeal operons and coding sequences.

    Science.gov (United States)

    Koide, Tie; Reiss, David J; Bare, J Christopher; Pang, Wyming Lee; Facciotti, Marc T; Schmid, Amy K; Pan, Min; Marzolf, Bruz; Van, Phu T; Lo, Fang-Yin; Pratap, Abhishek; Deutsch, Eric W; Peterson, Amelia; Martin, Dan; Baliga, Nitin S

    2009-01-01

    Despite the knowledge of complex prokaryotic-transcription mechanisms, generalized rules, such as the simplified organization of genes into operons with well-defined promoters and terminators, have had a significant role in systems analysis of regulatory logic in both bacteria and archaea. Here, we have investigated the prevalence of alternate regulatory mechanisms through genome-wide characterization of transcript structures of approximately 64% of all genes, including putative non-coding RNAs in Halobacterium salinarum NRC-1. Our integrative analysis of transcriptome dynamics and protein-DNA interaction data sets showed widespread environment-dependent modulation of operon architectures, transcription initiation and termination inside coding sequences, and extensive overlap in 3' ends of transcripts for many convergently transcribed genes. A significant fraction of these alternate transcriptional events correlate to binding locations of 11 transcription factors and regulators (TFs) inside operons and annotated genes-events usually considered spurious or non-functional. Using experimental validation, we illustrate the prevalence of overlapping genomic signals in archaeal transcription, casting doubt on the general perception of rigid boundaries between coding sequences and regulatory elements.

  13. Dynamic usage of transcription start sites within core promoters

    DEFF Research Database (Denmark)

    Kawaji, Hideya; Frith, Martin C; Katayama, Shintaro

    2006-01-01

    BACKGROUND: Mammalian promoters do not initiate transcription at single, well defined base pairs, but rather at multiple, alternative start sites spread across a region. We previously characterized the static structures of transcription start site usage within promoters at the base pair level......, based on large-scale sequencing of transcript 5' ends. RESULTS: In the present study we begin to explore the internal dynamics of mammalian promoters, and demonstrate that start site selection within many mouse core promoters varies among tissues. We also show that this dynamic usage of start sites...... is associated with CpG islands, broad and multimodal promoter structures, and imprinting. CONCLUSION: Our results reveal a new level of biologic complexity within promoters--fine-scale regulation of transcription starting events at the base pair level. These events are likely to be related to epigenetic...

  14. Viral DNA Replication Orientation and hnRNPs Regulate Transcription of the Human Papillomavirus 18 Late Promoter.

    Science.gov (United States)

    Wang, Xiaohong; Liu, Haibin; Ge, Hui; Ajiro, Masahiko; Sharma, Nishi R; Meyers, Craig; Morozov, Pavel; Tuschl, Thomas; Klar, Amar; Court, Donald; Zheng, Zhi-Ming

    2017-05-30

    The life cycle of human papillomaviruses (HPVs) is tightly linked to keratinocyte differentiation. Although expression of viral early genes is initiated immediately upon virus infection of undifferentiated basal cells, viral DNA amplification and late gene expression occur only in the mid to upper strata of the keratinocytes undergoing terminal differentiation. In this report, we show that the relative activity of HPV18 TATA-less late promoter P 811 depends on its orientation relative to that of the origin (Ori) of viral DNA replication and is sensitive to the eukaryotic DNA polymerase inhibitor aphidicolin. Additionally, transfected 70-nucleotide (nt)-long single-strand DNA oligonucleotides that are homologous to the region near Ori induce late promoter activity. We also found that promoter activation in raft cultures leads to production of the late promoter-associated, sense-strand transcription initiation RNAs (tiRNAs) and splice-site small RNAs (spliRNAs). Finally, a cis -acting AAGTATGCA core element that functions as a repressor to the promoter was identified. This element interacts with hnRNP D0B and hnRNP A/B factors. Point mutations in the core prevented binding of hnRNPs and increased the promoter activity. Confirming this result, knocking down the expression of both hnRNPs in keratinocytes led to increased promoter activity. Taking the data together, our study revealed the mechanism of how the HPV18 late promoter is regulated by DNA replication and host factors. IMPORTANCE It has been known for decades that the activity of viral late promoters is associated with viral DNA replication among almost all DNA viruses. However, the mechanism of how DNA replication activates the viral late promoter and what components of the replication machinery are involved remain largely unknown. In this study, we characterized the P 811 promoter region of HPV18 and demonstrated that its activation depends on the orientation of DNA replication. Using single

  15. Initial Study

    DEFF Research Database (Denmark)

    Torp, Kristian

    2009-01-01

    increased. In the initial study presented here, the time it takes to pass an intersection is studied in details. Two major signal-controlled four-way intersections in the center of the city Aalborg are studied in details to estimate the congestion levels in these intersections, based on the time it takes...

  16. Ethanol up-regulates nucleus accumbens neuronal activity dependent pentraxin (Narp): implications for alcohol-induced behavioral plasticity.

    Science.gov (United States)

    Ary, Alexis W; Cozzoli, Debra K; Finn, Deborah A; Crabbe, John C; Dehoff, Marlin H; Worley, Paul F; Szumlinski, Karen K

    2012-06-01

    Neuronal activity dependent pentraxin (Narp) interacts with α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate (AMPA) glutamate receptors to facilitate excitatory synapse formation by aggregating them at established synapses. Alcohol is well-characterized to influence central glutamatergic transmission, including AMPA receptor function. Herein, we examined the influence of injected and ingested alcohol upon Narp protein expression, as well as basal Narp expression in mouse lines selectively bred for high blood alcohol concentrations under limited access conditions. Alcohol up-regulated accumbens Narp levels, concomitant with increases in levels of the GluR1 AMPA receptor subunit. However, accumbens Narp or GluR1 levels did not vary as a function of selectively bred genotype. We next employed a Narp knock-out (KO) strategy to begin to understand the behavioral relevance of alcohol-induced changes in protein expression in several assays of alcohol reward. Compared to wild-type mice, Narp KO animals: fail to escalate daily intake of high alcohol concentrations under free-access conditions; shift their preference away from high alcohol concentrations with repeated alcohol experience; exhibit a conditioned place-aversion in response to the repeated pairing of 3 g/kg alcohol with a distinct environment and fail to exhibit alcohol-induced locomotor hyperactivity following repeated alcohol treatment. Narp deletion did not influence the daily intake of either food or water, nor did it alter any aspect of spontaneous or alcohol-induced motor activity, including the development of tolerance to its motor-impairing effects with repeated treatment. Taken together, these data indicate that Narp induction, and presumably subsequent aggregation of AMPA receptors, may be important for neuroplasticity within limbic subcircuits mediating or maintaining the rewarding properties of alcohol. Published by Elsevier Inc.

  17. Activity-dependent branching ratios in stocks, solar x-ray flux, and the Bak-Tang-Wiesenfeld sandpile model.

    Science.gov (United States)

    Martin, Elliot; Shreim, Amer; Paczuski, Maya

    2010-01-01

    We define an activity-dependent branching ratio that allows comparison of different time series X(t). The branching ratio b(x) is defined as b(x)=E[xi(x)/x]. The random variable xi(x) is the value of the next signal given that the previous one is equal to x, so xi(x)=[X(t+1) | X(t)=x]. If b(x)>1, the process is on average supercritical when the signal is equal to x, while if b(x)efficient market hypothesis." For stock volumes, solar x-ray flux intensities, and the Bak-Tang-Wiesenfeld (BTW) sandpile model, b(x) is supercritical for small values of activity and subcritical for the largest ones, indicating a tendency to return to a typical value. For stock volumes this tendency has an approximate power-law behavior. For solar x-ray flux and the BTW model, there is a broad regime of activity where b(x) approximately equal 1, which we interpret as an indicator of critical behavior. This is true despite different underlying probability distributions for X(t) and for xi(x). For the BTW model the distribution of xi(x) is Gaussian, for x sufficiently larger than 1, and its variance grows linearly with x. Hence, the activity in the BTW model obeys a central limit theorem when sampling over past histories. The broad region of activity where b(x) is close to one disappears once bulk dissipation is introduced in the BTW model-supporting our hypothesis that it is an indicator of criticality.

  18. Prominent fatigue in spinal muscular atrophy and spinal and bulbar muscular atrophy: evidence of activity-dependent conduction block.

    Science.gov (United States)

    Noto, Yu-ichi; Misawa, Sonoko; Mori, Masahiro; Kawaguchi, Naoki; Kanai, Kazuaki; Shibuya, Kazumoto; Isose, Sagiri; Nasu, Saiko; Sekiguchi, Yukari; Beppu, Minako; Ohmori, Shigeki; Nakagawa, Masanori; Kuwabara, Satoshi

    2013-09-01

    To clarify whether patients with spinal muscular atrophy (SMA) or spinal and bulbar muscular atrophy (SBMA) suffer disabling muscle fatigue, and whether activity-dependent conduction block (ADCB) contributes to their fatigue. ADCB is usually caused by reduced safety factor for impulse transmission in demyelinating diseases, whereas markedly increased axonal branching associated with collateral sprouting may reduce the safety factor in chronic lower motor neuron disorders. We assessed the fatigue severity scale (FSS) in 22 patients with SMA/SBMA, and in 100 disease controls (multiple sclerosis, myasthenia gravis, chronic inflammatory demyelinating polyneuropathy (CIDP), and axonal neuropathy). We then performed stimulated-single fibre electromyography (s-SFEMG) in the extensor digitorum communis (EDC) muscle of 21 SMA/SBMA patients, 6 CIDP patients, and 10 normal subjects. The FSS score was the highest in SMA/SBMA patients [4.9 ± 1.1 (mean ± SD)], with 81% of them complaining of disabling fatigue, compared with normal controls (3.5 ± 1.0), whereas patients with multiple sclerosis (4.3 ± 1.6), myasthenia gravis (4.0 ± 1.6) or CIDP (4.3 ± 1.4) also showed higher FSS score. When 2000 stimuli were delivered at 20 Hz in s-SFEMG, conduction block of single motor axons developed in 46% of patients with SMA/SBMA, and 40% of CIDP patients, but in none of the normal controls. SMA/SBMA patients frequently suffer from disabling fatigue presumably caused by ADCB induced by voluntary activity. ADCB could be the mechanism for muscle fatigue in chronic lower motor neuron diseases. Copyright © 2013 International Federation of Clinical Neurophysiology. Published by Elsevier Ireland Ltd. All rights reserved.

  19. Multi-timescale Modeling of Activity-Dependent Metabolic Coupling in the Neuron-Glia-Vasculature Ensemble

    KAUST Repository

    Jolivet, Renaud

    2015-02-26

    Glucose is the main energy substrate in the adult brain under normal conditions. Accumulating evidence, however, indicates that lactate produced in astrocytes (a type of glial cell) can also fuel neuronal activity. The quantitative aspects of this so-called astrocyte-neuron lactate shuttle (ANLS) are still debated. To address this question, we developed a detailed biophysical model of the brain’s metabolic interactions. Our model integrates three modeling approaches, the Buxton-Wang model of vascular dynamics, the Hodgkin-Huxley formulation of neuronal membrane excitability and a biophysical model of metabolic pathways. This approach provides a template for large-scale simulations of the neuron-glia-vasculature (NGV) ensemble, and for the first time integrates the respective timescales at which energy metabolism and neuronal excitability occur. The model is constrained by relative neuronal and astrocytic oxygen and glucose utilization, by the concentration of metabolites at rest and by the temporal dynamics of NADH upon activation. These constraints produced four observations. First, a transfer of lactate from astrocytes to neurons emerged in response to activity. Second, constrained by activity-dependent NADH transients, neuronal oxidative metabolism increased first upon activation with a subsequent delayed astrocytic glycolysis increase. Third, the model correctly predicted the dynamics of extracellular lactate and oxygen as observed in vivo in rats. Fourth, the model correctly predicted the temporal dynamics of tissue lactate, of tissue glucose and oxygen consumption, and of the BOLD signal as reported in human studies. These findings not only support the ANLS hypothesis but also provide a quantitative mathematical description of the metabolic activation in neurons and glial cells, as well as of the macroscopic measurements obtained during brain imaging.

  20. Multi-timescale Modeling of Activity-Dependent Metabolic Coupling in the Neuron-Glia-Vasculature Ensemble

    Science.gov (United States)

    Jolivet, Renaud; Coggan, Jay S.; Allaman, Igor; Magistretti, Pierre J.

    2015-01-01

    Glucose is the main energy substrate in the adult brain under normal conditions. Accumulating evidence, however, indicates that lactate produced in astrocytes (a type of glial cell) can also fuel neuronal activity. The quantitative aspects of this so-called astrocyte-neuron lactate shuttle (ANLS) are still debated. To address this question, we developed a detailed biophysical model of the brain’s metabolic interactions. Our model integrates three modeling approaches, the Buxton-Wang model of vascular dynamics, the Hodgkin-Huxley formulation of neuronal membrane excitability and a biophysical model of metabolic pathways. This approach provides a template for large-scale simulations of the neuron-glia-vasculature (NGV) ensemble, and for the first time integrates the respective timescales at which energy metabolism and neuronal excitability occur. The model is constrained by relative neuronal and astrocytic oxygen and glucose utilization, by the concentration of metabolites at rest and by the temporal dynamics of NADH upon activation. These constraints produced four observations. First, a transfer of lactate from astrocytes to neurons emerged in response to activity. Second, constrained by activity-dependent NADH transients, neuronal oxidative metabolism increased first upon activation with a subsequent delayed astrocytic glycolysis increase. Third, the model correctly predicted the dynamics of extracellular lactate and oxygen as observed in vivo in rats. Fourth, the model correctly predicted the temporal dynamics of tissue lactate, of tissue glucose and oxygen consumption, and of the BOLD signal as reported in human studies. These findings not only support the ANLS hypothesis but also provide a quantitative mathematical description of the metabolic activation in neurons and glial cells, as well as of the macroscopic measurements obtained during brain imaging. PMID:25719367

  1. Multi-timescale modeling of activity-dependent metabolic coupling in the neuron-glia-vasculature ensemble.

    Directory of Open Access Journals (Sweden)

    Renaud Jolivet

    2015-02-01

    Full Text Available Glucose is the main energy substrate in the adult brain under normal conditions. Accumulating evidence, however, indicates that lactate produced in astrocytes (a type of glial cell can also fuel neuronal activity. The quantitative aspects of this so-called astrocyte-neuron lactate shuttle (ANLS are still debated. To address this question, we developed a detailed biophysical model of the brain's metabolic interactions. Our model integrates three modeling approaches, the Buxton-Wang model of vascular dynamics, the Hodgkin-Huxley formulation of neuronal membrane excitability and a biophysical model of metabolic pathways. This approach provides a template for large-scale simulations of the neuron-glia-vasculature (NGV ensemble, and for the first time integrates the respective timescales at which energy metabolism and neuronal excitability occur. The model is constrained by relative neuronal and astrocytic oxygen and glucose utilization, by the concentration of metabolites at rest and by the temporal dynamics of NADH upon activation. These constraints produced four observations. First, a transfer of lactate from astrocytes to neurons emerged in response to activity. Second, constrained by activity-dependent NADH transients, neuronal oxidative metabolism increased first upon activation with a subsequent delayed astrocytic glycolysis increase. Third, the model correctly predicted the dynamics of extracellular lactate and oxygen as observed in vivo in rats. Fourth, the model correctly predicted the temporal dynamics of tissue lactate, of tissue glucose and oxygen consumption, and of the BOLD signal as reported in human studies. These findings not only support the ANLS hypothesis but also provide a quantitative mathematical description of the metabolic activation in neurons and glial cells, as well as of the macroscopic measurements obtained during brain imaging.

  2. A Transcription and Translation Protocol for Sensitive Cross-Cultural Team Research.

    Science.gov (United States)

    Clark, Lauren; Birkhead, Ana Sanchez; Fernandez, Cecilia; Egger, Marlene J

    2017-10-01

    Assurance of transcript accuracy and quality in interview-based qualitative research is foundational for data accuracy and study validity. Based on our experience in a cross-cultural ethnographic study of women's pelvic organ prolapse, we provide practical guidance to set up step-by-step interview transcription and translation protocols for team-based research on sensitive topics. Beginning with team decisions about level of detail in transcription, completeness, and accuracy, we operationalize the process of securing vendors to deliver the required quality of transcription and translation. We also share rubrics for assessing transcript quality and the team protocol for managing transcripts (assuring consistency of format, insertion of metadata, anonymization, and file labeling conventions) and procuring an acceptable initial translation of Spanish-language interviews. Accurate, complete, and systematically constructed transcripts in both source and target languages respond to the call for more transparency and reproducibility of scientific methods.

  3. Traveling Rocky Roads: The Consequences of Transcription-Blocking DNA Lesions on RNA Polymerase II.

    Science.gov (United States)

    Steurer, Barbara; Marteijn, Jurgen A

    2017-10-27

    The faithful transcription of eukaryotic genes by RNA polymerase II (RNAP2) is crucial for proper cell function and tissue homeostasis. However, transcription-blocking DNA lesions of both endogenous and environmental origin continuously challenge the progression of elongating RNAP2. The stalling of RNAP2 on a transcription-blocking lesion triggers a series of highly regulated events, including RNAP2 processing to make the lesion accessible for DNA repair, R-loop-mediated DNA damage signaling, and the initiation of transcription-coupled DNA repair. The correct execution and coordination of these processes is vital for resuming transcription following the successful repair of transcription-blocking lesions. Here, we outline recent insights into the molecular consequences of RNAP2 stalling on transcription-blocking DNA lesions and how these lesions are resolved to restore mRNA synthesis. Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.

  4. RNA-guided transcriptional regulation

    Science.gov (United States)

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

    2016-02-23

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

  5. Transcriptional control of megakaryocyte development.

    Science.gov (United States)

    Goldfarb, A N

    2007-10-15

    Megakaryocytes are highly specialized cells that arise from a bipotent megakaryocytic-erythroid progenitor (MEP). This developmental leap requires coordinated activation of megakaryocyte-specific genes, radical changes in cell cycle properties, and active prevention of erythroid differentiation. These programs result from upregulation of megakaryocyte-selective transcription factors, downregulation of erythroid-selective transcription factors and ongoing mediation of common erythro-megakaryocytic transcription factors. Unlike most developmental programs, no single lineage-unique family of master regulators exerts executive control over the megakaryocytic plan. Rather, an assemblage of non-unique factors and signals converge to determine lineage and differentiation. In human megakaryopoiesis, hereditary disorders of platelet production have confirmed contributions from three distinct transcription factor families. Murine models have extended this repertoire to include multiple additional factors. At a mechanistic level, the means by which these non-unique factors collaborate in the establishment of a perfectly unique cell type remains a central question.

  6. National Capital Planning Commission Meeting Transcripts

    Data.gov (United States)

    National Capital Planning Commission — Transcripts of the monthly (with the exception of August) National Capital Planning Commission meeting transcripts are provided for research to confirm actions taken...

  7. Recent behavioral history modifies coupling between cell activity and Arc gene transcription in hippocampal CA1 neurons.

    Science.gov (United States)

    Guzowski, John F; Miyashita, Teiko; Chawla, Monica K; Sanderson, Jennifer; Maes, Levi I; Houston, Frank P; Lipa, Peter; McNaughton, Bruce L; Worley, Paul F; Barnes, Carol A

    2006-01-24

    The ability of neurons to alter their transcriptional programs in response to synaptic input is of fundamental importance to the neuroplastic mechanisms underlying learning and memory. Because of technical limitations of conventional gene detection methods, the current view of activity-dependent neural transcription derives from experiments in which neurons are assumed quiescent until a signaling stimulus is given. The present study was designed to move beyond this static model by examining how earlier episodes of neural activity influence transcription of the immediate-early gene Arc. Using a sensitive FISH method that detects primary transcript at genomic alleles, the proportion of hippocampal CA1 neurons that activate transcription of Arc RNA was constant at approximately 40% in response to both a single novel exploration session and daily sessions repeated over 9 days. This proportion is similar to the percentage of active neurons defined electrophysiologically. However, this close correspondence was disrupted in rats exposed briefly, but repeatedly, to the same environment within a single day. Arc transcription in CA1 neurons declined dramatically after as few as four 5-min sessions, despite stable electrophysiological activity during all sessions. Additional experiments indicate that the decrement in Arc transcription occurred at the cellular, rather than synaptic level, and was not simply linked to habituation to novelty. Thus, the neural genomic response is governed by recent, but not remote, cell firing history in the behaving animal. This state-dependence of neuronal transcriptional coupling provides a mechanism of metaplasticity and may regulate capacity for synaptic modification in neural networks.

  8. A light- and calcium-gated transcription factor for imaging and manipulating activated neurons.

    Science.gov (United States)

    Wang, Wenjing; Wildes, Craig P; Pattarabanjird, Tanyaporn; Sanchez, Mateo I; Glober, Gordon F; Matthews, Gillian A; Tye, Kay M; Ting, Alice Y

    2017-09-01

    Activity remodels neurons, altering their molecular, structural, and electrical characteristics. To enable the selective characterization and manipulation of these neurons, we present FLARE, an engineered transcription factor that drives expression of fluorescent proteins, opsins, and other genetically encoded tools only in the subset of neurons that experienced activity during a user-defined time window. FLARE senses the coincidence of elevated cytosolic calcium and externally applied blue light, which together produce translocation of a membrane-anchored transcription factor to the nucleus to drive expression of any transgene. In cultured rat neurons, FLARE gives a light-to-dark signal ratio of 120 and a high- to low-calcium signal ratio of 10 after 10 min of stimulation. Opsin expression permitted functional manipulation of FLARE-marked neurons. In adult mice, FLARE also gave light- and motor-activity-dependent transcription in the cortex. Due to its modular design, minute-scale temporal resolution, and minimal dark-state leak, FLARE should be useful for the study of activity-dependent processes in neurons and other cells that signal with calcium.

  9. Activity-dependent formation of a vesicular inhibitory amino acid transporter gradient in the superior olivary complex of NMRI mice.

    Science.gov (United States)

    Ebbers, Lena; Weber, Maren; Nothwang, Hans Gerd

    2017-10-26

    factors involved in activity-dependent developmental processes in the auditory system. This would provide an important step forward concerning improved auditory rehabilitation in cases of congenital deafness.

  10. Genomic structure and cloning of two transcript isoforms of human Sp8.

    NARCIS (Netherlands)

    M.A. Milona (Maria-athina); J.E. Gough (Julie); A.J. Edgar (Alasdair)

    2004-01-01

    textabstractBACKGROUND: The Specificity proteins (Sp) are a family of transcription factors that have three highly conserved zinc-fingers located towards the carboxy-terminal that bind GC-boxes and assist in the initiation of gene transcription. Human Sp1-7 genes have been

  11. Activity-Dependent Bidirectional Regulation of GAD Expression in a Homeostatic Fashion Is Mediated by BDNF-Dependent and Independent Pathways

    Science.gov (United States)

    Hanno-Iijima, Yoko; Tanaka, Masami; Iijima, Takatoshi

    2015-01-01

    Homeostatic synaptic plasticity, or synaptic scaling, is a mechanism that tunes neuronal transmission to compensate for prolonged, excessive changes in neuronal activity. Both excitatory and inhibitory neurons undergo homeostatic changes based on synaptic transmission strength, which could effectively contribute to a fine-tuning of circuit activity. However, gene regulation that underlies homeostatic synaptic plasticity in GABAergic (GABA, gamma aminobutyric) neurons is still poorly understood. The present study demonstrated activity-dependent dynamic scaling in which NMDA-R (N-methyl-D-aspartic acid receptor) activity regulated the expression of GABA synthetic enzymes: glutamic acid decarboxylase 65 and 67 (GAD65 and GAD67). Results revealed that activity-regulated BDNF (brain-derived neurotrophic factor) release is necessary, but not sufficient, for activity-dependent up-scaling of these GAD isoforms. Bidirectional forms of activity-dependent GAD expression require both BDNF-dependent and BDNF-independent pathways, both triggered by NMDA-R activity. Additional results indicated that these two GAD genes differ in their responsiveness to chronic changes in neuronal activity, which could be partially caused by differential dependence on BDNF. In parallel to activity-dependent bidirectional scaling in GAD expression, the present study further observed that a chronic change in neuronal activity leads to an alteration in neurotransmitter release from GABAergic neurons in a homeostatic, bidirectional fashion. Therefore, the differential expression of GAD65 and 67 during prolonged changes in neuronal activity may be implicated in some aspects of bidirectional homeostatic plasticity within mature GABAergic presynapses. PMID:26241953

  12. Post-transcriptional trafficking and regulation of neuronal gene expression.

    Science.gov (United States)

    Goldie, Belinda J; Cairns, Murray J

    2012-02-01

    Intracellular messenger RNA (mRNA) traffic and translation must be highly regulated, both temporally and spatially, within eukaryotic cells to support the complex functional partitioning. This capacity is essential in neurons because it provides a mechanism for rapid input-restricted activity-dependent protein synthesis in individual dendritic spines. While this feature is thought to be important for synaptic plasticity, the structures and mechanisms that support this capability are largely unknown. Certainly specialized RNA binding proteins and binding elements in the 3' untranslated region (UTR) of translationally regulated mRNA are important, but the subtlety and complexity of this system suggests that an intermediate "specificity" component is also involved. Small non-coding microRNA (miRNA) are essential for CNS development and may fulfill this role by acting as the guide strand for mediating complex patterns of post-transcriptional regulation. In this review we examine post-synaptic gene regulation, mRNA trafficking and the emerging role of post-transcriptional gene silencing in synaptic plasticity.

  13. Method for determining transcriptional linkage by means of inhibition of deoxyribonucleic acid transcription by ultraviolet irradiation: evaluation in application to the investigation of in vivo transcription in bacteriophage T7

    International Nuclear Information System (INIS)

    Brautigam, A.R.

    1975-01-01

    A technique is presented for mapping promotor sites that utilizes the introduction of transcription-terminating lesions in DNA through uv irradiation which prevents transcription of genes in proportion to their distance from the promotor. This technique was applied to and evaluated in investigations of the transcriptional linkage of bacteriophage T7. All results substantiate the hypothesis that transcription in vivo does not proceed beyond the first uv lesion encountered in the template DNA and that such premature termination of transcription is the principal effect of the uv irradiation on the transcriptional template function of DNA. UV-induced inhibition of the initiation of transcription is insignificant by comparison. Uv inactivation of expression of individual T7 genes was found to follow pseudo first-order kinetics, allowing a gene-specific uv inactivation cross section to be evaluated for each gene. Promotor locations were inferred from the discontinuity in the numerical values of inactivation cross sections arising at the start of each new unit. By such analysis the bacteriophage T7 genome was found to consist of seven transcription units. In vivo E. coli RNA polymerase transcribes the T7 early region as a single unit from a pomotor region located at the left end of the genome. The T7 late region was found to consist of six transcription units, with promotors located just ahead of genes 1.7, 7, 9, 11, 13 and 17

  14. Reactive Oxygen Species-Mediated Loss of Synaptic Akt1 Signaling Leads to Deficient Activity-Dependent Protein Translation Early in Alzheimer's Disease.

    Science.gov (United States)

    Ahmad, Faraz; Singh, Kunal; Das, Debajyoti; Gowaikar, Ruturaj; Shaw, Eisha; Ramachandran, Arathy; Rupanagudi, Khader Valli; Kommaddi, Reddy Peera; Bennett, David A; Ravindranath, Vijayalakshmi

    2017-12-01

    Synaptic deficits are known to underlie the cognitive dysfunction seen in Alzheimer's disease (AD). Generation of reactive oxygen species (ROS) by β-amyloid has also been implicated in AD pathogenesis. However, it is unclear whether ROS contributes to synaptic dysfunction seen in AD pathogenesis and, therefore, we examined whether altered redox signaling could contribute to synaptic deficits in AD. Activity dependent but not basal translation was impaired in synaptoneurosomes from 1-month old presymptomatic APP Swe /PS1ΔE9 (APP/PS1) mice, and this deficit was sustained till middle age (MA, 9-10 months). ROS generation leads to oxidative modification of Akt1 in the synapse and consequent reduction in Akt1-mechanistic target of rapamycin (mTOR) signaling, leading to deficiency in activity-dependent protein translation. Moreover, we found a similar loss of activity-dependent protein translation in synaptoneurosomes from postmortem AD brains. Loss of activity-dependent protein translation occurs presymptomatically early in the pathogenesis of AD. This is caused by ROS-mediated loss of pAkt1, leading to reduced synaptic Akt1-mTOR signaling and is rescued by overexpression of Akt1. ROS-mediated damage is restricted to the synaptosomes, indicating selectivity. We demonstrate that ROS-mediated oxidative modification of Akt1 contributes to synaptic dysfunction in AD, seen as loss of activity-dependent protein translation that is essential for synaptic plasticity and maintenance. Therapeutic strategies promoting Akt1-mTOR signaling at synapses may provide novel target(s) for disease-modifying therapy in AD. Antioxid. Redox Signal. 27, 1269-1280.

  15. Nuclear cGMP-dependent kinase regulates gene expression via activity-dependent recruitment of a conserved histone deacetylase complex.

    Directory of Open Access Journals (Sweden)

    Yan Hao

    2011-05-01

    Full Text Available Elevation of the second messenger cGMP by nitric oxide (NO activates the cGMP-dependent protein kinase PKG, which is key in regulating cardiovascular, intestinal, and neuronal functions in mammals. The NO-cGMP-PKG signaling pathway is also a major therapeutic target for cardiovascular and male reproductive diseases. Despite widespread effects of PKG activation, few molecular targets of PKG are known. We study how EGL-4, the Caenorhabditis elegans PKG ortholog, modulates foraging behavior and egg-laying and seeks the downstream effectors of EGL-4 activity. Using a combination of unbiased forward genetic screen and proteomic analysis, we have identified a conserved SAEG-1/SAEG-2/HDA-2 histone deacetylase complex that is specifically recruited by activated nuclear EGL-4. Gene expression profiling by microarrays revealed >40 genes that are sensitive to EGL-4 activity in a SAEG-1-dependent manner. We present evidence that EGL-4 controls egg laying via one of these genes, Y45F10C.2, which encodes a novel protein that is expressed exclusively in the uterine epithelium. Our results indicate that, in addition to cytoplasmic functions, active EGL-4/PKG acts in the nucleus via a conserved Class I histone deacetylase complex to regulate gene expression pertinent to behavioral and physiological responses to cGMP. We also identify transcriptional targets of EGL-4 that carry out discrete components of the physiological response.

  16. Transcription of the soybean leghemoglobin genes during nodule development

    DEFF Research Database (Denmark)

    Marcker, Anne; Ø Jensen, Erik; Marcker, Kjeld A

    1984-01-01

    During the early stages of soybean nodule development the leghemoglobin (Lb) genes are activated sequentially in the opposite order to which they are arranged in the soybean genome. At a specific stage after the initial activation of all the Lb genes, a large increment occurs in the transcription...... of the Lb(c1), Lb(c3) and Lb(a) genes while the transcription of the Lb(c2) gene is not amplified to a similar extent. All the Lb genes retain significant activity for a long period during the lifetime of a nodule. Consequently the soybean Lb genes are not regulated by a developmental gene switching...

  17. Transcription factor Tbx3 is required for the specification of the atrioventricular conduction system

    NARCIS (Netherlands)

    Bakker, Martijn L.; Boukens, Bastiaan J.; Mommersteeg, Mathilda T. M.; Brons, Janynke F.; Wakker, Vincent; Moorman, Antoon F. M.; Christoffels, Vincent M.

    2008-01-01

    The cardiac conduction system consists of distinctive heart muscle cells that initiate and propagate the electric impulse required for coordinated contraction. The conduction system expresses the transcriptional repressor Tbx3, which is required for vertebrate development and controls the formation

  18. A mechanistic overview of herbal medicine and botanical compounds to target transcriptional factors in Breast cancer.

    Science.gov (United States)

    Zhao, Yingke; Liu, Yue

    2018-04-01

    The abnormalities of transcription factors, such as NF-κB, STAT, estrogen receptor, play a critical role in the initiation and progression of breast cancer. Due to the limitation of current treatment, transcription factors could be promising therapeutic targets, which have received close attention. In this review, we introduced herbal medicines, as well as botanical compounds that had been verified with anti-tumor properties via regulating transcription factors. Herbs, compounds, as well as formulae reported with various transcriptional targets, were summarized thoroughly, to provide implication for the future research on basic experiment and clinical application. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. The Mediator complex: a central integrator of transcription

    Science.gov (United States)

    Allen, Benjamin L.; Taatjes, Dylan J.

    2016-01-01

    The RNA polymerase II (pol II) enzyme transcribes all protein-coding and most non-coding RNA genes and is globally regulated by Mediator, a large, conformationally flexible protein complex with variable subunit composition (for example, a four-subunit CDK8 module can reversibly associate). These biochemical characteristics are fundamentally important for Mediator's ability to control various processes important for transcription, including organization of chromatin architecture and regulation of pol II pre-initiation, initiation, re-initiation, pausing, and elongation. Although Mediator exists in all eukaryotes, a variety of Mediator functions appear to be specific to metazoans, indicative of more diverse regulatory requirements. PMID:25693131

  20. Activity-dependent regulation of MHC class I expression in the developing primary visual cortex of the common marmoset monkey

    Directory of Open Access Journals (Sweden)

    Schlumbohm Christina

    2011-01-01

    Full Text Available Abstract Background Several recent studies have highlighted the important role of immunity-related molecules in synaptic plasticity processes in the developing and adult mammalian brains. It has been suggested that neuronal MHCI (major histocompatibility complex class I genes play a role in the refinement and pruning of synapses in the developing visual system. As a fast evolutionary rate may generate distinct properties of molecules in different mammalian species, we studied the expression of MHCI molecules in a nonhuman primate, the common marmoset monkey (Callithrix jacchus. Methods and results Analysis of expression levels of MHCI molecules in the developing visual cortex of the common marmoset monkeys revealed a distinct spatio-temporal pattern. High levels of expression were detected very early in postnatal development, at a stage when synaptogenesis takes place and ocular dominance columns are formed. To determine whether the expression of MHCI molecules is regulated by retinal activity, animals were subjected to monocular enucleation. Levels of MHCI heavy chain subunit transcripts in the visual cortex were found to be elevated in response to monocular enucleation. Furthermore, MHCI heavy chain immunoreactivity revealed a banded pattern in layer IV of the visual cortex in enucleated animals, which was not observed in control animals. This pattern of immunoreactivity indicated that higher expression levels were associated with retinal activity coming from the intact eye. Conclusions These data demonstrate that, in the nonhuman primate brain, expression of MHCI molecules is regulated by neuronal activity. Moreover, this study extends previous findings by suggesting a role for neuronal MHCI molecules during synaptogenesis in the visual cortex.

  1. Openness initiative

    International Nuclear Information System (INIS)

    Duncan, S.S.

    1995-01-01

    Although antinuclear campaigns seem to be effective, public communication and education efforts on low-level radioactive waste have mixed results. Attempts at public information programs on low-level radioactive waste still focus on influencing public opinion. A question then is: open-quotes Is it preferable to have a program focus on public education that will empower individuals to make informed decisions rather than trying to influence them in their decisions?close quotes To address this question, a case study with both quantitative and qualitative data will be used. The Ohio Low-Level Radioactive Waste Education Program has a goal to provide people with information they want/need to make their own decisions. The program initiated its efforts by conducting a statewide survey to determine information needed by people and where they turned for that information. This presentation reports data from the survey and then explores the program development process in which programs were designed and presented using the information. Pre and post data from the programs reveal attitude and knowledge shifts

  2. Openness initiative

    Energy Technology Data Exchange (ETDEWEB)

    Duncan, S.S. [Los Alamos National Lab., NM (United States)

    1995-12-31

    Although antinuclear campaigns seem to be effective, public communication and education efforts on low-level radioactive waste have mixed results. Attempts at public information programs on low-level radioactive waste still focus on influencing public opinion. A question then is: {open_quotes}Is it preferable to have a program focus on public education that will empower individuals to make informed decisions rather than trying to influence them in their decisions?{close_quotes} To address this question, a case study with both quantitative and qualitative data will be used. The Ohio Low-Level Radioactive Waste Education Program has a goal to provide people with information they want/need to make their own decisions. The program initiated its efforts by conducting a statewide survey to determine information needed by people and where they turned for that information. This presentation reports data from the survey and then explores the program development process in which programs were designed and presented using the information. Pre and post data from the programs reveal attitude and knowledge shifts.

  3. Initiative hard coal; Initiative Steinkohle

    Energy Technology Data Exchange (ETDEWEB)

    Leonhardt, J.

    2007-08-02

    In order to decrease the import dependence of hard coal in the European Union, the author has submitted suggestions to the director of conventional sources of energy (directorate general for energy and transport) of the European community, which found a positive resonance. These suggestions are summarized in an elaboration 'Initiative Hard Coal'. After clarifying the starting situation and defining the target the presupposition for a better use of hard coal deposits as raw material in the European Union are pointed out. On that basis concrete suggestions for measures are made. Apart from the conditions of the deposits it concerns thereby also new mining techniques and mining-economical developments, connected with tasks for the mining-machine industry. (orig.)

  4. Multiple promoters and alternative splicing: Hoxa5 transcriptional complexity in the mouse embryo.

    Directory of Open Access Journals (Sweden)

    Yan Coulombe

    2010-05-01

    Full Text Available The genomic organization of Hox clusters is fundamental for the precise spatio-temporal regulation and the function of each Hox gene, and hence for correct embryo patterning. Multiple overlapping transcriptional units exist at the Hoxa5 locus reflecting the complexity of Hox clustering: a major form of 1.8 kb corresponding to the two characterized exons of the gene and polyadenylated RNA species of 5.0, 9.5 and 11.0 kb. This transcriptional intricacy raises the question of the involvement of the larger transcripts in Hox function and regulation.We have undertaken the molecular characterization of the Hoxa5 larger transcripts. They initiate from two highly conserved distal promoters, one corresponding to the putative Hoxa6 promoter, and a second located nearby Hoxa7. Alternative splicing is also involved in the generation of the different transcripts. No functional polyadenylation sequence was found at the Hoxa6 locus and all larger transcripts use the polyadenylation site of the Hoxa5 gene. Some larger transcripts are potential Hoxa6/Hoxa5 bicistronic units. However, even though all transcripts could produce the genuine 270 a.a. HOXA5 protein, only the 1.8 kb form is translated into the protein, indicative of its essential role in Hoxa5 gene function. The Hoxa6 mutation disrupts the larger transcripts without major phenotypic impact on axial specification in their expression domain. However, Hoxa5-like skeletal anomalies are observed in Hoxa6 mutants and these defects can be explained by the loss of expression of the 1.8 kb transcript. Our data raise the possibility that the larger transcripts may be involved in Hoxa5 gene regulation.Our observation that the Hoxa5 larger transcripts possess a developmentally-regulated expression combined to the increasing sum of data on the role of long noncoding RNAs in transcriptional regulation suggest that the Hoxa5 larger transcripts may participate in the control of Hox gene expression.

  5. Transcriptional regulation of hepatic lipogenesis.

    Science.gov (United States)

    Wang, Yuhui; Viscarra, Jose; Kim, Sun-Joong; Sul, Hei Sook

    2015-11-01

    Fatty acid and fat synthesis in the liver is a highly regulated metabolic pathway that is important for very low-density lipoprotein (VLDL) production and thus energy distribution to other tissues. Having common features at their promoter regions, lipogenic genes are coordinately regulated at the transcriptional level. Transcription factors, such as upstream stimulatory factors (USFs), sterol regulatory element-binding protein 1C (SREBP1C), liver X receptors (LXRs) and carbohydrate-responsive element-binding protein (ChREBP) have crucial roles in this process. Recently, insights have been gained into the signalling pathways that regulate these transcription factors. After feeding, high blood glucose and insulin levels activate lipogenic genes through several pathways, including the DNA-dependent protein kinase (DNA-PK), atypical protein kinase C (aPKC) and AKT-mTOR pathways. These pathways control the post-translational modifications of transcription factors and co-regulators, such as phosphorylation, acetylation or ubiquitylation, that affect their function, stability and/or localization. Dysregulation of lipogenesis can contribute to hepatosteatosis, which is associated with obesity and insulin resistance.

  6. Transcription factor-based biosensor

    Science.gov (United States)

    Dietrich, Jeffrey A; Keasling, Jay D

    2013-10-08

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

  7. Orchestration of Floral Initiation by APETALA1

    NARCIS (Netherlands)

    Kaufmann, K.; Muino Acuna, J.M.

    2010-01-01

    The MADS-domain transcription factor APETALA1 (AP1) is a key regulator of Arabidopsis flower development. To understand the molecular mechanisms underlying AP1 function, we identified its target genes during floral initiation using a combination of gene expression profiling and genome-wide binding

  8. Transcriptional regulation by nonclassical action of thyroid hormone

    Directory of Open Access Journals (Sweden)

    Moeller Lars C

    2011-08-01

    Full Text Available Abstract Thyroid hormone (TH is essential for normal development, growth and metabolism. Its effects were thought to be principally mediated through triiodothyronine (T3, acting as a ligand for the nuclear TH receptors (TRs α and β residing on thyroid hormone response elements (TREs in the promoter of TH target genes. In this classical model of TH action, T3 binding to TRs leads to recruitment of basal transcription factors and increased transcription of TH responsive genes. Recently, the concept of TH action on gene expression has become more diverse and now includes nonclassical actions of T3 and T4: T3 has been shown to activate PI3K via the TRs, which ultimately increases transcription of certain genes, e.g. HIF-1α. Additionally, both T3 and thyroxine (T4 can bind to a membrane integrin, αvβ3, which leads to activation of the PI3K and MAPK signal transduction pathways and finally also increases gene transcription, e.g. of the FGF2 gene. Therefore, these initially nongenomic, nonclassical actions seem to serve as additional interfaces for transcriptional regulation by TH. Aim of this perspective is to summarize the genes that are currently known to be induced by nonclassical TH action and the mechanisms involved.

  9. Archaeal RNA polymerase arrests transcription at DNA lesions.

    Science.gov (United States)

    Gehring, Alexandra M; Santangelo, Thomas J

    2017-01-01

    Transcription elongation is not uniform and transcription is often hindered by protein-bound factors or DNA lesions that limit translocation and impair catalysis. Despite the high degree of sequence and structural homology of the multi-subunit RNA polymerases (RNAP), substantial differences in response to DNA lesions have been reported. Archaea encode only a single RNAP with striking structural conservation with eukaryotic RNAP II (Pol II). Here, we demonstrate that the archaeal RNAP from Thermococcus kodakarensis is sensitive to a variety of DNA lesions that pause and arrest RNAP at or adjacent to the site of DNA damage. DNA damage only halts elongation when present in the template strand, and the damage often results in RNAP arresting such that the lesion would be encapsulated with the transcription elongation complex. The strand-specific halt to archaeal transcription elongation on modified templates is supportive of RNAP recognizing DNA damage and potentially initiating DNA repair through a process akin to the well-described transcription-coupled DNA repair (TCR) pathways in Bacteria and Eukarya.

  10. Oncogenes Activate an Autonomous Transcriptional Regulatory Circuit That Drives Glioblastoma

    Directory of Open Access Journals (Sweden)

    Dinesh K. Singh

    2017-01-01

    Full Text Available Efforts to identify and target glioblastoma (GBM drivers have primarily focused on receptor tyrosine kinases (RTKs. Clinical benefits, however, have been elusive. Here, we identify an SRY-related box 2 (SOX2 transcriptional regulatory network that is independent of upstream RTKs and capable of driving glioma-initiating cells. We identified oligodendrocyte lineage transcription factor 2 (OLIG2 and zinc-finger E-box binding homeobox 1 (ZEB1, which are frequently co-expressed irrespective of driver mutations, as potential SOX2 targets. In murine glioma models, we show that different combinations of tumor suppressor and oncogene mutations can activate Sox2, Olig2, and Zeb1 expression. We demonstrate that ectopic co-expression of the three transcription factors can transform tumor-suppressor-deficient astrocytes into glioma-initiating cells in the absence of an upstream RTK oncogene. Finally, we demonstrate that the transcriptional inhibitor mithramycin downregulates SOX2 and its target genes, resulting in markedly reduced proliferation of GBM cells in vivo.

  11. Characterization of a novel radiation-inducible transcript, uscA, and analysis of its transcriptional regulation

    Energy Technology Data Exchange (ETDEWEB)

    Lim, Sang Yong; Kim, Dong Ho; Joe, Min Ho

    2010-03-15

    The transcriptional expression of the uscA promote (P{sub uscA}) only occurred under aerobic conditions and a dose of 2Gy maximally activated transcription of P{sub uscA}. However, various environmental stress including physical shocks (pH, temperature, osmotic shock), DNA damaging agents (UV and MMC) or oxidative stressagents (paraquat, menadione, and H{sub 2}O{sub 2}) didn't cause the transcriptional activationof P{sub uscA}. The transcription of uscA was initiated at 170 bp upstream of the cyoA start codon, and ended around the ampG stop codon. The size of uscA was determined through reverse transcription assay, approximately 250 bp. The deletion analysis of uscA promoter demonstrates that radiation inducibility of P{sub uscA} is mediated by sequences present between -20 and +111 relativeto +1 of P{sub uscA} and radiation causes P{sub uscA} activation thorough permitting the expression that is repressed under non-irradiated conditions

  12. Characterization of a novel radiation-inducible transcript, uscA, and analysis of its transcriptional regulation

    International Nuclear Information System (INIS)

    Lim, Sang Yong; Kim, Dong Ho; Joe, Min Ho

    2010-03-01

    The transcriptional expression of the uscA promote (P uscA ) only occurred under aerobic conditions and a dose of 2Gy maximally activated transcription of P uscA . However, various environmental stress including physical shocks (pH, temperature, osmotic shock), DNA damaging agents (UV and MMC) or oxidative stressagents (paraquat, menadione, and H 2 O 2 ) didn't cause the transcriptional activationof P uscA . The transcription of uscA was initiated at 170 bp upstream of the cyoA start codon, and ended around the ampG stop codon. The size of uscA was determined through reverse transcription assay, approximately 250 bp. The deletion analysis of uscA promoter demonstrates that radiation inducibility of P uscA is mediated by sequences present between -20 and +111 relativeto +1 of P uscA and radiation causes P uscA activation thorough permitting the expression that is repressed under non-irradiated conditions

  13. Alternative staffing services. Contract transcription.

    Science.gov (United States)

    Tessier, C

    1992-03-01

    Contract medical transcription services can be of great assistance in meeting the demands for transcription, without jeopardizing patient, physician, or institutional confidentiality. You simply must require the contract service to provide at least the same degree of protection and preservation of confidentiality that you should require inhouse. To achieve this you must make these requirements explicit, comprehensive, comprehensible, believable, and enforceable. Discuss the requirements with prospective contractors. Review them at least annually with existing contractors and when contracts are due for renewal. Be sure to specify the consequence of breaching confidentiality, and if there are breaches, enforce the terms of the contract. Consult your institution's legal counsel both in developing the contract and in enforcing its provisions. Take into consideration your department's and institution's policies, AHIMA's statement on confidentiality, as well as local, state, and federal laws. Above all, never lose sight of the patient. Ultimately, it is not patient information that you are obligated to protect. It is the patient.

  14. The post-transcriptional operon

    DEFF Research Database (Denmark)

    Tenenbaum, Scott A.; Christiansen, Jan; Nielsen, Henrik

    2011-01-01

    model (PTO) is used to describe data from an assortment of methods (e.g. RIP-Chip, CLIP-Chip, miRNA profiling, ribosome profiling) that globally address the functionality of mRNA. Several examples of post-transcriptional operons have been documented in the literature and demonstrate the usefulness...... of the model in identifying new participants in cellular pathways as well as in deepening our understanding of cellular responses....

  15. Does the benefit on survival from leisure time physical activity depend on physical activity at work? A prospective cohort study.

    Directory of Open Access Journals (Sweden)

    Andreas Holtermann

    Full Text Available PURPOSE: To investigate if persons with high physical activity at work have the same benefits from leisure time physical activity as persons with sedentary work. METHODS: In the Copenhagen City Heart Study, a prospective cohort of 7,411 males and 8,916 females aged 25-66 years without known cardiovascular disease at entry in 1976-78, 1981-83, 1991-94, or 2001-03, the authors analyzed with sex-stratified multivariate Cox proportional hazards regression the association between leisure time physical activity and cardiovascular and all-cause mortality among individuals with different levels of occupational physical activity. RESULTS: During a median follow-up of 22.4 years, 4,003 individuals died from cardiovascular disease and 8,935 from all-causes. Irrespective of level of occupational physical activity, a consistently lower risk with increasing leisure time physical activity was found for both cardiovascular and all-cause mortality among both men and women. Compared to low leisure time physical activity, the survival benefit ranged from 1.5-3.6 years for moderate and 2.6-4.7 years for high leisure time physical activity among the different levels of occupational physical activity. CONCLUSION: Public campaigns and initiatives for increasing physical activity in the working population should target everybody, irrespective of physical activity at work.

  16. Does the benefit on survival from leisure time physical activity depend on physical activity at work? A prospective cohort study.

    Science.gov (United States)

    Holtermann, Andreas; Marott, Jacob Louis; Gyntelberg, Finn; Søgaard, Karen; Suadicani, Poul; Mortensen, Ole Steen; Prescott, Eva; Schnohr, Peter

    2013-01-01

    To investigate if persons with high physical activity at work have the same benefits from leisure time physical activity as persons with sedentary work. In the Copenhagen City Heart Study, a prospective cohort of 7,411 males and 8,916 females aged 25-66 years without known cardiovascular disease at entry in 1976-78, 1981-83, 1991-94, or 2001-03, the authors analyzed with sex-stratified multivariate Cox proportional hazards regression the association between leisure time physical activity and cardiovascular and all-cause mortality among individuals with different levels of occupational physical activity. During a median follow-up of 22.4 years, 4,003 individuals died from cardiovascular disease and 8,935 from all-causes. Irrespective of level of occupational physical activity, a consistently lower risk with increasing leisure time physical activity was found for both cardiovascular and all-cause mortality among both men and women. Compared to low leisure time physical activity, the survival benefit ranged from 1.5-3.6 years for moderate and 2.6-4.7 years for high leisure time physical activity among the different levels of occupational physical activity. Public campaigns and initiatives for increasing physical activity in the working population should target everybody, irrespective of physical activity at work.

  17. Mutual interdependence of splicing and transcription elongation.

    Science.gov (United States)

    Brzyżek, Grzegorz; Świeżewski, Szymon

    2015-01-01

    Transcription and splicing are intrinsically linked, as splicing needs a pre-mRNA substrate to commence. The more nuanced view is that the rate of transcription contributes to splicing regulation. On the other hand there is accumulating evidence that splicing has an active role in controlling transcription elongation by DNA-dependent RNA polymerase II (RNAP II). We briefly review those mechanisms and propose a unifying model where splicing controls transcription elongation to provide an optimal timing for successive rounds of splicing.

  18. Transcription of minute virus of mice, an autonomous parvovirus, may be regulated by attenuation

    International Nuclear Information System (INIS)

    Ben-Asher, E.; Aloni, Y.

    1984-01-01

    To characterize the transcriptional organization and regulation of minute virus of mice, an autonomous parvovirus, viral transcriptional complexes were isolated and cleaved with restriction enzymes. The in vivo preinitiated nascent RNA was elongated in vitro in the presence of [alpha- 32 P]UTP to generate runoff transcripts. The lengths of the runoff transcripts were analyzed by gel electrophoresis under denaturing conditions. On the basis of the map locations of the restriction sites and the lengths of the runoff transcripts, the in vivo initiation sites were determined. Two major initiation sites having similar activities were thus identified at residues 201 +/- 5 and 2005 +/- 5; both of them were preceded by a TATAA sequence. When uncleaved viral transcriptional complexes or isolated nuclei were incubated in vitro in the presence of [alpha- 32 P]UTP or [alpha- 32 P]CTP, they synthesized labeled RNA that, as determined by polyacrylamide gel electrophoresis, contained a major band of 142 nucleotides. The RNA of the major band was mapped between the initiation site at residue 201 +/- 5 and residue 342. We noticed the potential of forming two mutually exclusive stem-and-loop structures in the 142-nucleotide RNA; one of them is followed by a string of uridylic acid residues typical of a procaryotic transcription termination signal. We propose that, as in the transcription of simian virus 40, RNA transcription in minute virus of mice may be regulated by attenuation and may involve eucaryotic polymerase B, which can respond to a transcription termination signal similar to that of the procaryotic polymerase

  19. Dissection of TALE-dependent gene activation reveals that they induce transcription cooperatively and in both orientations.

    Science.gov (United States)

    Streubel, Jana; Baum, Heidi; Grau, Jan; Stuttman, Johannes; Boch, Jens

    2017-01-01

    Plant-pathogenic Xanthomonas bacteria inject transcription activator-like effector proteins (TALEs) into host cells to specifically induce transcription of plant genes and enhance susceptibility. Although the DNA-binding mode is well-understood it is still ambiguous how TALEs initiate transcription and whether additional promoter elements are needed to support this. To systematically dissect prerequisites for transcriptional initiation the activity of one TALE was compared on different synthetic Bs4 promoter fragments. In addition, a large collection of artificial TALEs spanning the OsSWEET14 promoter was compared. We show that the presence of a TALE alone is not sufficient to initiate transcription suggesting the requirement of additional supporting promoter elements. At the OsSWEET14 promoter TALEs can initiate transcription from various positions, in a synergistic manner of multiple TALEs binding in parallel to the promoter, and even by binding in reverse orientation. TALEs are known to shift the transcriptional start site, but our data show that this shift depends on the individual position of a TALE within a promoter context. Our results implicate that TALEs function like classical enhancer-binding proteins and initiate transcription in both orientations which has consequences for in planta target gene prediction and design of artificial activators.

  20. Interplay between DNA supercoiling and transcription elongation.

    Science.gov (United States)

    Ma, Jie; Wang, Michelle

    2014-01-01

    Transcription-coupled DNA supercoiling has been shown to be an important regulator of transcription that is broadly present in the cell. Here we review experimental work which shows that RNA polymerase is a powerful torsional motor that can alter DNA topology and structure, and DNA supercoiling in turn directly affects transcription elongation.

  1. The DNA replication checkpoint directly regulates MBF-dependent G1/S transcription.

    Science.gov (United States)

    Dutta, Chaitali; Patel, Prasanta K; Rosebrock, Adam; Oliva, Anna; Leatherwood, Janet; Rhind, Nicholas

    2008-10-01

    The DNA replication checkpoint transcriptionally upregulates genes that allow cells to adapt to and survive replication stress. Our results show that, in the fission yeast Schizosaccharomyces pombe, the replication checkpoint regulates the entire G(1)/S transcriptional program by directly regulating MBF, the G(1)/S transcription factor. Instead of initiating a checkpoint-specific transcriptional program, the replication checkpoint targets MBF to maintain the normal G(1)/S transcriptional program during replication stress. We propose a mechanism for this regulation, based on in vitro phosphorylation of the Cdc10 subunit of MBF by the Cds1 replication-checkpoint kinase. Replacement of two potential phosphorylation sites with phosphomimetic amino acids suffices to promote the checkpoint transcriptional program, suggesting that Cds1 phosphorylation directly regulates MBF-dependent transcription. The conservation of MBF between fission and budding yeast, and recent results implicating MBF as a target of the budding yeast replication checkpoint, suggests that checkpoint regulation of the MBF transcription factor is a conserved strategy for coping with replication stress. Furthermore, the structural and regulatory similarity between MBF and E2F, the metazoan G(1)/S transcription factor, suggests that this checkpoint mechanism may be broadly conserved among eukaryotes.

  2. Directing traffic on DNA-How transcription factors relieve or induce transcriptional interference.

    Science.gov (United States)

    Hao, Nan; Palmer, Adam C; Dodd, Ian B; Shearwin, Keith E

    2017-03-15

    Transcriptional interference (TI) is increasingly recognized as a widespread mechanism of gene control, particularly given the pervasive nature of transcription, both sense and antisense, across all kingdoms of life. Here, we discuss how transcription factor binding kinetics strongly influence the ability of a transcription factor to relieve or induce TI.

  3. Insights into mRNP biogenesis provided by new genetic interactions among export and transcription factors

    Directory of Open Access Journals (Sweden)

    Estruch Francisco

    2012-09-01

    Full Text Available Abstract Background The various steps of mRNP biogenesis (transcription, processing and export are interconnected. It has been shown that the transcription machinery plays a pivotal role in mRNP assembly, since several mRNA export factors are recruited during transcription and physically interact with components of the transcription machinery. Although the shuttling DEAD-box protein Dbp5p is concentrated on the cytoplasmic fibrils of the NPC, previous studies demonstrated that it interacts physically and genetically with factors involved in transcription initiation. Results We investigated the effect of mutations affecting various components of the transcription initiation apparatus on the phenotypes of mRNA export mutant strains. Our results show that growth and mRNA export defects of dbp5 and mex67 mutant strains can be suppressed by mutation of specific transcription initiation components, but suppression was not observed for mutants acting in the very first steps of the pre-initiation complex (PIC formation. Conclusions Our results indicate that mere reduction in the amount of mRNP produced is not sufficient to suppress the defects caused by a defective mRNA export factor. Suppression occurs only with mutants affecting events within a narrow window of the mRNP biogenesis process. We propose that reducing the speed with which transcription converts from initiation and promoter clearance to elongation may have a positive effect on mRNP formation by permitting more effective recruitment of partially-functional mRNP proteins to the nascent mRNP.

  4. The Journey of a Transcription Factor

    DEFF Research Database (Denmark)

    Pireyre, Marie

    Plants have developed astonishing networks regulating their metabolism to adapt to their environment. The complexity of these networks is illustrated by the expansion of families of regulators such as transcription factors in the plant kingdom. Transcription factors specifically impact...... transcriptional networks by integrating exogenous and endogenous stimuli and regulating gene expression accordingly. Regulation of transcription factors and their activation is thus highly important to modulate the transcriptional programs and increase fitness of the plant in a given environment. Plant metabolism....... The biosynthetic machinery of GLS is governed by interplay of six MYB and three bHLH transcription factors. MYB28, MYB29 and MYB76 regulate methionine-derived GLS, and MYB51, MYB34 and MYB122 regulate tryptophan-derived GLS. The three bHLH transcription factors MYC2, MYC3 and MYC4 physically interact with all six...

  5. Repression of Meiotic Genes by Antisense Transcription and by Fkh2 Transcription Factor in Schizosaccharomyces pombe

    OpenAIRE

    Chen, Huei-Mei; Rosebrock, Adam P.; Khan, Sohail R.; Futcher, Bruce; Leatherwood, Janet K.

    2012-01-01

    In S. pombe, about 5% of genes are meiosis-specific and accumulate little or no mRNA during vegetative growth. Here we use Affymetrix tiling arrays to characterize transcripts in vegetative and meiotic cells. In vegetative cells, many meiotic genes, especially those induced in mid-meiosis, have abundant antisense transcripts. Disruption of the antisense transcription of three of these mid-meiotic genes allowed vegetative sense transcription. These results suggest that antisense transcription ...

  6. Initiation devices, initiation systems including initiation devices and related methods

    Energy Technology Data Exchange (ETDEWEB)

    Daniels, Michael A.; Condit, Reston A.; Rasmussen, Nikki; Wallace, Ronald S.

    2018-04-10

    Initiation devices may include at least one substrate, an initiation element positioned on a first side of the at least one substrate, and a spark gap electrically coupled to the initiation element and positioned on a second side of the at least one substrate. Initiation devices may include a plurality of substrates where at least one substrate of the plurality of substrates is electrically connected to at least one adjacent substrate of the plurality of substrates with at least one via extending through the at least one substrate. Initiation systems may include such initiation devices. Methods of igniting energetic materials include passing a current through a spark gap formed on at least one substrate of the initiation device, passing the current through at least one via formed through the at least one substrate, and passing the current through an explosive bridge wire of the initiation device.

  7. DNA Topoisomerases Maintain Promoters in a State Competent for Transcriptional Activation in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Pedersen, Jakob Madsen; Fredsøe, Jacob Christian; Rødgaard, Morten Terpager

    2012-01-01

    To investigate the role of DNA topoisomerases in transcription, we have studied global gene expression in Saccharomyces cerevisiae cells deficient for topoisomerases I and II and performed single-gene analyses to support our findings. The genome-wide studies show a general transcriptional down......-regulation upon lack of the enzymes, which correlates with gene activity but not gene length. Furthermore, our data reveal a distinct subclass of genes with a strong requirement for topoisomerases. These genes are characterized by high transcriptional plasticity, chromatin regulation, TATA box presence......-depth analysis of the inducible PHO5 gene reveals that topoisomerases are essential for binding of the Pho4p transcription factor to the PHO5 promoter, which is required for promoter nucleosome removal during activation. In contrast, topoisomerases are dispensable for constitutive transcription initiation...

  8. Regulation of transcription of cellulases- and hemicellulases-encoding genes in Aspergillus niger and Hypocrea jecorina (Trichoderma reesei)

    NARCIS (Netherlands)

    Stricker, A.R.; Mach, R.L.; Graaff, de L.H.

    2008-01-01

    The filamentous fungi Aspergillus niger and Hypocrea jecorina (Trichoderma reesei) have been the subject of many studies investigating the mechanism of transcriptional regulation of hemicellulase- and cellulase-encoding genes. The transcriptional regulator XlnR that was initially identified in A.

  9. Activity-dependent shedding of the NMDA receptor glycine binding site by matrix metalloproteinase 3: a PUTATIVE mechanism of postsynaptic plasticity.

    Directory of Open Access Journals (Sweden)

    Thorsten Pauly

    Full Text Available Functional and structural alterations of clustered postsynaptic ligand gated ion channels in neuronal cells are thought to contribute to synaptic plasticity and memory formation in the human brain. Here, we describe a novel molecular mechanism for structural alterations of NR1 subunits of the NMDA receptor. In cultured rat spinal cord neurons, chronic NMDA receptor stimulation induces disappearance of extracellular epitopes of NMDA receptor NR1 subunits, which was prevented by inhibiting matrix metalloproteinases (MMPs. Immunoblotting revealed the digestion of solubilized NR1 subunits by MMP-3 and identified a fragment of about 60 kDa as MMPs-activity-dependent cleavage product of the NR1 subunit in cultured neurons. The expression of MMP-3 in the spinal cord culture was shown by immunoblotting and immunofluorescence microscopy. Recombinant NR1 glycine binding protein was used to identify MMP-3 cleavage sites within the extracellular S1 and S2-domains. N-terminal sequencing and site-directed mutagenesis revealed S542 and L790 as two putative major MMP-3 cleavage sites of the NR1 subunit. In conclusion, our data indicate that MMPs, and in particular MMP-3, are involved in the activity dependent alteration of NMDA receptor structure at postsynaptic membrane specializations in the CNS.

  10. Activity-dependent shedding of the NMDA receptor glycine binding site by matrix metalloproteinase 3: a PUTATIVE mechanism of postsynaptic plasticity.

    Science.gov (United States)

    Pauly, Thorsten; Ratliff, Miriam; Pietrowski, Eweline; Neugebauer, Rainer; Schlicksupp, Andrea; Kirsch, Joachim; Kuhse, Jochen

    2008-07-16

    Functional and structural alterations of clustered postsynaptic ligand gated ion channels in neuronal cells are thought to contribute to synaptic plasticity and memory formation in the human brain. Here, we describe a novel molecular mechanism for structural alterations of NR1 subunits of the NMDA receptor. In cultured rat spinal cord neurons, chronic NMDA receptor stimulation induces disappearance of extracellular epitopes of NMDA receptor NR1 subunits, which was prevented by inhibiting matrix metalloproteinases (MMPs). Immunoblotting revealed the digestion of solubilized NR1 subunits by MMP-3 and identified a fragment of about 60 kDa as MMPs-activity-dependent cleavage product of the NR1 subunit in cultured neurons. The expression of MMP-3 in the spinal cord culture was shown by immunoblotting and immunofluorescence microscopy. Recombinant NR1 glycine binding protein was used to identify MMP-3 cleavage sites within the extracellular S1 and S2-domains. N-terminal sequencing and site-directed mutagenesis revealed S542 and L790 as two putative major MMP-3 cleavage sites of the NR1 subunit. In conclusion, our data indicate that MMPs, and in particular MMP-3, are involved in the activity dependent alteration of NMDA receptor structure at postsynaptic membrane specializations in the CNS.

  11. Replicative Stress Induces Intragenic Transcription of the ASE1 Gene that Negatively Regulates Ase1 Activity

    OpenAIRE

    McKnight, Kelly; Liu, Hong; Wang, Yanchang

    2014-01-01

    Intragenic transcripts initiate within the coding region of a gene, thereby producing shorter mRNAs and proteins. Although intragenic transcripts are widely expressed [1], their role in the functional regulation of genes remains largely unknown. In budding yeast, DNA replication stress activates the S-phase checkpoint that stabilizes replication forks and arrests cells in S-phase with a short spindle [2-4]. When yeast cells were treated with hydroxyurea (HU) to block DNA synthesis and induce ...

  12. Potential role of Arabidopsis PHP as an accessory subunit of the PAF1 transcriptional cofactor.

    Science.gov (United States)

    Park, Sunchung; Ek-Ramos, Maria Julissa; Oh, Sookyung; van Nocker, Steven

    2011-08-01

    Paf1C is a transcriptional cofactor that has been implicated in various transcription-associated mechanisms spanning initiation, elongation and RNA processing, and is important for multiple aspects of development in Arabidopsis. Our recent studies suggest Arabidopsis Paf1C is crucial for proper regulation of genes within H3K27me3-enriched chromatin, and that a protein named PHP may act as an accessory subunit of Paf1C that promotes this function.

  13. Mitotic Transcriptional Activation: Clearance of Actively Engaged Pol II via Transcriptional Elongation Control in Mitosis.

    Science.gov (United States)

    Liang, Kaiwei; Woodfin, Ashley R; Slaughter, Brian D; Unruh, Jay R; Box, Andrew C; Rickels, Ryan A; Gao, Xin; Haug, Jeffrey S; Jaspersen, Sue L; Shilatifard, Ali

    2015-11-05

    Although it is established that some general transcription factors are inactivated at mitosis, many details of mitotic transcription inhibition (MTI) and its underlying mechanisms are largely unknown. We have identified mitotic transcriptional activation (MTA) as a key regulatory step to control transcription in mitosis for genes with transcriptionally engaged RNA polymerase II (Pol II) to activate and transcribe until the end of the gene to clear Pol II from mitotic chromatin, followed by global impairment of transcription reinitiation through MTI. Global nascent RNA sequencing and RNA fluorescence in situ hybridization demonstrate the existence of transcriptionally engaged Pol II in early mitosis. Both genetic and chemical inhibition of P-TEFb in mitosis lead to delays in the progression of cell division. Together, our study reveals a mechanism for MTA and MTI whereby transcriptionally engaged Pol II can progress into productive elongation and finish transcription to allow proper cellular division. Copyright © 2015 Elsevier Inc. All rights reserved.

  14. Effect of uv irradiation on lambda DNA transcription

    Energy Technology Data Exchange (ETDEWEB)

    Ranade, S S [Cancer Research Inst., Bombay (India)

    1977-05-01

    The effect of uv irradiation of template DNA has been studied in vitro in the E.coli RNA polymerase system with native and uv treated lambda DNA. Lambda DNA was more susceptible to uv than was calf-thymus DNA, yet a residual activity was observed at a uv dose of 0.5 x 10/sup 4/ erg/mm/sup 2/. From the kinetic analysis of the reaction and the incorporation of lambda /sup 32/P-labelled nucleoside triphosphates, it seems reasonable to conclude that uv irradiation probably did not affect the DNA initiation sites, recognizable by RNA polymerase. The transcription products made with uv irradiated lambda DNA were asymmetrical, and hybridized to the right half (R) and the left half (L) of lambda DNA with the ratio of R/L=4/1, and they showed a lower hybridizability than the transcripts with native lambda DNA. The initiation sites recognizable by RNA polymerase seemed to be the same on both native and uv irradiated lambda DNA, though the transcription of uv treated lambda DNA appeared to terminate with rather short RNA chains.

  15. RNA Pol II promotes transcription of centromeric satellite DNA in beetles.

    Directory of Open Access Journals (Sweden)

    Zeljka Pezer

    Full Text Available Transcripts of centromeric satellite DNAs are known to play a role in heterochromatin formation as well as in establishment of the kinetochore. However, little is known about basic mechanisms of satellite DNA expression within constitutive heterochromatin and its regulation. Here we present comprehensive analysis of transcription of abundant centromeric satellite DNA, PRAT from beetle Palorus ratzeburgii (Coleoptera. This satellite is characterized by preservation and extreme sequence conservation among evolutionarily distant insect species. PRAT is expressed in all three developmental stages: larvae, pupae and adults at similar level. Transcripts are abundant comprising 0.033% of total RNA and are heterogeneous in size ranging from 0.5 kb up to more than 5 kb. Transcription proceeds from both strands but with 10 fold different expression intensity and transcripts are not processed into siRNAs. Most of the transcripts (80% are not polyadenylated and remain in the nucleus while a small portion is exported to the cytoplasm. Multiple, irregularly distributed transcription initiation sites as well as termination sites have been mapped within the PRAT sequence using primer extension and RLM-RACE. The presence of cap structure as well as poly(A tails in a portion of the transcripts indicate RNA polymerase II-dependent transcription and a putative polymerase II promoter site overlaps the most conserved part of the PRAT sequence. The treatment of larvae with alpha-amanitin decreases the level of PRAT transcripts at concentrations that selectively inhibit pol II activity. In conclusion, stable, RNA polymerase II dependant transcripts of abundant centromeric satellite DNA, not regulated by RNAi, have been identified and characterized. This study offers a basic understanding of expression of highly abundant heterochromatic DNA which in beetle species constitutes up to 50% of the genome.

  16. Discriminative identification of transcriptional responses of promoters and enhancers after stimulus

    KAUST Repository

    Kleftogiannis, Dimitrios A.

    2016-10-17

    Promoters and enhancers regulate the initiation of gene expression and maintenance of expression levels in spatial and temporal manner. Recent findings stemming from the Cap Analysis of Gene Expression (CAGE) demonstrate that promoters and enhancers, based on their expression profiles after stimulus, belong to different transcription response subclasses. One of the most promising biological features that might explain the difference in transcriptional response between subclasses is the local chromatin environment. We introduce a novel computational framework, PEDAL, for distinguishing effectively transcriptional profiles of promoters and enhancers using solely histone modification marks, chromatin accessibility and binding sites of transcription factors and co-activators. A case study on data from MCF-7 cell-line reveals that PEDAL can identify successfully the transcription response subclasses of promoters and enhancers from two different stimulations. Moreover, we report subsets of input markers that discriminate with minimized classification error MCF-7 promoter and enhancer transcription response subclasses. Our work provides a general computational approach for identifying effectively cell-specific and stimulation-specific promoter and enhancer transcriptional profiles, and thus, contributes to improve our understanding of transcriptional activation in human.

  17. Promoter proximal polyadenylation sites reduce transcription activity

    DEFF Research Database (Denmark)

    Andersen, Pia Kjølhede; Lykke-Andersen, Søren; Jensen, Torben Heick

    2012-01-01

    Gene expression relies on the functional communication between mRNA processing and transcription. We previously described the negative impact of a point-mutated splice donor (SD) site on transcription. Here we demonstrate that this mutation activates an upstream cryptic polyadenylation (CpA) site......, which in turn causes reduced transcription. Functional depletion of U1 snRNP in the context of the wild-type SD triggers the same CpA event accompanied by decreased RNA levels. Thus, in accordance with recent findings, U1 snRNP can shield premature pA sites. The negative impact of unshielded pA sites...... on transcription requires promoter proximity, as demonstrated using artificial constructs and supported by a genome-wide data set. Importantly, transcription down-regulation can be recapitulated in a gene context devoid of splice sites by placing a functional bona fide pA site/transcription terminator within ∼500...

  18. Transcription and recombination: when RNA meets DNA.

    Science.gov (United States)

    Aguilera, Andrés; Gaillard, Hélène

    2014-08-01

    A particularly relevant phenomenon in cell physiology and proliferation is the fact that spontaneous mitotic recombination is strongly enhanced by transcription. The most accepted view is that transcription increases the occurrence of double-strand breaks and/or single-stranded DNA gaps that are repaired by recombination. Most breaks would arise as a consequence of the impact that transcription has on replication fork progression, provoking its stalling and/or breakage. Here, we discuss the mechanisms responsible for the cross talk between transcription and recombination, with emphasis on (1) the transcription-replication conflicts as the main source of recombinogenic DNA breaks, and (2) the formation of cotranscriptional R-loops as a major cause of such breaks. The new emerging questions and perspectives are discussed on the basis of the interference between transcription and replication, as well as the way RNA influences genome dynamics. Copyright © 2014 Cold Spring Harbor Laboratory Press; all rights reserved.

  19. Specificity and robustness in transcription control networks.

    Science.gov (United States)

    Sengupta, Anirvan M; Djordjevic, Marko; Shraiman, Boris I

    2002-02-19

    Recognition by transcription factors of the regulatory DNA elements upstream of genes is the fundamental step in controlling gene expression. How does the necessity to provide stability with respect to mutation constrain the organization of transcription control networks? We examine the mutation load of a transcription factor interacting with a set of n regulatory response elements as a function of the factor/DNA binding specificity and conclude on theoretical grounds that the optimal specificity decreases with n. The predicted correlation between variability of binding sites (for a given transcription factor) and their number is supported by the genomic data for Escherichia coli. The analysis of E. coli genomic data was carried out using an algorithm suggested by the biophysical model of transcription factor/DNA binding. Complete results of the search for candidate transcription factor binding sites are available at http://www.physics.rockefeller.edu/~boris/public/search_ecoli.

  20. Transcriptional analysis of apple fruit proanthocyanidin biosynthesis

    Science.gov (United States)

    Henry-Kirk, Rebecca A.

    2012-01-01

    Proanthocyanidins (PAs) are products of the flavonoid pathway, which also leads to the production of anthocyanins and flavonols. Many flavonoids have antioxidant properties and may have beneficial effects for human health. PAs are found in the seeds and fruits of many plants. In apple fruit (Malus × domestica Borkh.), the flavonoid biosynthetic pathway is most active in the skin, with the flavan-3-ols, catechin, and epicatechin acting as the initiating units for the synthesis of PA polymers. This study examined the genes involved in the production of PAs in three apple cultivars: two heritage apple cultivars, Hetlina and Devonshire Quarrenden, and a commercial cultivar, Royal Gala. HPLC analysis shows that tree-ripe fruit from Hetlina and Devonshire Quarrenden had a higher phenolic content than Royal Gala. Epicatechin and catechin biosynthesis is under the control of the biosynthetic enzymes anthocyanidin reductase (ANR) and leucoanthocyanidin reductase (LAR1), respectively. Counter-intuitively, real-time quantitative PCR analysis showed that the expression levels of Royal Gala LAR1 and ANR were significantly higher than those of both Devonshire Quarrenden and Hetlina. This suggests that a compensatory feedback mechanism may be active, whereby low concentrations of PAs may induce higher expression of gene transcripts. Further investigation is required into the regulation of these key enzymes in apple. Abbreviations:ANOVAanalysis of varianceANRanthocyanidin reductaseDADdiode array detectorDAFBdays after full bloomDFRdihydroflavonol reductaseLARleucoanthocyanidin reductaseLC-MSliquid chromatography/mass spectrometryPAproanthocyanidinqPCRreal-time quantitative PCR PMID:22859681

  1. Coordinating repair of oxidative DNA damage with transcription and replication

    International Nuclear Information System (INIS)

    Cooper, P.K.

    2003-01-01

    Transcription-coupled repair (TCR) preferentially removes DNA lesions from template strands of active genes. Defects in TCR, which acts both on lesions removed by nucleotide excision repair (NER) and on oxidative lesions removed by base excision repair (BER), underlie the fatal developmental disorder Cockayne syndrome. Although its detailed mechanism remains unknown, TCR involves recognition of a stalled RNA polymerase (RNAP), removal or remodeling of RNAP to allow access to the lesion, and recruitment of repair enzymes. At a minimum, these early steps require a non-enzymatic function of the multifunctional repair protein XPG, the CSB protein with ATP-dependent chromatin remodeling activity, and the TFIIH complex (including the XPB and XPD helicases) that is also required for basal transcription initiation and NER. XPG exists in the cell in a complex with TFIIH, and in vitro evidence has suggested that it interacts with CSB. To address the mechanism of TCR, we are characterizing protein-DNA and protein-protein interactions of XPG. We show that XPG preferentially binds to double-stranded DNA containing bubbles resembling in size the unpaired regions associated with transcription. Two distinct domains of XPG are required for the observed strong binding specificity and stability. XPG both interacts directly with CSB and synergistically binds with it to bubble DNA, and it strongly stimulates the bubble DNA-dependent ATPase activity of CSB. Significantly for TCR, XPG also interacts directly with RNAP II, binds both the protein and nucleic acid components (the R-loop) of a stalled RNA polymerase, and forms a ternary complex with CSB and the stalled RNAP. These results are consistent with the model that XPG and CSB jointly interact with the DNA/chromatin structure in the vicinity of the stalled transcriptional apparatus and with the transcriptional machinery itself to remodel the chromatin and either move or remodel the blocked RNA polymerase to expose the lesion

  2. Transcriptional networks and chromatin remodeling controlling adipogenesis

    DEFF Research Database (Denmark)

    Siersbæk, Rasmus; Nielsen, Ronni; Mandrup, Susanne

    2012-01-01

    Adipocyte differentiation is tightly controlled by a transcriptional cascade, which directs the extensive reprogramming of gene expression required to convert fibroblast-like precursor cells into mature lipid-laden adipocytes. Recent global analyses of transcription factor binding and chromatin...... remodeling have revealed 'snapshots' of this cascade and the chromatin landscape at specific time-points of differentiation. These studies demonstrate that multiple adipogenic transcription factors co-occupy hotspots characterized by an open chromatin structure and specific epigenetic modifications....... Such transcription factor hotspots are likely to represent key signaling nodes which integrate multiple adipogenic signals at specific chromatin sites, thereby facilitating coordinated action on gene expression....

  3. Targeted deficiency of the transcriptional activator Hnf1alpha alters subnuclear positioning of its genomic targets.

    Directory of Open Access Journals (Sweden)

    Reini F Luco

    2008-05-01

    Full Text Available DNA binding transcriptional activators play a central role in gene-selective regulation. In part, this is mediated by targeting local covalent modifications of histone tails. Transcriptional regulation has also been associated with the positioning of genes within the nucleus. We have now examined the role of a transcriptional activator in regulating the positioning of target genes. This was carried out with primary beta-cells and hepatocytes freshly isolated from mice lacking Hnf1alpha, an activator encoded by the most frequently mutated gene in human monogenic diabetes (MODY3. We show that in Hnf1a-/- cells inactive endogenous Hnf1alpha-target genes exhibit increased trimethylated histone H3-Lys27 and reduced methylated H3-Lys4. Inactive Hnf1alpha-targets in Hnf1a-/- cells are also preferentially located in peripheral subnuclear domains enriched in trimethylated H3-Lys27, whereas active targets in wild-type cells are positioned in more central domains enriched in methylated H3-Lys4 and RNA polymerase II. We demonstrate that this differential positioning involves the decondensation of target chromatin, and show that it is spatially restricted rather than a reflection of non-specific changes in the nuclear organization of Hnf1a-deficient cells. This study, therefore, provides genetic evidence that a single transcriptional activator can influence the subnuclear location of its endogenous genomic targets in primary cells, and links activator-dependent changes in local chromatin structure to the spatial organization of the genome. We have also revealed a defect in subnuclear gene positioning in a model of a human transcription factor disease.

  4. Transcription-based model for the induction of chromosomal exchange events by ionising radiation

    International Nuclear Information System (INIS)

    Radford, I.A.

    2003-01-01

    The mechanistic basis for chromosomal aberration formation, following exposure of mammalian cells to ionising radiation, has long been debated. Although chromosomal aberrations are probably initiated by DNA double-strand breaks (DSB), little is understood about the mechanisms that generate and modulate DNA rearrangement. Based on results from our laboratory and data from the literature, a novel model of chromosomal aberration formation has been suggested (Radford 2002). The basic postulates of this model are that: (1) DSB, primarily those involving multiple individual damage sites (i.e. complex DSB), are the critical initiating lesion; (2) only those DSB occurring in transcription units that are associated with transcription 'factories' (complexes containing multiple transcription units) induce chromosomal exchange events; (3) such DSB are brought into contact with a DNA topoisomerase I molecule through RNA polymerase II catalysed transcription and give rise to trapped DNA-topo I cleavage complexes; and (4) trapped complexes interact with another topo I molecule on a temporarily inactive transcription unit at the same transcription factory leading to DNA cleavage and subsequent strand exchange between the cleavage complexes. We have developed a method using inverse PCR that allows the detection and sequencing of putative ionising radiation-induced DNA rearrangements involving different regions of the human genome (Forrester and Radford 1998). The sequences detected by inverse PCR can provide a test of the prediction of the transcription-based model that ionising radiation-induced DNA rearrangements occur between sequences in active transcription units. Accordingly, reverse transcriptase PCR was used to determine if sequences involved in rearrangements were transcribed in the test cells. Consistent with the transcription-based model, nearly all of the sequences examined gave a positive result to reverse transcriptase PCR (Forrester and Radford unpublished)

  5. The functionalized amino acid (S-Lacosamide subverts CRMP2-mediated tubulin polymerization to prevent constitutive and activity-dependent increase in neurite outgrowth

    Directory of Open Access Journals (Sweden)

    Sarah M Wilson

    2014-07-01

    Full Text Available Activity-dependent neurite outgrowth is a highly complex, regulated process with important implications for neuronal circuit remodeling in development as well as in seizure-induced sprouting in epilepsy. Recent work has linked outgrowth to collapsin response mediator protein 2 (CRMP2, an intracellular phosphoprotein originally identified as axon guidance and growth cone collapse protein. The neurite outgrowth promoting function of CRMP2 is regulated by its phosphorylation state. In this study, depolarization (potassium chloride-driven activity increased the level of active CRMP2 by decreasing its phosphorylation by GSK3β via a reduction in priming by Cdk5. To determine the contribution of CRMP2 in activity-driven neurite outgrowth, we screened a limited set of compounds for their ability to reduce neurite outgrowth but not modify voltage-gated sodium channel (VGSC biophysical properties. This led to the identification of (S-lacosamide ((S-LCM, a stereoisomer of the clinically used antiepileptic drug (R-LCM (Vimpat®, as a novel tool for preferentially targeting CRMP2-mediated neurite outgrowth. Whereas (S-LCM was ineffective in targeting VGSCs, the presumptive pharmacological targets of (R-LCM, (S-LCM was more efficient than (R-LCM in subverting neurite outgrowth. Biomolecular interaction analyses revealed that (S-LCM bound to wildtype CRMP2 with low micromolar affinity, similar to (R-LCM. Through the use of this novel tool, the activity-dependent increase in neurite outgrowth observed following depolarization was characterized to be reliant on CRMP2 function. Knockdown of CRMP2 by siRNA in cortical neurons resulted in reduced CRMP2-dependent neurite outgrowth; incubation with (S-LCM phenocopied this effect. Other CRMP2-mediated processes were unaffected. (S-LCM subverted neurite outgrowth not by affecting the canonical CRMP2-tubulin association but rather by impairing the ability of CRMP2 to promote tubulin polymerization, events that are

  6. Transcriptional repression of BODENLOS by HD-ZIP transcription factor HB5 in Arabidopsis thaliana.

    NARCIS (Netherlands)

    Smet, De I.; Lau, S.; Ehrismann, J.S.; Axiotis, I.; Kolb, M.; Kientz, M.; Weijers, D.; Jürgens, G.

    2013-01-01

    In Arabidopsis thaliana, the phytohormone auxin is an important patterning agent during embryogenesis and post-embryonic development, exerting effects through transcriptional regulation. The main determinants of the transcriptional auxin response machinery are AUXIN RESPONSE FACTOR (ARF)

  7. A long HBV transcript encoding pX is inefficiently exported from the nucleus

    International Nuclear Information System (INIS)

    Doitsh, Gilad; Shaul, Yosef

    2003-01-01

    The longest hepatitis B virus transcript is a 3.9-kb mRNA whose function remained unclear. In this study, we wished to identify the translation products and physiological role of this viral transcript. This transcript initiates from the X promoter region ignoring the inefficient and noncanonical viral polyadenylation signal at the first round of transcription. However, an HBV mutant with canonical polyadenylation signal continues, though with lower efficiency, to program the synthesis of this long transcript, indicating that the deviated HBV polyadenylation signal is important but not essential to enable transcription of the 3.9-kb species. The 3.9-kb RNA contains two times the X open reading frame (ORF). The X ORF at the 5'-end is positioned upstream of the CORE gene. By generating an HBV DNA mutant in which the X and Core ORFs are fused, we demonstrated the production of a 40-kDa X-Core fusion protein that must be encoded by the 3.9-kb transcript. Mutagenesis studies revealed that the production of this protein depends on the 5' X ORF ATG, suggesting that the 3.9-kb RNA is active in translation of the X ORF. Based on these features, the 3.9-kb transcript was designated lxRNA for long X RNA. Unlike other HBV transcripts, lxRNA harbors two copies of PRE, the posttranscriptional regulatory element that controls the nuclear export of HBV mRNAs. Unexpectedly, despite the presence of PRE sequences, RNA fractionation analysis revealed that lxRNA barely accumulates in the cytoplasm, suggesting that nuclear export of lxRNA is poor. Collectively, our data suggest that two distinct HBV mRNA species encode pX and that the HBV transcripts are differentially regulated at the level of nuclear export

  8. Overlapping transcription structure of human cytomegalovirus

    Indian Academy of Sciences (India)

    Transcription of human cytomegalovirus UL/b′ region has been studied extensively for some genes. In this study, transcripts of the UL140 and UL141, two of the UL/b′ genes, were identified in late RNAs of three HCMV isolates using Northern blot hybridization, cDNA library screening and RACE-PCR. At least three ...

  9. Overlapping transcription structure of human cytomegalovirus ...

    Indian Academy of Sciences (India)

    2013-01-21

    Jan 21, 2013 ... Transcription of human cytomegalovirus UL/b′ region has been studied extensively for some genes. In this study, transcripts of the UL140 and UL141, two of the UL/b′ genes, were identified in late RNAs of three HCMV isolates using Northern blot hybridization, cDNA library screening and RACE-PCR.

  10. Transcription of Byzantine Chant - Problems, Possibilities, Formats

    DEFF Research Database (Denmark)

    Troelsgård, Christian

    2007-01-01

    Discusses the problems and possibilities for transsription of Byzantine chant on the basis of medieval musical manuscripts. A relatively 'neutral' style of transcription is suggested for musicological purposes.......Discusses the problems and possibilities for transsription of Byzantine chant on the basis of medieval musical manuscripts. A relatively 'neutral' style of transcription is suggested for musicological purposes....

  11. Regulation of transcription in hyperthermophilic archaea

    NARCIS (Netherlands)

    Brinkman, A.B.

    2002-01-01

    The aim of the research presented here was to insight in the mechanisms by which transcription in hyperthermophilic archaea is regulated. To accomplish this, we have aimed (I) to identify transcriptional regulatory proteins from hyperthermophilic archaea, (II) to characterize these

  12. 45 CFR 99.27 - Official transcript.

    Science.gov (United States)

    2010-10-01

    ... 45 Public Welfare 1 2010-10-01 2010-10-01 false Official transcript. 99.27 Section 99.27 Public Welfare DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL ADMINISTRATION PROCEDURE FOR HEARINGS FOR THE CHILD CARE AND DEVELOPMENT FUND Hearing Procedures § 99.27 Official transcript. The Department will...

  13. RNA polymerase II collision interrupts convergent transcription

    DEFF Research Database (Denmark)

    Hobson, David J; Wei, Wu; Steinmetz, Lars M

    2012-01-01

    Antisense noncoding transcripts, genes-within-genes, and convergent gene pairs are prevalent among eukaryotes. The existence of such transcription units raises the question of what happens when RNA polymerase II (RNAPII) molecules collide head-to-head. Here we use a combination of biochemical...

  14. TcoF-DB v2: update of the database of human and mouse transcription co-factors and transcription factor interactions

    KAUST Repository

    Schmeier, Sebastian

    2016-10-17

    Transcription factors (TFs) play a pivotal role in transcriptional regulation, making them crucial for cell survival and important biological functions. For the regulation of transcription, interactions of different regulatory proteins known as transcription co-factors (TcoFs) and TFs are essential in forming necessary protein complexes. Although TcoFs themselves do not bind DNA directly, their influence on transcriptional regulation and initiation, although indirect, has been shown to be significant, with the functionality of TFs strongly influenced by the presence of TcoFs. In the TcoF-DB v2 database, we collect information on TcoFs. In this article, we describe updates and improvements implemented in TcoF-DB v2. TcoF-DB v2 provides several new features that enables exploration of the roles of TcoFs. The content of the database has significantly expanded, and is enriched with information from Gene Ontology, biological pathways, diseases and molecular signatures. TcoF-DB v2 now includes many more TFs; has substantially increased the number of human TcoFs to 958, and now includes information on mouse (418 new TcoFs). TcoF-DB v2 enables the exploration of information on TcoFs and allows investigations into their influence on transcriptional regulation in humans and mice. TcoF-DB v2 can be accessed at http://tcofdb.org/.

  15. TcoF-DB v2: update of the database of human and mouse transcription co-factors and transcription factor interactions

    KAUST Repository

    Schmeier, Sebastian; Alam, Tanvir; Essack, Magbubah; Bajic, Vladimir B.

    2016-01-01

    Transcription factors (TFs) play a pivotal role in transcriptional regulation, making them crucial for cell survival and important biological functions. For the regulation of transcription, interactions of different regulatory proteins known as transcription co-factors (TcoFs) and TFs are essential in forming necessary protein complexes. Although TcoFs themselves do not bind DNA directly, their influence on transcriptional regulation and initiation, although indirect, has been shown to be significant, with the functionality of TFs strongly influenced by the presence of TcoFs. In the TcoF-DB v2 database, we collect information on TcoFs. In this article, we describe updates and improvements implemented in TcoF-DB v2. TcoF-DB v2 provides several new features that enables exploration of the roles of TcoFs. The content of the database has significantly expanded, and is enriched with information from Gene Ontology, biological pathways, diseases and molecular signatures. TcoF-DB v2 now includes many more TFs; has substantially increased the number of human TcoFs to 958, and now includes information on mouse (418 new TcoFs). TcoF-DB v2 enables the exploration of information on TcoFs and allows investigations into their influence on transcriptional regulation in humans and mice. TcoF-DB v2 can be accessed at http://tcofdb.org/.

  16. Mapping the transcription termination region of the mouse immunoglobulin kappa gene

    International Nuclear Information System (INIS)

    Xu, M.; Garrard, W.T.

    1986-01-01

    To define the transcription termination region of the mouse immunoglobulin kappa gene, they have subcloned single copy DNA sequences corresponding to both the template and the non-template strands of this locus. In vitro nuclear transcription with isolated MPC-11 nuclei was performed and the resulting 32 P-labeled RNA was hybridized to slot-blotted, single-stranded M13 probes covering regions within and flanking the kappa gene. The hybridization pattern for the template-strand reveals that transcription terminates within the region between 1.1 to 2.3 kb downstream from the poly(A) site. Ten different short sequences (8-13 bp) reside within 460 bp of this region that exhibit homology with sequences found in the termination regions of mouse β-globin and chicken ovalbumin genes. Transcription of the non-template strand occurs on either side of this termination region. They note that no transcription is detectable on the non-template strand downstream of the enhancer, indicating that if RNA polymerase II enters at this site, it does not initiate transcription during transit to the promoter region. They conclude that transcription of the kappa gene passes the poly(A) addition site and terminates within 2.3 Kb downstream

  17. Recent Advancements in DNA Damage-Transcription Crosstalk and High-Resolution Mapping of DNA Breaks.

    Science.gov (United States)

    Vitelli, Valerio; Galbiati, Alessandro; Iannelli, Fabio; Pessina, Fabio; Sharma, Sheetal; d'Adda di Fagagna, Fabrizio

    2017-08-31

    Until recently, DNA damage arising from physiological DNA metabolism was considered a detrimental by-product for cells. However, an increasing amount of evidence has shown that DNA damage could have a positive role in transcription activation. In particular, DNA damage has been detected in transcriptional elements following different stimuli. These physiological DNA breaks are thought to be instrumental for the correct expression of genomic loci through different mechanisms. In this regard, although a plethora of methods are available to precisely map transcribed regions and transcription start sites, commonly used techniques for mapping DNA breaks lack sufficient resolution and sensitivity to draw a robust correlation between DNA damage generation and transcription. Recently, however, several methods have been developed to map DNA damage at single-nucleotide resolution, thus providing a new set of tools to correlate DNA damage and transcription. Here, we review how DNA damage can positively regulate transcription initiation, the current techniques for mapping DNA breaks at high resolution, and how these techniques can benefit future studies of DNA damage and transcription.

  18. Altered minor-groove hydrogen bonds in DNA block transcription elongation by T7 RNA polymerase.

    Science.gov (United States)

    Tanasova, Marina; Goeldi, Silvan; Meyer, Fabian; Hanawalt, Philip C; Spivak, Graciela; Sturla, Shana J

    2015-05-26

    DNA transcription depends upon the highly efficient and selective function of RNA polymerases (RNAPs). Modifications in the template DNA can impact the progression of RNA synthesis, and a number of DNA adducts, as well as abasic sites, arrest or stall transcription. Nonetheless, data are needed to understand why certain modifications to the structure of DNA bases stall RNA polymerases while others are efficiently bypassed. In this study, we evaluate the impact that alterations in dNTP/rNTP base-pair geometry have on transcription. T7 RNA polymerase was used to study transcription over modified purines and pyrimidines with altered H-bonding capacities. The results suggest that introducing wobble base-pairs into the DNA:RNA heteroduplex interferes with transcriptional elongation and stalls RNA polymerase. However, transcriptional stalling is not observed if mismatched base-pairs do not H-bond. Together, these studies show that RNAP is able to discriminate mismatches resulting in wobble base-pairs, and suggest that, in cases of modifications with minor steric impact, DNA:RNA heteroduplex geometry could serve as a controlling factor for initiating transcription-coupled DNA repair. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. ABA signaling is necessary but not sufficient for RD29B transcriptional memory during successive dehydration stresses in Arabidopsis thaliana.

    Science.gov (United States)

    Virlouvet, Laetitia; Ding, Yong; Fujii, Hiroaki; Avramova, Zoya; Fromm, Michael

    2014-07-01

    Plants subjected to a prior dehydration stress were seen to have altered transcriptional responses during a subsequent dehydration stress for up to 5 days after the initial stress. The abscisic acid (ABA) inducible RD29B gene of Arabidopsis thaliana was strongly induced after the first stress and displayed transcriptional memory with transcript levels nine-fold higher during the second dehydration stress. These increased transcript levels were due to an increased rate of transcription and are associated with an altered chromatin template during the recovery interval between the dehydration stresses. Here we use a combination of promoter deletion/substitutions, mutants in the trans-acting transcription factors and their upstream protein kinases, and treatments with exogenous ABA or dehydration stress to advance our understanding of the features required for transcriptional memory of RD29B. ABA Response Elements (ABREs) are sufficient to confer transcriptional memory on a minimal promoter, although there is a context effect from flanking sequences. Different mutations in Snf1 Related Protein Kinase 2 (SnRK2) genes positively and negatively affected the response, suggesting that this effect is important for transcriptional memory. Although exogenous ABA treatments could prime transcriptional memory, a second ABA treatment was not sufficient to activate transcriptional memory. Therefore, we concluded that transcriptional memory requires ABA and an ABA-independent factor that is induced or activated by a subsequent dehydration stress and directly or indirectly results in a more active RD29B chromatin template. These results advance our knowledge of the cis- and trans-acting factors that are required for transcriptional memory of RD29B. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

  20. The Drosophila Helicase MLE Targets Hairpin Structures in Genomic Transcripts.

    Directory of Open Access Journals (Sweden)

    Simona Cugusi

    2016-01-01

    Full Text Available RNA hairpins are a common type of secondary structures that play a role in every aspect of RNA biochemistry including RNA editing, mRNA stability, localization and translation of transcripts, and in the activation of the RNA interference (RNAi and microRNA (miRNA pathways. Participation in these functions often requires restructuring the RNA molecules by the association of single-strand (ss RNA-binding proteins or by the action of helicases. The Drosophila MLE helicase has long been identified as a member of the MSL complex responsible for dosage compensation. The complex includes one of two long non-coding RNAs and MLE was shown to remodel the roX RNA hairpin structures in order to initiate assembly of the complex. Here we report that this function of MLE may apply to the hairpins present in the primary RNA transcripts that generate the small molecules responsible for RNA interference. Using stocks from the Transgenic RNAi Project and the Vienna Drosophila Research Center, we show that MLE specifically targets hairpin RNAs at their site of transcription. The association of MLE at these sites is independent of sequence and chromosome location. We use two functional assays to test the biological relevance of this association and determine that MLE participates in the RNAi pathway.

  1. RNA polymerase II mediated transcription from the polymerase III promoters in short hairpin RNA expression vector

    International Nuclear Information System (INIS)

    Rumi, Mohammad; Ishihara, Shunji; Aziz, Monowar; Kazumori, Hideaki; Ishimura, Norihisa; Yuki, Takafumi; Kadota, Chikara; Kadowaki, Yasunori; Kinoshita, Yoshikazu

    2006-01-01

    RNA polymerase III promoters of human ribonuclease P RNA component H1, human U6, and mouse U6 small nuclear RNA genes are commonly used in short hairpin RNA (shRNA) expression vectors due their precise initiation and termination sites. During transient transfection of shRNA vectors, we observed that H1 or U6 promoters also express longer transcripts enough to express several reporter genes including firefly luciferase, green fluorescent protein EGFP, and red fluorescent protein JRed. Expression of such longer transcripts was augmented by upstream RNA polymerase II enhancers and completely inhibited by downstream polyA signal sequences. Moreover, the transcription of firefly luciferase from human H1 promoter was sensitive to RNA polymerase II inhibitor α-amanitin. Our findings suggest that commonly used polymerase III promoters in shRNA vectors are also prone to RNA polymerase II mediated transcription, which may have negative impacts on their targeted use

  2. High-resolution structure of TBP with TAF1 reveals anchoring patterns in transcriptional regulation.

    Science.gov (United States)

    Anandapadamanaban, Madhanagopal; Andresen, Cecilia; Helander, Sara; Ohyama, Yoshifumi; Siponen, Marina I; Lundström, Patrik; Kokubo, Tetsuro; Ikura, Mitsuhiko; Moche, Martin; Sunnerhagen, Maria

    2013-08-01

    The general transcription factor TFIID provides a regulatory platform for transcription initiation. Here we present the crystal structure (1.97 Å) and NMR analysis of yeast TAF1 N-terminal domains TAND1 and TAND2 bound to yeast TBP, together with mutational data. We find that yeast TAF1-TAND1, which in itself acts as a transcriptional activator, binds TBP's concave DNA-binding surface by presenting similar anchor residues to TBP as does Mot1 but from a distinct structural scaffold. Furthermore, we show how TAF1-TAND2 uses an aromatic and acidic anchoring pattern to bind a conserved TBP surface groove traversing the basic helix region, and we find highly similar TBP-binding motifs also presented by the structurally distinct TFIIA, Mot1 and Brf1 proteins. Our identification of these anchoring patterns, which can be easily disrupted or enhanced, provides insight into the competitive multiprotein TBP interplay critical to transcriptional regulation.

  3. Transcriptional analysis of left-sided colitis, pancolitis, and ulcerative colitis-associated dysplasia

    DEFF Research Database (Denmark)

    Bjerrum, Jacob T; Nielsen, Ole H; Riis, Lene B

    2014-01-01

    to identify potential biomarkers and transcripts of importance for the carcinogenic behavior of chronic inflammation. METHODS: The Affymetrix GeneChip Human Genome U133 Plus 2.0 was applied on colonic biopsies from UC patients with left-sided UC, pancolitis, dysplasia, and controls. Reverse transcription...... polymerase chain reaction and immunohistochemistry were performed for validating selected transcripts in the initial cohort and in 2 independent cohorts of patients with UC. Microarray data were analyzed by principal component analysis, and reverse transcription polymerase chain reaction...... and immunohistochemistry data by the Wilcoxon's rank-sum test. RESULTS: The principal component analysis results revealed separate clusters for left-sided UC, pancolitis, dysplasia, and controls. Close clustering of dysplastic and pancolitic samples indicated similarities in gene expression. Indeed, 101 and 656 parallel...

  4. Mapping of gene transcripts by nuclease protection assays and cDNA primer extension

    International Nuclear Information System (INIS)

    Calzone, F.J.; Britten, R.J.; Davidson, E.J.

    1987-01-01

    An important problem often faced in the molecular characterization of genes is the precise mapping of those genomic sequences transcribed into RNA. This requires identification of the genomic site initiating gene transcription, the location of genomic sequences removed from the primary gene transcript during RNA processing, and knowledge of sequences terminating the processed gene transcript. The objective of the protocols described here is the generation of transcription maps utilizing relatively uncharacterized gene fragments. The basic approach is hybridization of a single-stranded DNA probe with cellular RNA, followed by treatment with a single-strand-specific nuclease that does not attack DNA-RNA hybrids, in order to destroy any unreacted probe sequences. Thus the probe sequences included in the hybrid duplexes are protected from nuclease digestion. The sizes of the protected probe fragments determined by gel electrophoresis correspond to the lengths of the hybridized sequence elements

  5. The logic of communication: roles for mobile transcription factors in plants.

    Science.gov (United States)

    Long, Yuchen; Scheres, Ben; Blilou, Ikram

    2015-02-01

    Mobile transcription factors play many roles in plant development. Here, we compare the use of mobile transcription factors as signals with some canonical signal transduction processes in prokaryotes and eukaryotes. After an initial survey, we focus on the SHORT-ROOT pathway in Arabidopsis roots to show that, despite the simplicity of the concept of mobile transcription factor signalling, many lines of evidence reveal a surprising complexity in control mechanisms linked to this process. We argue that these controls bestow precision, robustness, and versatility on mobile transcription factor signalling. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  6. Dual Regulation of Bacillus subtilis kinB Gene Encoding a Sporulation Trigger by SinR through Transcription Repression and Positive Stringent Transcription Control.

    Science.gov (United States)

    Fujita, Yasutaro; Ogura, Mitsuo; Nii, Satomi; Hirooka, Kazutake

    2017-01-01

    It is known that transcription of kinB encoding a trigger for Bacillus subtilis sporulation is under repression by SinR, a master repressor of biofilm formation, and under positive stringent transcription control depending on the adenine species at the transcription initiation nucleotide (nt). Deletion and base substitution analyses of the kinB promoter (P kinB ) region using lacZ fusions indicated that either a 5-nt deletion (Δ5, nt -61/-57, +1 is the transcription initiation nt) or the substitution of G at nt -45 with A (G-45A) relieved kinB repression. Thus, we found a pair of SinR-binding consensus sequences (GTTCTYT; Y is T or C) in an inverted orientation (SinR-1) between nt -57/-42, which is most likely a SinR-binding site for kinB repression. This relief from SinR repression likely requires SinI, an antagonist of SinR. Surprisingly, we found that SinR is essential for positive stringent transcription control of P kinB . Electrophoretic mobility shift assay (EMSA) analysis indicated that SinR bound not only to SinR-1 but also to SinR-2 (nt -29/-8) consisting of another pair of SinR consensus sequences in a tandem repeat arrangement; the two sequences partially overlap the '-35' and '-10' regions of P kinB . Introduction of base substitutions (T-27C C-26T) in the upstream consensus sequence of SinR-2 affected positive stringent transcription control of P kinB , suggesting that SinR binding to SinR-2 likely causes this positive control. EMSA also implied that RNA polymerase and SinR are possibly bound together to SinR-2 to form a transcription initiation complex for kinB transcription. Thus, it was suggested in this work that derepression of kinB from SinR repression by SinI induced by Spo0A∼P and occurrence of SinR-dependent positive stringent transcription control of kinB might induce effective sporulation cooperatively, implying an intimate interplay by stringent response, sporulation, and biofilm formation.

  7. Tonic 5nM DA stabilizes neuronal output by enabling bidirectional activity-dependent regulation of the hyperpolarization activated current via PKA and calcineurin.

    Science.gov (United States)

    Krenz, Wulf-Dieter C; Rodgers, Edmund W; Baro, Deborah J

    2015-01-01

    Volume transmission results in phasic and tonic modulatory signals. The actions of tonic dopamine (DA) at type 1 DA receptors (D1Rs) are largely undefined. Here we show that tonic 5nM DA acts at D1Rs to stabilize neuronal output over minutes by enabling activity-dependent regulation of the hyperpolarization activated current (I h). In the presence but not absence of 5nM DA, I h maximal conductance (G max) was adjusted according to changes in slow wave activity in order to maintain spike timing. Our study on the lateral pyloric neuron (LP), which undergoes rhythmic oscillations in membrane potential with depolarized plateaus, demonstrated that incremental, bi-directional changes in plateau duration produced corresponding alterations in LP I hG max when preparations were superfused with saline containing 5nM DA. However, when preparations were superfused with saline alone there was no linear correlation between LP I hGmax and duty cycle. Thus, tonic nM DA modulated the capacity for activity to modulate LP I h G max; this exemplifies metamodulation (modulation of modulation). Pretreatment with the Ca2+-chelator, BAPTA, or the specific PKA inhibitor, PKI, prevented all changes in LP I h in 5nM DA. Calcineurin inhibitors blocked activity-dependent changes enabled by DA and revealed a PKA-mediated, activity-independent enhancement of LP I hG max. These data suggested that tonic 5nM DA produced two simultaneous, PKA-dependent effects: a direct increase in LP I h G max and a priming event that permitted calcineurin regulation of LP I h. The latter produced graded reductions in LP I hG max with increasing duty cycles. We also demonstrated that this metamodulation preserved the timing of LP's first spike when network output was perturbed with bath-applied 4AP. In sum, 5nM DA permits slow wave activity to provide feedback that maintains spike timing, suggesting that one function of low-level, tonic modulation is to stabilize specific features of a dynamic output.

  8. Tonic 5nM DA stabilizes neuronal output by enabling bidirectional activity-dependent regulation of the hyperpolarization activated current via PKA and calcineurin.

    Directory of Open Access Journals (Sweden)

    Wulf-Dieter C Krenz

    Full Text Available Volume transmission results in phasic and tonic modulatory signals. The actions of tonic dopamine (DA at type 1 DA receptors (D1Rs are largely undefined. Here we show that tonic 5nM DA acts at D1Rs to stabilize neuronal output over minutes by enabling activity-dependent regulation of the hyperpolarization activated current (I h. In the presence but not absence of 5nM DA, I h maximal conductance (G max was adjusted according to changes in slow wave activity in order to maintain spike timing. Our study on the lateral pyloric neuron (LP, which undergoes rhythmic oscillations in membrane potential with depolarized plateaus, demonstrated that incremental, bi-directional changes in plateau duration produced corresponding alterations in LP I hG max when preparations were superfused with saline containing 5nM DA. However, when preparations were superfused with saline alone there was no linear correlation between LP I hGmax and duty cycle. Thus, tonic nM DA modulated the capacity for activity to modulate LP I h G max; this exemplifies metamodulation (modulation of modulation. Pretreatment with the Ca2+-chelator, BAPTA, or the specific PKA inhibitor, PKI, prevented all changes in LP I h in 5nM DA. Calcineurin inhibitors blocked activity-dependent changes enabled by DA and revealed a PKA-mediated, activity-independent enhancement of LP I hG max. These data suggested that tonic 5nM DA produced two simultaneous, PKA-dependent effects: a direct increase in LP I h G max and a priming event that permitted calcineurin regulation of LP I h. The latter produced graded reductions in LP I hG max with increasing duty cycles. We also demonstrated that this metamodulation preserved the timing of LP's first spike when network output was perturbed with bath-applied 4AP. In sum, 5nM DA permits slow wave activity to provide feedback that maintains spike timing, suggesting that one function of low-level, tonic modulation is to stabilize specific features of a dynamic output.

  9. Colon cancer associated transcripts in human cancers.

    Science.gov (United States)

    Chen, Yincong; Xie, Haibiao; Gao, Qunjun; Zhan, Hengji; Xiao, Huizhong; Zou, Yifan; Zhang, Fuyou; Liu, Yuchen; Li, Jianfa

    2017-10-01

    Long non-coding RNAs serve as important regulators in complicated cellular activities, including cell differentiation, proliferation and death. Dysregulation of long non-coding RNAs occurs in the formation and progression of cancers. The family of colon cancer associated transcripts, long non-coding RNAs colon cancer associated transcript-1 and colon cancer associated transcript-2 are known as oncogenes involved in various cancers. Colon cancer associated transcript-1 is a novel lncRNA located in 8q24.2, and colon cancer associated transcript-2 maps to the 8q24.21 region encompassing rs6983267. Colon cancer associated transcripts have close associations with clinical characteristics, such as lymph node metastasis, high TNM stage and short overall survival. Knockdown of them can reverse the malignant phenotypes of cancer cells, including proliferation, migration, invasion and apoptosis. Moreover, they can increase the expression level of c-MYC and oncogenic microRNAs via activating a series of complex mechanisms. In brief, the family of colon cancer associated transcripts may serve as potential biomarkers or therapeutic targets for human cancers. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  10. From reverse transcription to human brain tumors

    Directory of Open Access Journals (Sweden)

    Dmitrenko V. V.

    2013-05-01

    Full Text Available Reverse transcriptase from avian myeloblastosis virus (AMV was the subject of the study, from which the investi- gations of the Department of biosynthesis of nucleic acids were started. Production of AMV in grams quantities and isolation of AMV reverse transcriptase were established in the laboratory during the seventies of the past cen- tury and this initiated research on the cDNA synthesis, cloning and investigation of the structure and functions of the eukaryotic genes. Structures of salmon insulin and insulin-like growth factor (IGF family genes and their transcripts were determined during long-term investigations. Results of two modern techniques, microarray-ba- sed hybridization and SAGE, were used for the identification of the genes differentially expressed in astrocytic gliomas and human normal brain. Comparison of SAGE results on the genes overexpressed in glioblastoma with the results of microarray analysis revealed a limited number of common genes. 105 differentially expressed genes, common to both methods, can be included in the list of candidates for the molecular typing of glioblastoma. The first experiments on the classification of glioblastomas based on the data of the 20 genes expression were conducted by using of artificial neural network analysis. The results of these experiments showed that the expression profiles of these genes in 224 glioblastoma samples and 74 normal brain samples could be according to the Koho- nen’s maps. The CHI3L1 and CHI3L2 genes of chitinase-like cartilage protein were revealed among the most overexpressed genes in glioblastoma, which could have prognostic and diagnostic potential. Results of in vitro experiments demonstrated that both proteins, CHI3L1 and CHI3L2, may initiate the phosphorylation of ERK1/ ERK2 and AKT kinases leading to the activation of MAPK/ERK1/2 and PI3K/AKT signaling cascades in human embryonic kidney 293 cells, human glioblastoma U87MG, and U373 cells. The new human cell line

  11. Transcriptional mapping of rabies virus in vivo

    International Nuclear Information System (INIS)

    Flamand, A.; Delagneau, J.F.

    1978-01-01

    Synthesis of the proteins of rabies virus was studied in hamster cell infected with uv-irradiated virus. The uv target size of genes L, N, M 1 , and M 2 was measured during primary transcription. Except for N, the target size of the remaining genes was considerably larger than that of their physical sizes. The data fit the hypothesis that four genes occupy a single transcriptional unit and that transcription of rabies virus proceeds in the order N, M 1 , M 2 , and L

  12. NAC transcription factors: structurally distinct, functionally diverse

    DEFF Research Database (Denmark)

    Olsen, Addie Nina; Ernst, Heidi A; Leggio, Leila Lo

    2005-01-01

    level and localization, and to the first indications of NAC participation in transcription factor networks. The recent determination of the DNA and protein binding NAC domain structure offers insight into the molecular functions of the protein family. Research into NAC transcription factors has......NAC proteins constitute one of the largest families of plant-specific transcription factors, and the family is present in a wide range of land plants. Here, we summarize the biological and molecular functions of the NAC family, paying particular attention to the intricate regulation of NAC protein...

  13. Initialized Fractional Calculus

    Science.gov (United States)

    Lorenzo, Carl F.; Hartley, Tom T.

    2000-01-01

    This paper demonstrates the need for a nonconstant initialization for the fractional calculus and establishes a basic definition set for the initialized fractional differintegral. This definition set allows the formalization of an initialized fractional calculus. Two basis calculi are considered; the Riemann-Liouville and the Grunwald fractional calculi. Two forms of initialization, terminal and side are developed.

  14. Simplified Method for Predicting a Functional Class of Proteins in Transcription Factor Complexes

    KAUST Repository

    Piatek, Marek J.

    2013-07-12

    Background:Initiation of transcription is essential for most of the cellular responses to environmental conditions and for cell and tissue specificity. This process is regulated through numerous proteins, their ligands and mutual interactions, as well as interactions with DNA. The key such regulatory proteins are transcription factors (TFs) and transcription co-factors (TcoFs). TcoFs are important since they modulate the transcription initiation process through interaction with TFs. In eukaryotes, transcription requires that TFs form different protein complexes with various nuclear proteins. To better understand transcription regulation, it is important to know the functional class of proteins interacting with TFs during transcription initiation. Such information is not fully available, since not all proteins that act as TFs or TcoFs are yet annotated as such, due to generally partial functional annotation of proteins. In this study we have developed a method to predict, using only sequence composition of the interacting proteins, the functional class of human TF binding partners to be (i) TF, (ii) TcoF, or (iii) other nuclear protein. This allows for complementing the annotation of the currently known pool of nuclear proteins. Since only the knowledge of protein sequences is required in addition to protein interaction, the method should be easily applicable to many species.Results:Based on experimentally validated interactions between human TFs with different TFs, TcoFs and other nuclear proteins, our two classification systems (implemented as a web-based application) achieve high accuracies in distinguishing TFs and TcoFs from other nuclear proteins, and TFs from TcoFs respectively.Conclusion:As demonstrated, given the fact that two proteins are capable of forming direct physical interactions and using only information about their sequence composition, we have developed a completely new method for predicting a functional class of TF interacting protein partners

  15. Activity-Based Anorexia Alters the Expression of BDNF Transcripts in the Mesocorticolimbic Reward Circuit.

    Science.gov (United States)

    Ho, Emily V; Klenotich, Stephanie J; McMurray, Matthew S; Dulawa, Stephanie C

    2016-01-01

    Anorexia nervosa (AN) is a complex eating disorder with severe dysregulation of appetitive behavior. The activity-based anorexia (ABA) paradigm is an animal model in which rodents exposed to both running wheels and scheduled feeding develop aspects of AN including paradoxical hypophagia, dramatic weight loss, and hyperactivity, while animals exposed to only one condition maintain normal body weight. Brain-derived neurotrophic factor (BDNF), an activity-dependent modulator of neuronal plasticity, is reduced in the serum of AN patients, and is a known regulator of feeding and weight maintenance. We assessed the effects of scheduled feeding, running wheel access, or both on the expression of BDNF transcripts within the mesocorticolimbic pathway. We also assessed the expression of neuronal cell adhesion molecule 1 (NCAM1) to explore the specificity of effects on BDNF within the mesocorticolimbic pathway. Scheduled feeding increased the levels of both transcripts in the hippocampus (HPC), increased NCAM1 mRNA expression in the ventral tegmental area (VTA), and decreased BDNF mRNA levels in the medial prefrontal cortex (mPFC). In addition, wheel running increased BDNF mRNA expression in the VTA. No changes in either transcript were observed in the nucleus accumbens (NAc). Furthermore, no changes in either transcript were induced by the combined scheduled feeding and wheel access condition. These data indicate that scheduled feeding or wheel running alter BDNF and NCAM1 expression levels in specific regions of the mesocorticolimbic pathway. These findings contribute to our current knowledge of the molecular alterations induced by ABA and may help elucidate possible mechanisms of AN pathology.

  16. Activity-dependent expression of miR-132 regulates immediate-early gene induction during olfactory learning in the greater short-nosed fruit bat, Cynopterus sphinx.

    Science.gov (United States)

    Mukilan, Murugan; Ragu Varman, Durairaj; Sudhakar, Sivasubramaniam; Rajan, Koilmani Emmanuvel

    2015-04-01

    The activity-dependent expression of immediate-early genes (IEGs) and microRNA (miR)-132 has been implicated in synaptic plasticity and the formation of long-term memory (LTM). In the present study, we show that olfactory training induces the expression of IEGs (EGR-1, C-fos, C-jun) and miR-132 at similar time scale in olfactory bulb (OB) of Cynopterus sphinx. We examined the role of miR-132 in the OB using antisense oligodeoxynucleotide (AS-ODN) and demonstrated that a local infusion of AS-ODN in the OB 2h prior to training impaired olfactory memory formation in C. sphinx. However, the infusion of AS-ODN post-training did not cause a deficit in memory formation. Furthermore, the inhibition of miR-132 reduced the olfactory training-induced expression of IEGs and post synaptic density protein-95 (PSD-95) in the OB. Additionally, we show that miR-132 regulates the activation of calcium/calmodulin-dependent protein kinase-II (CaMKII) and cAMP response element binding protein (CREB), possibly through miR-148a. These data suggest that olfactory training induces the expression of miR-132 and IEGs, which in turn activates post-synaptic proteins that regulate olfactory memory formation. Copyright © 2015 Elsevier Inc. All rights reserved.

  17. Phospholipase A2 activity-dependent and -independent fusogenic activity of Naja nigricollis CMS-9 on zwitterionic and anionic phospholipid vesicles.

    Science.gov (United States)

    Chiou, Yi-Ling; Chen, Ying-Jung; Lin, Shinne-Ren; Chang, Long-Sen

    2011-11-01

    CMS-9, a phospholipase A(2) (PLA(2)) from Naja nigricollis venom, induced the death of human breast cancer MCF-7 cells accompanied with the formation of cell clumps without clear boundaries between cells. Annexin V-FITC staining indicated that abundant phosphatidylserine appeared on the outer membrane of MCF-7 cell clumps, implying the possibility that CMS-9 may promote membrane fusion via anionic phospholipids. To validate this proposition, fusogenic activity of CMS-9 on vesicles composed of zwitterionic phospholipid alone or a combination of zwitterionic and anionic phospholipids was examined. Although CMS-9-induced fusion of zwitterionic phospholipid vesicles depended on PLA(2) activity, CMS-9-induced fusion of vesicles containing anionic phospholipids could occur without the involvement of PLA(2) activity. Membrane-damaging activity of CMS-9 was associated with its fusogenicity. Moreover, CMS-9 induced differently membrane leakage and membrane fusion of vesicles with different compositions. Membrane fluidity and binding capability with phospholipid vesicles were not related to the fusogenicity of CMS-9. However, membrane-bound conformation and mode of CMS-9 depended on phospholipid compositions. Collectively, our data suggest that PLA(2) activity-dependent and -independent fusogenicity of CMS-9 are closely related to its membrane-bound modes and targeted membrane compositions. Copyright © 2011 Elsevier Ltd. All rights reserved.

  18. Unveiling clusters of RNA transcript pairs associated with markers of Alzheimer's disease progression.

    Directory of Open Access Journals (Sweden)

    Ahmed Shamsul Arefin

    Full Text Available BACKGROUND: One primary goal of transcriptomic studies is identifying gene expression patterns correlating with disease progression. This is usually achieved by considering transcripts that independently pass an arbitrary threshold (e.g. p<0.05. In diseases involving severe perturbations of multiple molecular systems, such as Alzheimer's disease (AD, this univariate approach often results in a large list of seemingly unrelated transcripts. We utilised a powerful multivariate clustering approach to identify clusters of RNA biomarkers strongly associated with markers of AD progression. We discuss the value of considering pairs of transcripts which, in contrast to individual transcripts, helps avoid natural human transcriptome variation that can overshadow disease-related changes. METHODOLOGY/PRINCIPAL FINDINGS: We re-analysed a dataset of hippocampal transcript levels in nine controls and 22 patients with varying degrees of AD. A large-scale clustering approach determined groups of transcript probe sets that correlate strongly with measures of AD progression, including both clinical and neuropathological measures and quantifiers of the characteristic transcriptome shift from control to severe AD. This enabled identification of restricted groups of highly correlated probe sets from an initial list of 1,372 previously published by our group. We repeated this analysis on an expanded dataset that included all pair-wise combinations of the 1,372 probe sets. As clustering of this massive dataset is unfeasible using standard computational tools, we adapted and re-implemented a clustering algorithm that uses external memory algorithmic approach. This identified various pairs that strongly correlated with markers of AD progression and highlighted important biological pathways potentially involved in AD pathogenesis. CONCLUSIONS/SIGNIFICANCE: Our analyses demonstrate that, although there exists a relatively large molecular signature of AD progression, only

  19. Breaking the mold: transcription factors in the anucleate platelet and platelet-derived microparticles

    Directory of Open Access Journals (Sweden)

    Katie L Lannan

    2015-02-01

    Full Text Available Platelets are small anucleate blood cells derived from megakaryocytes. In addition to their pivotal roles in hemostasis, platelets are the smallest, yet most abundant, immune cell and regulate inflammation, immunity, and disease progression. Although platelets lack DNA, and thus no functional transcriptional activities, they are nonetheless rich sources of RNAs, possess an intact spliceosome, and are thus capable of synthesizing proteins. Previously, it was thought that platelet RNAs and translational machinery were remnants from the megakaryocyte. We now know that the initial description of platelets as cellular fragments is an antiquated notion, as mounting evidence suggests otherwise. Therefore, it is reasonable to hypothesize that platelet transcription factors are not vestigial remnants from megakaryoctes, but have important, if only partly understood functions. Proteins play multiple cellular roles to minimize energy expenditure for maximum cellular function; thus, the same can be expected for transcription factors. In fact, numerous transcription factors have non-genomic roles, both in platelets and in nucleated cells. Our lab and others have discovered the presence and nongenomic roles of transcription factors in platelets, such as the nuclear factor kappa β (NFκB family of proteins and peroxisome proliferator activated receptor gamma (PPARγ. In addition to numerous roles in regulating platelet activation, functional transcription factors can be transferred to vascular and immune cells through platelet microparticles. This method of transcellular delivery of key immune molecules may be a vital mechanism by which platelet transcription factors regulate inflammation and immunity. At the very least, platelets are an ideal model cell to dissect out the nongenomic roles of transcription factors in nucleated cells. There is abundant evidence to suggest that transcription factors in platelets play key roles in regulating inflammatory and

  20. Repression of meiotic genes by antisense transcription and by Fkh2 transcription factor in Schizosaccharomyces pombe.

    Science.gov (United States)

    Chen, Huei-Mei; Rosebrock, Adam P; Khan, Sohail R; Futcher, Bruce; Leatherwood, Janet K

    2012-01-01

    In S. pombe, about 5% of genes are meiosis-specific and accumulate little or no mRNA during vegetative growth. Here we use Affymetrix tiling arrays to characterize transcripts in vegetative and meiotic cells. In vegetative cells, many meiotic genes, especially those induced in mid-meiosis, have abundant antisense transcripts. Disruption of the antisense transcription of three of these mid-meiotic genes allowed vegetative sense transcription. These results suggest that antisense transcription represses sense transcription of meiotic genes in vegetative cells. Although the mechanism(s) of antisense mediated transcription repression need to be further explored, our data indicates that RNAi machinery is not required for repression. Previously, we and others used non-strand specific methods to study splicing regulation of meiotic genes and concluded that 28 mid-meiotic genes are spliced only in meiosis. We now demonstrate that the "unspliced" signal in vegetative cells comes from the antisense RNA, not from unspliced sense RNA, and we argue against the idea that splicing regulates these mid-meiotic genes. Most of these mid-meiotic genes are induced in mid-meiosis by the forkhead transcription factor Mei4. Interestingly, deletion of a different forkhead transcription factor, Fkh2, allows low levels of sense expression of some mid-meiotic genes in vegetative cells. We propose that vegetative expression of mid-meiotic genes is repressed at least two independent ways: antisense transcription and Fkh2 repression.

  1. Repression of Meiotic Genes by Antisense Transcription and by Fkh2 Transcription Factor in Schizosaccharomyces pombe

    Science.gov (United States)

    Chen, Huei-Mei; Rosebrock, Adam P.; Khan, Sohail R.; Futcher, Bruce; Leatherwood, Janet K.

    2012-01-01

    In S. pombe, about 5% of genes are meiosis-specific and accumulate little or no mRNA during vegetative growth. Here we use Affymetrix tiling arrays to characterize transcripts in vegetative and meiotic cells. In vegetative cells, many meiotic genes, especially those induced in mid-meiosis, have abundant antisense transcripts. Disruption of the antisense transcription of three of these mid-meiotic genes allowed vegetative sense transcription. These results suggest that antisense transcription represses sense transcription of meiotic genes in vegetative cells. Although the mechanism(s) of antisense mediated transcription repression need to be further explored, our data indicates that RNAi machinery is not required for repression. Previously, we and others used non-strand specific methods to study splicing regulation of meiotic genes and concluded that 28 mid-meiotic genes are spliced only in meiosis. We now demonstrate that the “unspliced” signal in vegetative cells comes from the antisense RNA, not from unspliced sense RNA, and we argue against the idea that splicing regulates these mid-meiotic genes. Most of these mid-meiotic genes are induced in mid-meiosis by the forkhead transcription factor Mei4. Interestingly, deletion of a different forkhead transcription factor, Fkh2, allows low levels of sense expression of some mid-meiotic genes in vegetative cells. We propose that vegetative expression of mid-meiotic genes is repressed at least two independent ways: antisense transcription and Fkh2 repression. PMID:22238674

  2. Repression of meiotic genes by antisense transcription and by Fkh2 transcription factor in Schizosaccharomyces pombe.

    Directory of Open Access Journals (Sweden)

    Huei-Mei Chen

    Full Text Available In S. pombe, about 5% of genes are meiosis-specific and accumulate little or no mRNA during vegetative growth. Here we use Affymetrix tiling arrays to characterize transcripts in vegetative and meiotic cells. In vegetative cells, many meiotic genes, especially those induced in mid-meiosis, have abundant antisense transcripts. Disruption of the antisense transcription of three of these mid-meiotic genes allowed vegetative sense transcription. These results suggest that antisense transcription represses sense transcription of meiotic genes in vegetative cells. Although the mechanism(s of antisense mediated transcription repression need to be further explored, our data indicates that RNAi machinery is not required for repression. Previously, we and others used non-strand specific methods to study splicing regulation of meiotic genes and concluded that 28 mid-meiotic genes are spliced only in meiosis. We now demonstrate that the "unspliced" signal in vegetative cells comes from the antisense RNA, not from unspliced sense RNA, and we argue against the idea that splicing regulates these mid-meiotic genes. Most of these mid-meiotic genes are induced in mid-meiosis by the forkhead transcription factor Mei4. Interestingly, deletion of a different forkhead transcription factor, Fkh2, allows low levels of sense expression of some mid-meiotic genes in vegetative cells. We propose that vegetative expression of mid-meiotic genes is repressed at least two independent ways: antisense transcription and Fkh2 repression.

  3. In silico and wet lab approaches to study transcriptional regulation

    NARCIS (Netherlands)

    Hestand, Matthew Scott

    2010-01-01

    Gene expression is a complicated process with multiple types of regulation, including binding of proteins termed transcription factors. This thesis looks at transcription factors and transcription factor binding site discovery through computational predictions and wet lab work to better elucidate

  4. Synthesis of in vitro Co1E1 transcripts with 5'-terminal ribonucleotides that exhibit noncomplementarity with the DNA template

    International Nuclear Information System (INIS)

    Parker, R.C.

    1986-01-01

    A region that forms the S1 nuclease site in Co1E1 DNA is shown to code for an in vitro transcript, called S1 RNA-B, which contains a 5'-terminal GTP residue that exhibits noncomplementarity with the template's DNA sequence. The synthesis of S1 RNA-B initiates four bases upstream from the start point for S1 RNA-C. The initial four bases in S1 RNA-B and S1 RNA-C are identical. The relative synthesis of S1 RNA-B to S1 RNA-C is sensitive to the concentration of GTP, a substrate that is required for elongation past the +4 position in S1 RNA-C. Dinucleotides that are expected to only initiate synthesis of S1 RNA-C yield two transcripts that appear to initiate from the S1 RNA-C and S1 RNA-B start sites. In vitro studies involving other Co1E1 transcripts, RNA-B and RNA-C, provide similar observations concerning the noncomplementary initiation phenomenon. A model involving transcriptional slippage is suggested to explain the noncomplementary initiation phenomenon. The model proposes that the cycling reaction of Escherichia coli RNA polymerase produces tetranucleotides that are transposed to nearby upstream sequences for priming transcription

  5. Comparison of Transcription Factor Binding Site Models

    KAUST Repository

    Bhuyan, Sharifulislam

    2012-05-01

    Modeling of transcription factor binding sites (TFBSs) and TFBS prediction on genomic sequences are important steps to elucidate transcription regulatory mechanism. Dependency of transcription regulation on a great number of factors such as chemical specificity, molecular structure, genomic and epigenetic characteristics, long distance interaction, makes this a challenging problem. Different experimental procedures generate evidence that DNA-binding domains of transcription factors show considerable DNA sequence specificity. Probabilistic modeling of TFBSs has been moderately successful in identifying patterns from a family of sequences. In this study, we compare performances of different probabilistic models and try to estimate their efficacy over experimental TFBSs data. We build a pipeline to calculate sensitivity and specificity from aligned TFBS sequences for several probabilistic models, such as Markov chains, hidden Markov models, Bayesian networks. Our work, containing relevant statistics and evaluation for the models, can help researchers to choose the most appropriate model for the problem at hand.

  6. Battles and hijacks: Noncoding transcription in plants

    KAUST Repository

    Ariel, Federico; Romero-Barrios, Natali; Jé gu, Teddy; Benhamed, Moussa; Crespi, Martin

    2015-01-01

    splicing, fine-tuning of miRNA activity, and the control of mRNA translation or accumulation. Recently, dual noncoding transcription by alternative RNA polymerases was implicated in epigenetic and chromatin conformation dynamics. This review integrates

  7. Salmonella Typhimurium transcription profiles in space flight

    Data.gov (United States)

    National Aeronautics and Space Administration — Salmonella transcription profiles were obtained from samples flown on space shuttle mission STS-115 and compared to profiles from Salmonella grown under identical...

  8. Transcript for Understanding Medical Words: A Tutorial

    Science.gov (United States)

    ... medlineplus.gov/medicalwordstranscript.html Transcript for Understanding Medical Words: A Tutorial To use the sharing features on ... get to what those mean in a minute. Word Roots Word Roots. Let's begin with body parts. ...

  9. The specificity and flexibility of l1 reverse transcription priming at imperfect T-tracts.

    Directory of Open Access Journals (Sweden)

    Clément Monot

    2013-05-01

    Full Text Available L1 retrotransposons have a prominent role in reshaping mammalian genomes. To replicate, the L1 ribonucleoprotein particle (RNP first uses its endonuclease (EN to nick the genomic DNA. The newly generated DNA end is subsequently used as a primer to initiate reverse transcription within the L1 RNA poly(A tail, a process known as target-primed reverse transcription (TPRT. Prior studies demonstrated that most L1 insertions occur into sequences related to the L1 EN consensus sequence (degenerate 5'-TTTT/A-3' sites and frequently preceded by imperfect T-tracts. However, it is currently unclear whether--and to which degree--the liberated 3'-hydroxyl extremity on the genomic DNA needs to be accessible and complementary to the poly(A tail of the L1 RNA for efficient priming of reverse transcription. Here, we employed a direct assay for the initiation of L1 reverse transcription to define the molecular rules that guide this process. First, efficient priming is detected with as few as 4 matching nucleotides at the primer 3' end. Second, L1 RNP can tolerate terminal mismatches if they are compensated within the 10 last bases of the primer by an increased number of matching nucleotides. All terminal mismatches are not equally detrimental to DNA extension, a C being extended at higher levels than an A or a G. Third, efficient priming in the context of duplex DNA requires a 3' overhang. This suggests the possible existence of additional DNA processing steps, which generate a single-stranded 3' end to allow L1 reverse transcription. Based on these data we propose that the specificity of L1 reverse transcription initiation contributes, together with the specificity of the initial EN cleavage, to the distribution of new L1 insertions within the human genome.

  10. The specificity and flexibility of l1 reverse transcription priming at imperfect T-tracts.

    Science.gov (United States)

    Monot, Clément; Kuciak, Monika; Viollet, Sébastien; Mir, Ashfaq Ali; Gabus, Caroline; Darlix, Jean-Luc; Cristofari, Gaël

    2013-05-01

    L1 retrotransposons have a prominent role in reshaping mammalian genomes. To replicate, the L1 ribonucleoprotein particle (RNP) first uses its endonuclease (EN) to nick the genomic DNA. The newly generated DNA end is subsequently used as a primer to initiate reverse transcription within the L1 RNA poly(A) tail, a process known as target-primed reverse transcription (TPRT). Prior studies demonstrated that most L1 insertions occur into sequences related to the L1 EN consensus sequence (degenerate 5'-TTTT/A-3' sites) and frequently preceded by imperfect T-tracts. However, it is currently unclear whether--and to which degree--the liberated 3'-hydroxyl extremity on the genomic DNA needs to be accessible and complementary to the poly(A) tail of the L1 RNA for efficient priming of reverse transcription. Here, we employed a direct assay for the initiation of L1 reverse transcription to define the molecular rules that guide this process. First, efficient priming is detected with as few as 4 matching nucleotides at the primer 3' end. Second, L1 RNP can tolerate terminal mismatches if they are compensated within the 10 last bases of the primer by an increased number of matching nucleotides. All terminal mismatches are not equally detrimental to DNA extension, a C being extended at higher levels than an A or a G. Third, efficient priming in the context of duplex DNA requires a 3' overhang. This suggests the possible existence of additional DNA processing steps, which generate a single-stranded 3' end to allow L1 reverse transcription. Based on these data we propose that the specificity of L1 reverse transcription initiation contributes, together with the specificity of the initial EN cleavage, to the distribution of new L1 insertions within the human genome.

  11. APOBEC3G inhibits elongation of HIV-1 reverse transcripts.

    Directory of Open Access Journals (Sweden)

    Kate N Bishop

    2008-12-01

    Full Text Available APOBEC3G (A3G is a host cytidine deaminase that, in the absence of Vif, restricts HIV-1 replication and reduces the amount of viral DNA that accumulates in cells. Initial studies determined that A3G induces extensive mutation of nascent HIV-1 cDNA during reverse transcription. It has been proposed that this triggers the degradation of the viral DNA, but there is now mounting evidence that this mechanism may not be correct. Here, we use a natural endogenous reverse transcriptase assay to show that, in cell-free virus particles, A3G is able to inhibit HIV-1 cDNA accumulation not only in the absence of hypermutation but also without the apparent need for any target cell factors. We find that although reverse transcription initiates in the presence of A3G, elongation of the cDNA product is impeded. These data support the model that A3G reduces HIV-1 cDNA levels by inhibiting synthesis rather than by inducing degradation.

  12. Transcriptional Waves in the Yeast Cell Cycle

    OpenAIRE

    Oliva, Anna; Rosebrock, Adam; Ferrezuelo, Francisco; Pyne, Saumyadipta; Chen, Haiying; Skiena, Steve; Futcher, Bruce; Leatherwood, Janet

    2005-01-01

    Many genes are regulated as an innate part of the eukaryotic cell cycle, and a complex transcriptional network helps enable the cyclic behavior of dividing cells. This transcriptional network has been studied in Saccharomyces cerevisiae (budding yeast) and elsewhere. To provide more perspective on these regulatory mechanisms, we have used microarrays to measure gene expression through the cell cycle of Schizosaccharomyces pombe (fission yeast). The 750 genes with the most significant oscillat...

  13. RNA polymerase gate loop guides the nontemplate DNA strand in transcription complexes.

    Science.gov (United States)

    NandyMazumdar, Monali; Nedialkov, Yuri; Svetlov, Dmitri; Sevostyanova, Anastasia; Belogurov, Georgiy A; Artsimovitch, Irina

    2016-12-27

    Upon RNA polymerase (RNAP) binding to a promoter, the σ factor initiates DNA strand separation and captures the melted nontemplate DNA, whereas the core enzyme establishes interactions with the duplex DNA in front of the active site that stabilize initiation complexes and persist throughout elongation. Among many core RNAP elements that participate in these interactions, the β' clamp domain plays the most prominent role. In this work, we investigate the role of the β gate loop, a conserved and essential structural element that lies across the DNA channel from the clamp, in transcription regulation. The gate loop was proposed to control DNA loading during initiation and to interact with NusG-like proteins to lock RNAP in a closed, processive state during elongation. We show that the removal of the gate loop has large effects on promoter complexes, trapping an unstable intermediate in which the RNAP contacts with the nontemplate strand discriminator region and the downstream duplex DNA are not yet fully established. We find that although RNAP lacking the gate loop displays moderate defects in pausing, transcript cleavage, and termination, it is fully responsive to the transcription elongation factor NusG. Together with the structural data, our results support a model in which the gate loop, acting in concert with initiation or elongation factors, guides the nontemplate DNA in transcription complexes, thereby modulating their regulatory properties.

  14. A biophysical model for transcription factories

    International Nuclear Information System (INIS)

    Canals-Hamann, Ana Z; Neves, Ricardo Pires das; Reittie, Joyce E; Iñiguez, Carlos; Soneji, Shamit; Enver, Tariq; Buckle, Veronica J; Iborra, Francisco J

    2013-01-01

    Transcription factories are nuclear domains where gene transcription takes place although the molecular basis for their formation and maintenance are unknown. In this study, we explored how the properties of chromatin as a polymer may contribute to the structure of transcription factories. We found that transcriptional active chromatin contains modifications like histone H4 acetylated at Lysine 16 (H4K16ac). Single fibre analysis showed that this modification spans the entire body of the gene. Furthermore, H4K16ac genes cluster in regions up to 500 Kb alternating active and inactive chromatin. The introduction of H4K16ac in chromatin induces stiffness in the chromatin fibre. The result of this change in flexibility is that chromatin could behave like a multi-block copolymer with repetitions of stiff-flexible (active-inactive chromatin) components. Copolymers with such structure self-organize through spontaneous phase separation into microdomains. Consistent with such model H4K16ac chromatin form foci that associates with nascent transcripts. We propose that transcription factories are the result of the spontaneous concentration of H4K16ac chromatin that are in proximity, mainly in cis

  15. MADS-box gene evolution - structure and transcription patterns

    DEFF Research Database (Denmark)

    Johansen, Bo; Pedersen, Louise Buchholt; Skipper, Martin

    2002-01-01

    Mads-box genes, ABC model, Evolution, Phylogeny, Transcription patterns, Gene structure, Conserved motifs......Mads-box genes, ABC model, Evolution, Phylogeny, Transcription patterns, Gene structure, Conserved motifs...

  16. Initiation of DNA replication requires actin dynamics and formin activity.

    Science.gov (United States)

    Parisis, Nikolaos; Krasinska, Liliana; Harker, Bethany; Urbach, Serge; Rossignol, Michel; Camasses, Alain; Dewar, James; Morin, Nathalie; Fisher, Daniel

    2017-11-02

    Nuclear actin regulates transcriptional programmes in a manner dependent on its levels and polymerisation state. This dynamics is determined by the balance of nucleocytoplasmic shuttling, formin- and redox-dependent filament polymerisation. Here, using Xenopus egg extracts and human somatic cells, we show that actin dynamics and formins are essential for DNA replication. In proliferating cells, formin inhibition abolishes nuclear transport and initiation of DNA replication, as well as general transcription. In replicating nuclei from transcriptionally silent Xenopus egg extracts, we identified numerous actin regulators, and disruption of actin dynamics abrogates nuclear transport, preventing NLS (nuclear localisation signal)-cargo release from RanGTP-importin complexes. Nuclear formin activity is further required to promote loading of cyclin-dependent kinase (CDK) and proliferating cell nuclear antigen (PCNA) onto chromatin, as well as initiation and elongation of DNA replication. Therefore, actin dynamics and formins control DNA replication by multiple direct and indirect mechanisms. © 2017 The Authors.

  17. Dynamic analysis of stochastic transcription cycles.

    Directory of Open Access Journals (Sweden)

    Claire V Harper

    2011-04-01

    Full Text Available In individual mammalian cells the expression of some genes such as prolactin is highly variable over time and has been suggested to occur in stochastic pulses. To investigate the origins of this behavior and to understand its functional relevance, we quantitatively analyzed this variability using new mathematical tools that allowed us to reconstruct dynamic transcription rates of different reporter genes controlled by identical promoters in the same living cell. Quantitative microscopic analysis of two reporter genes, firefly luciferase and destabilized EGFP, was used to analyze the dynamics of prolactin promoter-directed gene expression in living individual clonal and primary pituitary cells over periods of up to 25 h. We quantified the time-dependence and cyclicity of the transcription pulses and estimated the length and variation of active and inactive transcription phases. We showed an average cycle period of approximately 11 h and demonstrated that while the measured time distribution of active phases agreed with commonly accepted models of transcription, the inactive phases were differently distributed and showed strong memory, with a refractory period of transcriptional inactivation close to 3 h. Cycles in transcription occurred at two distinct prolactin-promoter controlled reporter genes in the same individual clonal or primary cells. However, the timing of the cycles was independent and out-of-phase. For the first time, we have analyzed transcription dynamics from two equivalent loci in real-time in single cells. In unstimulated conditions, cells showed independent transcription dynamics at each locus. A key result from these analyses was the evidence for a minimum refractory period in the inactive-phase of transcription. The response to acute signals and the result of manipulation of histone acetylation was consistent with the hypothesis that this refractory period corresponded to a phase of chromatin remodeling which significantly

  18. Intracellular ATP influences synaptic plasticity in area CA1 of rat hippocampus via metabolism to adenosine and activity-dependent activation of adenosine A1 receptors.

    Science.gov (United States)

    zur Nedden, Stephanie; Hawley, Simon; Pentland, Naomi; Hardie, D Grahame; Doney, Alexander S; Frenguelli, Bruno G

    2011-04-20

    The extent to which brain slices reflect the energetic status of the in vivo brain has been a subject of debate. We addressed this issue to investigate the recovery of energetic parameters and adenine nucleotides in rat hippocampal slices and the influence this has on synaptic transmission and plasticity. We show that, although adenine nucleotide levels recover appreciably within 10 min of incubation, it takes 3 h for a full recovery of the energy charge (to ≥ 0.93) and that incubation of brain slices at 34°C results in a significantly higher ATP/AMP ratio and a threefold lower activity of AMP-activated protein kinase compared with slices incubated at room temperature. Supplementation of artificial CSF with d-ribose and adenine (Rib/Ade) increased the total adenine nucleotide pool of brain slices, which, when corrected for the influence of the dead cut edges, closely approached in vivo values. Rib/Ade did not affect basal synaptic transmission or paired-pulse facilitation but did inhibit long-term potentiation (LTP) induced by tetanic or weak theta-burst stimulation. This decrease in LTP was reversed by strong theta-burst stimulation or antagonizing the inhibitory adenosine A(1) receptor suggesting that the elevated tissue ATP levels had resulted in greater activity-dependent adenosine release during LTP induction. This was confirmed by direct measurement of adenosine release with adenosine biosensors. These observations provide new insight into the recovery of adenine nucleotides after slice preparation, the sources of loss of such compounds in brain slices, the means by which to restore them, and the functional consequences of doing so.

  19. Dioxin Exposure Initiative

    Science.gov (United States)

    The Dioxin Exposure Initiative (DEI) is no longer active. This page contains a summary of the dioxin exposure initiative with illustrations, contact and background information.Originally supported by scientist Matthew Lorber, who retired in Mar 2017.

  20. A deeper look into transcription regulatory code by preferred pair distance templates for transcription factor binding sites

    KAUST Repository

    Kulakovskiy, Ivan V.; Belostotsky, A. A.; Kasianov, Artem S.; Esipova, Natalia G.; Medvedeva, Yulia; Eliseeva, Irina A.; Makeev, Vsevolod J.

    2011-01-01

    Motivation: Modern experimental methods provide substantial information on protein-DNA recognition. Studying arrangements of transcription factor binding sites (TFBSs) of interacting transcription factors (TFs) advances understanding

  1. Interference of transcription across H-NS binding sites and repression by H-NS.

    Science.gov (United States)

    Rangarajan, Aathmaja Anandhi; Schnetz, Karin

    2018-05-01

    Nucleoid-associated protein H-NS represses transcription by forming extended DNA-H-NS complexes. Repression by H-NS operates mostly at the level of transcription initiation. Less is known about how DNA-H-NS complexes interfere with transcription elongation. In vitro H-NS has been shown to enhance RNA polymerase pausing and to promote Rho-dependent termination, while in vivo inhibition of Rho resulted in a decrease of the genome occupancy by H-NS. Here we show that transcription directed across H-NS binding regions relieves H-NS (and H-NS/StpA) mediated repression of promoters in these regions. Further, we observed a correlation of transcription across the H-NS-bound region and de-repression. The data suggest that the transcribing RNA polymerase is able to remodel the H-NS complex and/or dislodge H-NS from the DNA and thus relieve repression. Such an interference of transcription and H-NS mediated repression may imply that poorly transcribed AT-rich loci are prone to be repressed by H-NS, while efficiently transcribed loci escape repression. © 2018 John Wiley & Sons Ltd.

  2. A transcript cleavage factor of Mycobacterium tuberculosis important for its survival.

    Directory of Open Access Journals (Sweden)

    Arnab China

    Full Text Available After initiation of transcription, a number of proteins participate during elongation and termination modifying the properties of the RNA polymerase (RNAP. Gre factors are one such group conserved across bacteria. They regulate transcription by projecting their N-terminal coiled-coil domain into the active center of RNAP through the secondary channel and stimulating hydrolysis of the newly synthesized RNA in backtracked elongation complexes. Rv1080c is a putative gre factor (MtbGre in the genome of Mycobacterium tuberculosis. The protein enhanced the efficiency of promoter clearance by lowering abortive transcription and also rescued arrested and paused elongation complexes on the GC rich mycobacterial template. Although MtbGre is similar in domain organization and shares key residues for catalysis and RNAP interaction with the Gre factors of Escherichia coli, it could not complement an E. coli gre deficient strain. Moreover, MtbGre failed to rescue E. coli RNAP stalled elongation complexes, indicating the importance of specific protein-protein interactions for transcript cleavage. Decrease in the level of MtbGre reduced the bacterial survival by several fold indicating its essential role in mycobacteria. Another Gre homolog, Rv3788 was not functional in transcript cleavage activity indicating that a single Gre is sufficient for efficient transcription of the M. tuberculosis genome.

  3. New insights into the promoterless transcription of DNA coligo templates by RNA polymerase III.

    Science.gov (United States)

    Lama, Lodoe; Seidl, Christine I; Ryan, Kevin

    2014-01-01

    Chemically synthesized DNA can carry small RNA sequence information but converting that information into small RNA is generally thought to require large double-stranded promoters in the context of plasmids, viruses and genes. We previously found evidence that circularized oligodeoxynucleotides (coligos) containing certain sequences and secondary structures can template the synthesis of small RNA by RNA polymerase III in vitro and in human cells. By using immunoprecipitated RNA polymerase III we now report corroborating evidence that this enzyme is the sole polymerase responsible for coligo transcription. The immobilized polymerase enabled experiments showing that coligo transcripts can be formed through transcription termination without subsequent 3' end trimming. To better define the determinants of productive transcription, a structure-activity relationship study was performed using over 20 new coligos. The results show that unpaired nucleotides in the coligo stem facilitate circumtranscription, but also that internal loops and bulges should be kept small to avoid secondary transcription initiation sites. A polymerase termination sequence embedded in the double-stranded region of a hairpin-encoding coligo stem can antagonize transcription. Using lessons learned from new and old coligos, we demonstrate how to convert poorly transcribed coligos into productive templates. Our findings support the possibility that coligos may prove useful as chemically synthesized vectors for the ectopic expression of small RNA in human cells.

  4. p44 and p34 subunits of the BTF2/TFIIH transcription factor have homologies with SSL1, a yeast protein involved in DNA repair.

    NARCIS (Netherlands)

    S. Humbert; H. van Vuuren; Y. Lutz; J.H.J. Hoeijmakers (Jan); J-M. Egly (Jean-Marc); V. Moncollin

    1994-01-01

    textabstractThe human BTF2 (TFIIH) transcription factor is a multisubunit protein involved in transcription initiation by RNA polymerase II (B) as well as in DNA repair. In addition to the previously characterized p62 and p89/ERCC3 subunits, we have cloned two other subunits of BTF2, p44 and p34.

  5. Tall Buildings Initiative

    Science.gov (United States)

    Initiative 2017 TBI Guidelines Version 2.03 Now Available Screen Shot 2017-10-10 at 3.05.10 PM PEER has just initiative to develop design criteria that will ensure safe and usable tall buildings following future earthquakes. Download the primary product of this initiative: Guidelines for Performance-Based Seismic Design

  6. Lack of promoter IV-driven BDNF transcription results in depression-like behavior.

    Science.gov (United States)

    Sakata, K; Jin, L; Jha, S

    2010-10-01

    Transcription of Bdnf is controlled by multiple promoters, in which promoter IV contributes significantly to activity-dependent Bdnf transcription. We have generated promoter IV mutant mice [brain-derived neurotrophic factor (BDNF)-KIV] in which promoter IV-driven expression of BDNF is selectively disrupted by inserting a green fluorescent protein (GFP)-STOP cassette within the Bdnf exon IV locus. BDNF-KIV animals exhibited depression-like behavior as shown by the tail suspension test (TST), sucrose preference test (SPT) and learned helplessness test (LHT). In addition, BDNF-KIV mice showed reduced activity in the open field test (OFT) and reduced food intake in the novelty-suppressed feeding test (NSFT). The mutant mice did not display anxiety-like behavior in the light and dark box test and elevated plus maze tests. Interestingly, the mutant mice showed defective response inhibition in the passive avoidance test (PAT) even though their learning ability was intact when measured with the active avoidance test (AAT). These results suggest that promoter IV-dependent BDNF expression plays a critical role in the control of mood-related behaviors. This is the first study that directly addressed the effects of endogenous promoter-driven expression of BDNF in depression-like behavior. © 2010 The Authors. Genes, Brain and Behavior © 2010 Blackwell Publishing Ltd and International Behavioural and Neural Genetics Society.

  7. Transcriptional interference networks coordinate the expression of functionally related genes clustered in the same genomic loci.

    Science.gov (United States)

    Boldogköi, Zsolt

    2012-01-01

    The regulation of gene expression is essential for normal functioning of biological systems in every form of life. Gene expression is primarily controlled at the level of transcription, especially at the phase of initiation. Non-coding RNAs are one of the major players at every level of genetic regulation, including the control of chromatin organization, transcription, various post-transcriptional processes, and translation. In this study, the Transcriptional Interference Network (TIN) hypothesis was put forward in an attempt to explain the global expression of antisense RNAs and the overall occurrence of tandem gene clusters in the genomes of various biological systems ranging from viruses to mammalian cells. The TIN hypothesis suggests the existence of a novel layer of genetic regulation, based on the interactions between the transcriptional machineries of neighboring genes at their overlapping regions, which are assumed to play a fundamental role in coordinating gene expression within a cluster of functionally linked genes. It is claimed that the transcriptional overlaps between adjacent genes are much more widespread in genomes than is thought today. The Waterfall model of the TIN hypothesis postulates a unidirectional effect of upstream genes on the transcription of downstream genes within a cluster of tandemly arrayed genes, while the Seesaw model proposes a mutual interdependence of gene expression between the oppositely oriented genes. The TIN represents an auto-regulatory system with an exquisitely timed and highly synchronized cascade of gene expression in functionally linked genes located in close physical proximity to each other. In this study, we focused on herpesviruses. The reason for this lies in the compressed nature of viral genes, which allows a tight regulation and an easier investigation of the transcriptional interactions between genes. However, I believe that the same or similar principles can be applied to cellular organisms too.

  8. Similarities in transcription factor IIIC subunits that bind to the posterior regions of internal promoters for RNA polymerase III

    OpenAIRE

    Matsutani Sachiko

    2004-01-01

    Abstract Background In eukaryotes, RNA polymerase III (RNAP III) transcribes the genes for small RNAs like tRNAs, 5S rRNA, and several viral RNAs, and short interspersed repetitive elements (SINEs). The genes for these RNAs and SINEs have internal promoters that consist of two regions. These two regions are called the A and B blocks. The multisubunit transcription factor TFIIIC is required for transcription initiation of RNAP III; in transcription of tRNAs, the B-block binding subunit of TFII...

  9. Characterization of herpes simplex virus 2 primary microRNA Transcript regulation.

    Science.gov (United States)

    Tang, Shuang; Bosch-Marce, Marta; Patel, Amita; Margolis, Todd P; Krause, Philip R

    2015-05-01

    In order to understand factors that may influence latency-associated transcription and latency-associated transcript (LAT) phenotypes, we studied the expression of the herpes simplex virus 2 (HSV-2) LAT-associated microRNAs (miRNAs). We mapped the transcription initiation sites of all three primary miRNA transcripts and identified the ICP4-binding sequences at the transcription initiation sites of both HSV-2 LAT (pri-miRNA for miR-I and miR-II, which target ICP34.5, and miR-III, which targets ICP0) and L/ST (a pri-miRNA for miR-I and miR-II) but not at that of the primary miR-H6 (for which the target is unknown). We confirmed activity of the putative HSV-2 L/ST promoter and found that ICP4 trans-activates the L/ST promoter when the ICP4-binding site at its transcription initiation site is mutated, suggesting that ICP4 may play a dual role in regulating transcription of L/ST and, consequently, of miR-I and miR-II. LAT exon 1 (containing LAT enhancer sequences), together with the LAT promoter region, comprises a bidirectional promoter required for the expression of both LAT-encoded miRNAs and miR-H6 in latently infected mouse ganglia. The ability of ICP4 to suppress ICP34.5-targeting miRNAs and to activate lytic viral genes suggests that ICP4 could play a key role in the switch between latency and reactivation. The HSV-2 LAT and viral miRNAs expressed in the LAT region are the most abundant viral transcripts during HSV latency. The balance between the expression of LAT and LAT-associated miRNAs and the expression of lytic viral transcripts from the opposite strand appears to influence whether individual HSV-infected neurons will be latently or productively infected. The outcome of neuronal infection may thus depend on regulation of gene expression of the corresponding primary miRNAs. In the present study, we characterize promoter sequences responsible for miRNA expression, including identification of the primary miRNA 5' ends and evaluation of ICP4 response. These

  10. Manuscript Transcription by Crowdsourcing: Transcribe Bentham

    Directory of Open Access Journals (Sweden)

    Martin Moyle

    2011-02-01

    Full Text Available Transcribe Bentham is testing the feasibility of outsourcing the work of manuscript transcription to members of the public. UCL Library Services holds 60,000 folios of manuscripts of the philosopher and jurist Jeremy Bentham (1748–1832. Transcribe Bentham will digitise 12,500 Bentham folios, and, through a wiki-based interface, allow volunteer transcribers to take temporary ownership of manuscript images and to create TEI-encoded transcription text for final approval by UCL experts. Approved transcripts will be stored and preserved, with the manuscript images, in UCL’s public Digital Collections repository. The project makes innovative use of traditional library material. It will stimulate public engagement with UCL’s scholarly archive collections and the challenges of palaeography and manuscript transcription; it will raise the profile of the work and thought of Jeremy Bentham; and it will create new digital resources for future use by professional researchers. Towards the end of the project, the transcription tool will be made available to other projects and services. This paper is based on a presentation given by the lead author at LIBER’s 39th Annual General Conference in Aarhus, Denmark, 2010.

  11. The transcript release factor PTRF augments ribosomal gene transcription by facilitating reinitiation of RNA polymerase I

    Czech Academy of Sciences Publication Activity Database

    Jansa, Petr; Burek, C.; Sander, E. E.; Grummt, I.

    2001-01-01

    Roč. 29, č. 2 (2001), s. 423-429 ISSN 0305-1048 Institutional research plan: CEZ:AV0Z5052915 Keywords : rDNA transcription * PTRF * transcription reinitiation Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 6.373, year: 2001

  12. Transcription-induced DNA supercoiling: New roles of intranucleosomal DNA loops in DNA repair and transcription.

    Science.gov (United States)

    Gerasimova, N S; Pestov, N A; Kulaeva, O I; Clark, D J; Studitsky, V M

    2016-05-26

    RNA polymerase II (Pol II) transcription through chromatin is accompanied by formation of small intranucleosomal DNA loops. Pol II captured within a small loop drives accumulation of DNA supercoiling, facilitating further transcription. DNA breaks relieve supercoiling and induce Pol II arrest, allowing detection of DNA damage hidden in chromatin structure.

  13. A novel mode for transcription inhibition mediated by PNA-induced R-loops with a model in vitro system.

    Science.gov (United States)

    D'Souza, Alicia D; Belotserkovskii, Boris P; Hanawalt, Philip C

    2018-02-01

    The selective inhibition of transcription of a chosen gene by an artificial agent has numerous applications. Usually, these agents are designed to bind a specific nucleotide sequence in the promoter or within the transcribed region of the chosen gene. However, since optimal binding sites might not exist within the gene, it is of interest to explore the possibility of transcription inhibition when the agent is designed to bind at other locations. One of these possibilities arises when an additional transcription initiation site (e.g. secondary promoter) is present upstream from the primary promoter of the target gene. In this case, transcription inhibition might be achieved by inducing the formation of an RNA-DNA hybrid (R-loop) upon transcription from the secondary promoter. The R-loop could extend into the region of the primary promoter, to interfere with promoter recognition by RNA polymerase and thereby inhibit transcription. As a sequence-specific R-loop-inducing agent, a peptide nucleic acid (PNA) could be designed to facilitate R-loop formation by sequestering the non-template DNA strand. To investigate this mode for transcription inhibition, we have employed a model system in which a PNA binding site is localized between the T3 and T7 phage RNA polymerase promoters, which respectively assume the roles of primary and secondary promoters. In accord with our model, we have demonstrated that with PNA-bound DNA substrates, transcription from the T7 promoter reduces transcription from the T3 promoter by 30-fold, while in the absence of PNA binding there is no significant effect of T7 transcription upon T3 transcription. Copyright © 2018 Elsevier B.V. All rights reserved.

  14. The Intertwined Roles of DNA Damage and Transcription

    OpenAIRE

    Di Palo, Giacomo

    2016-01-01

    DNA damage and transcription are two interconnected events. Transcription can induce damage and scheduled DNA damage can be required for transcription. Here, we analyzed genome-wide distribution of 8oxodG-marked oxidative DNA damage obtained by OxiDIP-Seq, and we found a correlation with transcription of protein coding genes.

  15. NF-kB activity-dependent P-selectin involved in ox-LDL-induced foam cell formation in U937 cell

    International Nuclear Information System (INIS)

    Wang, Yi; Wang, Xiang; Sun, Minghui; Zhang, Zhenyu; Cao, Heng; Chen, Xiaoqing

    2011-01-01

    Highlights: → Ox-LDL induced foam cell formation in the human U937 promonocytic cell line in a dose- and time-dependent manner. → Ox-LDL induced expression of P-selectin through degradation of IkBa and augment of NF-kB activity and protein level during macrophage-derived foam cell formation. → P-selectin and NF-kB may be identified as pivotal regulators of ox-LDL-induced foam cell formation. → Therapy based on the inhibition of P-selectin and NF-kB may complement conventional treatments to prevent atherosclerosis. -- Abstract: Oxidized low-density lipoprotein (ox-LDL) plays a critical role in regulation of atherosclerosis. However, little is known about the role of Nuclear factor kB (NF-kB) activity-dependent P-selectin in ox-LDL-induced foam cell formation during atherosclerosis. In this study, we first investigated ox-LDL induced foam cell formation in the human U937 promonocytic cell line in a dose- and time-dependent manner. Treatment of U937 cells with ox-LDL increased lipid accumulation as well as intracellular cholesterol content. Next, a comparative analysis of gene expression profiling using cDNA microarray and Real-time-PCR indicated that ox-LDL exposure induced, in three treated groups, an extremely marked increase in the mRNA level of P-selectin. Protein levels of P-selectin and its upstream regulators IkBa and NF-kB showed that NF-kB pathway is involved in the ox-LDL-induced foam cell formation. Finally, overexpression of NF-kB significantly accelerated, whereas, inhibition of NF-kB with siRNA remarkably attenuated ox-LDL-induced macrophage-derived foam cell formation. It was concluded that the activity of NF-kB is augmented during macrophage-derived foam cell formation. Activation of NF-kB increased, whereas, inhibition of NF-kB decreased ox-LDL-induced P-selectin expression and lipid accumulation in macrophages, suggesting ox-LDL induced expression of P-selectin through degradation of IkBa and activation of NF-kB in the regulation of foam

  16. Soluble ICAM-5, a product of activity dependent proteolysis, increases mEPSC frequency and dendritic expression of GluA1.

    Directory of Open Access Journals (Sweden)

    Irina Lonskaya

    Full Text Available Matrix metalloproteinases (MMPs are zinc dependent endopeptidases that can be released from neurons in an activity dependent manner to play a role in varied forms of learning and memory. MMP inhibitors impair hippocampal long term potentiation (LTP, spatial memory, and behavioral correlates of drug addiction. Since MMPs are thought to influence LTP through a β1 integrin dependent mechanism, it has been suggested that these enzymes cleave specific substrates to generate integrin binding ligands. In previously published work, we have shown that neuronal activity stimulates rapid MMP dependent shedding of intercellular adhesion molecule-5 (ICAM-5, a synaptic adhesion molecule expressed on dendrites of the telencephalon. We have also shown that the ICAM-5 ectodomain can interact with β1 integrins to stimulate integrin dependent phosphorylation of cofilin, an event that occurs with dendritic spine maturation and LTP. In the current study, we investigate the potential for the ICAM-5 ectodomain to stimulate changes in α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate receptor (AMPAR dependent glutamatergic transmission. Single cell recordings show that the ICAM-5 ectodomain stimulates an increase in the frequency, but not the amplitude, of AMPA mini excitatory post synaptic currents (mEPSCs. With biotinylation and precipitation assays, we also show that the ICAM-5 ectodomain stimulates an increase in membrane levels of GluA1, but not GluA2, AMPAR subunits. In addition, we observe an ICAM-5 associated increase in GluA1 phosphorylation at serine 845. Concomitantly, ICAM-5 affects an increase in GluA1 surface staining along dendrites without affecting an increase in dendritic spine number. Together these data are consistent with the possibility that soluble ICAM-5 increases glutamatergic transmission and that post-synaptic changes, including increased phosphorylation and dendritic insertion of GluA1, could contribute. We suggest that future studies

  17. NF-kB activity-dependent P-selectin involved in ox-LDL-induced foam cell formation in U937 cell

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yi, E-mail: wangyi2004a@126.com [Department of Cardiology, Shanghai First People' s Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200080 (China); Wang, Xiang; Sun, Minghui; Zhang, Zhenyu; Cao, Heng; Chen, Xiaoqing [Department of Cardiology, Shanghai First People' s Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200080 (China)

    2011-08-05

    Highlights: {yields} Ox-LDL induced foam cell formation in the human U937 promonocytic cell line in a dose- and time-dependent manner. {yields} Ox-LDL induced expression of P-selectin through degradation of IkBa and augment of NF-kB activity and protein level during macrophage-derived foam cell formation. {yields} P-selectin and NF-kB may be identified as pivotal regulators of ox-LDL-induced foam cell formation. {yields} Therapy based on the inhibition of P-selectin and NF-kB may complement conventional treatments to prevent atherosclerosis. -- Abstract: Oxidized low-density lipoprotein (ox-LDL) plays a critical role in regulation of atherosclerosis. However, little is known about the role of Nuclear factor kB (NF-kB) activity-dependent P-selectin in ox-LDL-induced foam cell formation during atherosclerosis. In this study, we first investigated ox-LDL induced foam cell formation in the human U937 promonocytic cell line in a dose- and time-dependent manner. Treatment of U937 cells with ox-LDL increased lipid accumulation as well as intracellular cholesterol content. Next, a comparative analysis of gene expression profiling using cDNA microarray and Real-time-PCR indicated that ox-LDL exposure induced, in three treated groups, an extremely marked increase in the mRNA level of P-selectin. Protein levels of P-selectin and its upstream regulators IkBa and NF-kB showed that NF-kB pathway is involved in the ox-LDL-induced foam cell formation. Finally, overexpression of NF-kB significantly accelerated, whereas, inhibition of NF-kB with siRNA remarkably attenuated ox-LDL-induced macrophage-derived foam cell formation. It was concluded that the activity of NF-kB is augmented during macrophage-derived foam cell formation. Activation of NF-kB increased, whereas, inhibition of NF-kB decreased ox-LDL-induced P-selectin expression and lipid accumulation in macrophages, suggesting ox-LDL induced expression of P-selectin through degradation of IkBa and activation of NF-kB in the

  18. Novel core promoter elements and a cognate transcription factor in the divergent unicellular eukaryote Trichomonas vaginalis.

    Science.gov (United States)

    Smith, Alias J; Chudnovsky, Lorissa; Simoes-Barbosa, Augusto; Delgadillo-Correa, Maria G; Jonsson, Zophonias O; Wohlschlegel, James A; Johnson, Patricia J

    2011-04-01

    A highly conserved DNA initiator (Inr) element has been the only core promoter element described in the divergent unicellular eukaryote Trichomonas vaginalis, although genome analyses reveal that only ∼75% of protein-coding genes appear to contain an Inr. In search of another core promoter element(s), a nonredundant database containing 5' untranslated regions of expressed T. vaginalis genes was searched for overrepresented DNA motifs and known eukaryotic core promoter elements. In addition to identifying the Inr, two elements that lack sequence similarity to the known protein-coding gene core promoter, motif 3 (M3) and motif 5 (M5), were identified. Mutational and functional analyses demonstrate that both are novel core promoter elements. M3 [(A/G/T)(A/G)C(G/C)G(T/C)T(T/A/G)] resembles a Myb recognition element (MRE) and is bound specifically by a unique protein with a Myb-like DNA binding domain. The M5 element (CCTTT) overlaps the transcription start site and replaces the Inr as an alternative, gene-specific initiator element. Transcription specifically initiates at the second cytosine within M5, in contrast to characteristic initiation by RNA polymerase II at an adenosine. In promoters that combine M3 with either M5 or Inr, transcription initiation is regulated by the M3 motif.

  19. Novel Core Promoter Elements and a Cognate Transcription Factor in the Divergent Unicellular Eukaryote Trichomonas vaginalis▿

    Science.gov (United States)

    Smith, Alias J.; Chudnovsky, Lorissa; Simoes-Barbosa, Augusto; Delgadillo-Correa, Maria G.; Jonsson, Zophonias O.; Wohlschlegel, James A.; Johnson, Patricia J.

    2011-01-01

    A highly conserved DNA initiator (Inr) element has been the only core promoter element described in the divergent unicellular eukaryote Trichomonas vaginalis, although genome analyses reveal that only ∼75% of protein-coding genes appear to contain an Inr. In search of another core promoter element(s), a nonredundant database containing 5′ untranslated regions of expressed T. vaginalis genes was searched for overrepresented DNA motifs and known eukaryotic core promoter elements. In addition to identifying the Inr, two elements that lack sequence similarity to the known protein-coding gene core promoter, motif 3 (M3) and motif 5 (M5), were identified. Mutational and functional analyses demonstrate that both are novel core promoter elements. M3 [(A/G/T)(A/G)C(G/C)G(T/C)T(T/A/G)] resembles a Myb recognition element (MRE) and is bound specifically by a unique protein with a Myb-like DNA binding domain. The M5 element (CCTTT) overlaps the transcription start site and replaces the Inr as an alternative, gene-specific initiator element. Transcription specifically initiates at the second cytosine within M5, in contrast to characteristic initiation by RNA polymerase II at an adenosine. In promoters that combine M3 with either M5 or Inr, transcription initiation is regulated by the M3 motif. PMID:21245378

  20. Battles and hijacks: Noncoding transcription in plants

    KAUST Repository

    Ariel, Federico

    2015-06-01

    Noncoding RNAs have emerged as major components of the eukaryotic transcriptome. Genome-wide analyses revealed the existence of thousands of long noncoding RNAs (lncRNAs) in several plant species. Plant lncRNAs are transcribed by the plant-specific RNA polymerases Pol IV and Pol V, leading to transcriptional gene silencing, as well as by Pol II. They are involved in a wide range of regulatory mechanisms impacting on gene expression, including chromatin remodeling, modulation of alternative splicing, fine-tuning of miRNA activity, and the control of mRNA translation or accumulation. Recently, dual noncoding transcription by alternative RNA polymerases was implicated in epigenetic and chromatin conformation dynamics. This review integrates the current knowledge on the regulatory mechanisms acting through plant noncoding transcription. © 2015 Elsevier Ltd.

  1. Phonemic Transcriptions in British and American Dictionaries

    Directory of Open Access Journals (Sweden)

    Rastislav Šuštaršič

    2005-06-01

    Full Text Available In view of recent criticisms concerning vowel symbols in some British English dictionaries (in particular by J. Windsor Lewis in JIPA (Windsor Lewis, 2003, with regard to the Oxford Dictionary of Pronunciation (Upton, 2001, this article extends the discussion on English phonemic transcriptions by including those that typically occur in standard American dictionaries, and by comparing the most common conventions of British and American dictionaries. In addition to symbols for both vowels and consonants, the paper also deals with the different representations of word accentuation and the issue of consistency regarding application of phonemic (systemic, broad, rather than phonetic (allophonic, narrow transcription. The different transcriptions are assessed from the points of view of their departures from the International Phonetic Alphabet, their overlapping with orthographic representation (spelling and their appropriateness in terms of reflecting actual pronunciation in standard British and/or American pronunciation.

  2. Crowdsourcing for quantifying transcripts: An exploratory study.

    Science.gov (United States)

    Azzam, Tarek; Harman, Elena

    2016-02-01

    This exploratory study attempts to demonstrate the potential utility of crowdsourcing as a supplemental technique for quantifying transcribed interviews. Crowdsourcing is the harnessing of the abilities of many people to complete a specific task or a set of tasks. In this study multiple samples of crowdsourced individuals were asked to rate and select supporting quotes from two different transcripts. The findings indicate that the different crowdsourced samples produced nearly identical ratings of the transcripts, and were able to consistently select the same supporting text from the transcripts. These findings suggest that crowdsourcing, with further development, can potentially be used as a mixed method tool to offer a supplemental perspective on transcribed interviews. Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. Transcription regulatory networks analysis using CAGE

    KAUST Repository

    Tegnér, Jesper N.

    2009-10-01

    Mapping out cellular networks in general and transcriptional networks in particular has proved to be a bottle-neck hampering our understanding of biological processes. Integrative approaches fusing computational and experimental technologies for decoding transcriptional networks at a high level of resolution is therefore of uttermost importance. Yet, this is challenging since the control of gene expression in eukaryotes is a complex multi-level process influenced by several epigenetic factors and the fine interplay between regulatory proteins and the promoter structure governing the combinatorial regulation of gene expression. In this chapter we review how the CAGE data can be integrated with other measurements such as expression, physical interactions and computational prediction of regulatory motifs, which together can provide a genome-wide picture of eukaryotic transcriptional regulatory networks at a new level of resolution. © 2010 by Pan Stanford Publishing Pte. Ltd. All rights reserved.

  4. Runx transcription factors in neuronal development

    Directory of Open Access Journals (Sweden)

    Shiga Takashi

    2008-08-01

    Full Text Available Abstract Runt-related (Runx transcription factors control diverse aspects of embryonic development and are responsible for the pathogenesis of many human diseases. In recent years, the functions of this transcription factor family in the nervous system have just begun to be understood. In dorsal root ganglion neurons, Runx1 and Runx3 play pivotal roles in the development of nociceptive and proprioceptive sensory neurons, respectively. Runx appears to control the transcriptional regulation of neurotrophin receptors, numerous ion channels and neuropeptides. As a consequence, Runx contributes to diverse aspects of the sensory system in higher vertebrates. In this review, we summarize recent progress in determining the role of Runx in neuronal development.

  5. Transcriptional inhibition by the retinoblastoma protein

    DEFF Research Database (Denmark)

    Fattaey, A; Helin, K; Harlow, E

    1993-01-01

    The retinoblastoma protein, pRB, appears to play a key role in coordinating the regulation of cell cycle position and transcriptional events. pRB undergoes specific cell-cycle-dependent phosphorylation, being underphosphorylated in G1 and heavily phosphorylated in S, G2, and M. The underphosphory......The retinoblastoma protein, pRB, appears to play a key role in coordinating the regulation of cell cycle position and transcriptional events. pRB undergoes specific cell-cycle-dependent phosphorylation, being underphosphorylated in G1 and heavily phosphorylated in S, G2, and M......-mediated transcription would be lost by mutation in the retinoblastoma gene in human tumours, by pRB's interaction with DNA tumour virus oncoproteins, or by phosphorylation during the cell cycle....

  6. Deciphering the Innate Lymphoid Cell Transcriptional Program

    Directory of Open Access Journals (Sweden)

    Cyril Seillet

    2016-10-01

    Full Text Available Innate lymphoid cells (ILCs are enriched at mucosal surfaces, where they provide immune surveillance. All ILC subsets develop from a common progenitor that gives rise to pre-committed progenitors for each of the ILC lineages. Currently, the temporal control of gene expression that guides the emergence of these progenitors is poorly understood. We used global transcriptional mapping to analyze gene expression in different ILC progenitors. We identified PD-1 to be specifically expressed in PLZF+ ILCp and revealed that the timing and order of expression of the transcription factors NFIL3, ID2, and TCF-1 was critical. Importantly, induction of ILC lineage commitment required only transient expression of NFIL3 prior to ID2 and TCF-1 expression. These findings highlight the importance of the temporal program that permits commitment of progenitors to the ILC lineage, and they expand our understanding of the core transcriptional program by identifying potential regulators of ILC development.

  7. Transcription as a Threat to Genome Integrity.

    Science.gov (United States)

    Gaillard, Hélène; Aguilera, Andrés

    2016-06-02

    Genomes undergo different types of sporadic alterations, including DNA damage, point mutations, and genome rearrangements, that constitute the basis for evolution. However, these changes may occur at high levels as a result of cell pathology and trigger genome instability, a hallmark of cancer and a number of genetic diseases. In the last two decades, evidence has accumulated that transcription constitutes an important natural source of DNA metabolic errors that can compromise the integrity of the genome. Transcription can create the conditions for high levels of mutations and recombination by its ability to open the DNA structure and remodel chromatin, making it more accessible to DNA insulting agents, and by its ability to become a barrier to DNA replication. Here we review the molecular basis of such events from a mechanistic perspective with particular emphasis on the role of transcription as a genome instability determinant.

  8. Molecular imaging of transcriptional regulation during inflammation

    Directory of Open Access Journals (Sweden)

    Carlsen Harald

    2010-04-01

    Full Text Available Abstract Molecular imaging enables non-invasive visualization of the dynamics of molecular processes within living organisms in vivo. Different imaging modalities as MRI, SPECT, PET and optic imaging are used together with molecular probes specific for the biological process of interest. Molecular imaging of transcription factor activity is done in animal models and mostly in transgenic reporter mice, where the transgene essentially consists of a promoter that regulates a reporter gene. During inflammation, the transcription factor NF-κB is widely involved in orchestration and regulation of the immune system and almost all imaging studies in this field has revolved around the role and regulation of NF-κB. We here present a brief introduction to experimental use and design of transgenic reporter mice and a more extensive review of the various studies where molecular imaging of transcriptional regulation has been applied during inflammation.

  9. Widespread anti-sense transcription in apple is correlated with siRNA production and indicates a large potential for transcriptional and/or post-transcriptional control.

    Science.gov (United States)

    Celton, Jean-Marc; Gaillard, Sylvain; Bruneau, Maryline; Pelletier, Sandra; Aubourg, Sébastien; Martin-Magniette, Marie-Laure; Navarro, Lionel; Laurens, François; Renou, Jean-Pierre

    2014-07-01

    Characterizing the transcriptome of eukaryotic organisms is essential for studying gene regulation and its impact on phenotype. The realization that anti-sense (AS) and noncoding RNA transcription is pervasive in many genomes has emphasized our limited understanding of gene transcription and post-transcriptional regulation. Numerous mechanisms including convergent transcription, anti-correlated expression of sense and AS transcripts, and RNAi remain ill-defined. Here, we have combined microarray analysis and high-throughput sequencing of small RNAs (sRNAs) to unravel the complexity of transcriptional and potential post-transcriptional regulation in eight organs of apple (Malus × domestica). The percentage of AS transcript expression is higher than that identified in annual plants such as rice and Arabidopsis thaliana. Furthermore, we show that a majority of AS transcripts are transcribed beyond 3'UTR regions, and may cover a significant portion of the predicted sense transcripts. Finally we demonstrate at a genome-wide scale that anti-sense transcript expression is correlated with the presence of both short (21-23 nt) and long (> 30 nt) siRNAs, and that the sRNA coverage depth varies with the level of AS transcript expression. Our study provides a new insight on the functional role of anti-sense transcripts at the genome-wide level, and a new basis for the understanding of sRNA biogenesis in plants. © 2014 INRA. New Phytologist © 2014 New Phytologist Trust.

  10. The global transcriptional response of fission yeast to hydrogen sulfide.

    Directory of Open Access Journals (Sweden)

    Xu Jia

    Full Text Available BACKGROUND: Hydrogen sulfide (H(2S is a newly identified member of the small family of gasotransmitters that are endogenous gaseous signaling molecules that have a fundamental role in human biology and disease. Although it is a relatively recent discovery and the mechanism of H(2S activity is not completely understood, it is known to be involved in a number of cellular processes; H(2S can affect ion channels, transcription factors and protein kinases in mammals. METHODOLOGY/PRINCIPAL FINDINGS: In this paper, we have used fission yeast as a model organism to study the global gene expression profile in response to H(2S by microarray. We initially measured the genome-wide transcriptional response of fission yeast to H(2S. Through the functional classification of genes whose expression profile changed in response to H(2S, we found that H(2S mainly influences genes that encode putative or known stress proteins, membrane transporters, cell cycle/meiotic proteins, transcription factors and respiration protein in the mitochondrion. Our analysis showed that there was a significant overlap between the genes affected by H(2S and the stress response. We identified that the target genes of the MAPK pathway respond to H(2S; we also identified that a number of transporters respond to H(2S, these include sugar/carbohydrate transporters, ion transporters, and amino acid transporters. We found many mitochondrial genes to be down regulated upon H(2S treatment and that H(2S can reduce mitochondrial oxygen consumption. CONCLUSION/SIGNIFICANCE: This study identifies potential molecular targets of the signaling molecule H(2S in fission yeast and provides clues about the identity of homologues human proteins and will further the understanding of the cellular role of H(2S in human diseases.

  11. Leaderless Transcripts and Small Proteins Are Common Features of the Mycobacterial Translational Landscape.

    Directory of Open Access Journals (Sweden)

    Scarlet S Shell

    2015-11-01

    Full Text Available RNA-seq technologies have provided significant insight into the transcription networks of mycobacteria. However, such studies provide no definitive information on the translational landscape. Here, we use a combination of high-throughput transcriptome and proteome-profiling approaches to more rigorously understand protein expression in two mycobacterial species. RNA-seq and ribosome profiling in Mycobacterium smegmatis, and transcription start site (TSS mapping and N-terminal peptide mass spectrometry in Mycobacterium tuberculosis, provide complementary, empirical datasets to examine the congruence of transcription and translation in the Mycobacterium genus. We find that nearly one-quarter of mycobacterial transcripts are leaderless, lacking a 5' untranslated region (UTR and Shine-Dalgarno ribosome-binding site. Our data indicate that leaderless translation is a major feature of mycobacterial genomes and is comparably robust to leadered initiation. Using translational reporters to systematically probe the cis-sequence requirements of leaderless translation initiation in mycobacteria, we find that an ATG or GTG at the mRNA 5' end is both necessary and sufficient. This criterion, together with our ribosome occupancy data, suggests that mycobacteria encode hundreds of small, unannotated proteins at the 5' ends of transcripts. The conservation of small proteins in both mycobacterial species tested suggests that some play important roles in mycobacterial physiology. Our translational-reporter system further indicates that mycobacterial leadered translation initiation requires a Shine Dalgarno site in the 5' UTR and that ATG, GTG, TTG, and ATT codons can robustly initiate translation. Our combined approaches provide the first comprehensive view of mycobacterial gene structures and their non-canonical mechanisms of protein expression.

  12. Transcription and the aspect ratio of DNA

    DEFF Research Database (Denmark)

    Olsen, Kasper Wibeck; Bohr, Jakob

    2013-01-01

    analysis of transcription. It is shown that under certain reasonable assumptions transcription is only possible if the aspect ratio is in the regime corresponding to further twisting. We find this constraint to be in agreement with long-established crystallographic studies of DNA.......Two separate regimes exist for the aspect ratio of DNA. A low aspect regime where DNA will twist further under strain and a high aspect regime where DNA will untwist under strain. The question of the overall geometry, i.e. the aspect ratio, of DNA is revisited from the perspective of a geometrical...

  13. Supplemental figures for ``Constraints on jet quenching in p-Pb collisions at $\\sqrt{s_{\\rm NN}} =5.02$ TeV measured by the event-activity dependence of semi-inclusive hadron-jet distributions''

    CERN Document Server

    2017-01-01

    This note presents supplemental figures that support the analysis and discussion in the ALICE paper ``Constraints on jet quenching in p-Pb collisions at $\\sqrt{s_{\\rm NN}} =5.02$ TeV measured by the event-activity dependence of semi-inclusive hadron-jet distributions"

  14. Emerging Pathogens Initiative (EPI)

    Data.gov (United States)

    Department of Veterans Affairs — The Emerging Pathogens Initiative (EPI) database contains emerging pathogens information from the local Veterans Affairs Medical Centers (VAMCs). The EPI software...

  15. Mycobacterium tuberculosis Transcription Machinery: Ready To Respond to Host Attacks

    Science.gov (United States)

    Flentie, Kelly; Garner, Ashley L.

    2016-01-01

    Regulating responses to stress is critical for all bacteria, whether they are environmental, commensal, or pathogenic species. For pathogenic bacteria, successful colonization and survival in the host are dependent on adaptation to diverse conditions imposed by the host tissue architecture and the immune response. Once the bacterium senses a hostile environment, it must enact a change in physiology that contributes to the organism's survival strategy. Inappropriate responses have consequences; hence, the execution of the appropriate response is essential for survival of the bacterium in its niche. Stress responses are most often regulated at the level of gene expression and, more specifically, transcription. This minireview focuses on mechanisms of regulating transcription initiation that are required by Mycobacterium tuberculosis to respond to the arsenal of defenses imposed by the host during infection. In particular, we highlight how certain features of M. tuberculosis physiology allow this pathogen to respond swiftly and effectively to host defenses. By enacting highly integrated and coordinated gene expression changes in response to stress, M. tuberculosis is prepared for battle against the host defense and able to persist within the human population. PMID:26883824

  16. Effect of ARA9 on dioxin receptor mediated transcription

    International Nuclear Information System (INIS)

    Lees, M.J.; Whitelaw, M.L.

    2002-01-01

    The dioxin (Aryl hydrocarbon) receptor (DR) is a unique bHLH transcription factor which is activated by binding of planar aromatic hydrocarbons typified by dioxin (TCDD). The active receptor is key to metabolism of aryl hydrocarbon xenobiotics by being a potent inducer of CYP1A1 gene activity. Chlorinated dioxins are inert to metabolism and initiate multifarious toxicities, including potent tumour promotion. These ill-effects are mediated by the activated DR and we are studying the mechanisms by which the ligand binding domain of the DR controls activity of the protein. The DR ligand binding domain resides within a PAS (Per/Arnt/Sim homology) region which is contiguous with the bHLH. The latent bHLH/PAS dioxin receptor (DR) is found in the cytoplasm of most mammalian cell types in a complex with heat shock protein 90, a novel immunophilin like protein termed ARA9/XAP2/AIP, and the co-chaperone p23. Here we use antisense ARA9 constructs to reveal that in the absence of ARA9, the DR is unable to form a transcriptionally active complex. Co-expression of antisense ARA9 with a form of the DR which is constitutively targeted to the nucleus leads to dramatically decreased levels of the nuclear DR protein, implying that ARA9 may function beyond its currently proposed role in cytoplasmic retention of the latent DR

  17. A critique on nuclear factor-kappa B and signal transducer and activator of transcription 3: The key transcription factors in periodontal pathogenesis

    Directory of Open Access Journals (Sweden)

    Ranjith Ambili

    2017-01-01

    Full Text Available Periodontal disease is initiated by microorganisms in dental plaque, and host immunoinflammatory response to the microbial challenge helps in disease progression. Conventional periodontal therapy was mainly targeted on the elimination of microbial component. However, a better understanding of molecular aspects in host response will enable the clinicians to formulate effective host modulation therapy (HMT for the periodontal management. Inflammatory mediators were the main targets for HMT in the past. Transcription factors can regulate the production of multiple mediators simultaneously, and inhibition of these factors will be more beneficial than blocking individual molecule. Two important transcription factors implicated in chronic inflammatory diseases are nuclear factor kappa B (NF-κB and signal transducers and activators of transcription 3. The role of these factors in periodontal disease is a less explored area. This comprehensive review is aimed at unveiling the critical role of NF-κB and signal transducers and activators of transcription 3 in periodontal pathogenesis. An online search was performed using MEDLINE/PubMed database. All publications till 2016 related to NF-κB, signal transducer and activator of transcription 3 (STAT3, and inflammation were included in writing this review. A total of 27,390 references were published based on the search terms used. Out of these, 507 were related to the periodontal research published in English till 2016. Relevant papers were chosen after carefully reading the abstract. This review has attempted to comprehend the existing knowledge regarding the role of transcription factors NF-κB and STAT3 in periodontal disease. Moreover, it also provides a connecting molecular link for the periodontal medicine concept.

  18. Transcriptional regulators of legume-rhizobia symbiosis: nuclear factors Ys and GRAS are two for tango.

    Science.gov (United States)

    Rípodas, Carolina; Clúa, Joaquín; Battaglia, Marina; Baudin, Maël; Niebel, Andreas; Zanetti, María Eugenia; Blanco, Flavio

    2014-01-01

    Transcription factors are DNA binding proteins that regulate gene expression. The nitrogen fixing symbiosis established between legume plants and soil bacteria is a complex interaction, in which plants need to integrate signals derived from the symbiont and the surrounding environment to initiate the developmental program of nodule organogenesis and the infection process. Several transcription factors that play critical roles in these processes have been reported in the past decade, including proteins of the GRAS and NF-Y families. Recently, we reported the characterization of a new GRAS domain containing-protein that interacts with a member of the C subunit of the NF-Y family, which plays an important role in nodule development and the progression of bacterial infection during the symbiotic interaction. The connection between transcription factors of these families highlights the significance of multimeric complexes in the fabulous capacity of plants to integrate and respond to multiple environmental stimuli.

  19. Subunit architecture and functional modular rearrangements of the transcriptional mediator complex.

    Science.gov (United States)

    Tsai, Kuang-Lei; Tomomori-Sato, Chieri; Sato, Shigeo; Conaway, Ronald C; Conaway, Joan W; Asturias, Francisco J

    2014-06-05

    The multisubunit Mediator, comprising ∼30 distinct proteins, plays an essential role in gene expression regulation by acting as a bridge between DNA-binding transcription factors and the RNA polymerase II (RNAPII) transcription machinery. Efforts to uncover the Mediator mechanism have been hindered by a poor understanding of its structure, subunit organization, and conformational rearrangements. By overcoming biochemical and image analysis hurdles, we obtained accurate EM structures of yeast and human Mediators. Subunit localization experiments, docking of partial X-ray structures, and biochemical analyses resulted in comprehensive mapping of yeast Mediator subunits and a complete reinterpretation of our previous Mediator organization model. Large-scale Mediator rearrangements depend on changes at the interfaces between previously described Mediator modules, which appear to be facilitated by factors conducive to transcription initiation. Conservation across eukaryotes of Mediator structure, subunit organization, and RNA polymerase II interaction suggest conservation of fundamental aspects of the Mediator mechanism. Copyright © 2014 Elsevier Inc. All rights reserved.

  20. Transcriptional regulation of the grape cytochrome P450 monooxygenase gene CYP736B expression in response to Xylella fastidiosa infection

    Directory of Open Access Journals (Sweden)

    Walker M Andrew

    2010-07-01

    Full Text Available Abstract Background Plant cytochrome P450 monooxygenases (CYP mediate synthesis and metabolism of many physiologically important primary and secondary compounds that are related to plant defense against a range of pathogenic microbes and insects. To determine if cytochrome P450 monooxygenases are involved in defense response to Xylella fastidiosa (Xf infection, we investigated expression and regulatory mechanisms of the cytochrome P450 monooxygenase CYP736B gene in both disease resistant and susceptible grapevines. Results Cloning of genomic DNA and cDNA revealed that the CYP736B gene was composed of two exons and one intron with GT as a donor site and AG as an acceptor site. CYP736B transcript was up-regulated in PD-resistant plants and down-regulated in PD-susceptible plants 6 weeks after Xf inoculation. However, CYP736B expression was very low in stem tissues at all evaluated time points. 5'RACE and 3'RACE sequence analyses revealed that there were three candidate transcription start sites (TSS in the upstream region and three candidate polyadenylation (PolyA sites in the downstream region of CYP736B. Usage frequencies of each transcription initiation site and each polyadenylation site varied depending on plant genotype, developmental stage, tissue, and treatment. These results demonstrate that expression of CYP736B is regulated developmentally and in response to Xf infection at both transcriptional and post-transcriptional levels. Multiple transcription start and polyadenylation sites contribute to regulation of CYP736B expression. Conclusions This report provides evidence that the cytochrome P450 monooxygenase CYP736B gene is involved in defense response at a specific stage of Xf infection in grapevines; multiple transcription initiation and polyadenylation sites exist for CYP736B in grapevine; and coordinative and selective use of transcription initiation and polyadenylation sites play an important role in regulation of CYP736B expression

  1. Mitochondrial transcription factor A protects human retinal ...

    African Journals Online (AJOL)

    Purpose: To investigate the impact of mitochondrial transcription factor A (TFAM), as a modulator of NF-κB, on proliferation of hypoxia-induced human retinal endothelial cell (HREC), and the probable mechanism. Methods: After exposure to hypoxia (1 % O2) for 5 days, cell proliferation and cell cycle of HREC were ...

  2. RNA Polymerase II–The Transcription Machine

    Indian Academy of Sciences (India)

    Home; Journals; Resonance – Journal of Science Education; Volume 12; Issue 3. RNA Polymerase II – The Transcription Machine - Nobel Prize in Chemistry 2006. Jiyoti Verma Aruna Naorem Anand Kumar Manimala Sen Parag Sadhale. General Article Volume 12 Issue 3 March 2007 pp 47-53 ...

  3. Cross-Family Transcription Factor Interactions

    NARCIS (Netherlands)

    Bemer, Marian; Dijk, van Aalt-Jan; Immink, Richard G.H.; Angenent, Gerco C.

    2017-01-01

    Specific and dynamic gene expression strongly depends on transcription factor (TF) activity and most plant TFs function in a combinatorial fashion. They can bind to DNA and control the expression of the corresponding gene in an additive fashion or cooperate by physical interactions, forming larger

  4. Corticosteroid receptors adopt distinct cyclical transcriptional signatures.

    Science.gov (United States)

    Le Billan, Florian; Amazit, Larbi; Bleakley, Kevin; Xue, Qiong-Yao; Pussard, Eric; Lhadj, Christophe; Kolkhof, Peter; Viengchareun, Say; Fagart, Jérôme; Lombès, Marc

    2018-05-07

    Mineralocorticoid receptors (MRs) and glucocorticoid receptors (GRs) are two closely related hormone-activated transcription factors that regulate major pathophysiologic functions. High homology between these receptors accounts for the crossbinding of their corresponding ligands, MR being activated by both aldosterone and cortisol and GR essentially activated by cortisol. Their coexpression and ability to bind similar DNA motifs highlight the need to investigate their respective contributions to overall corticosteroid signaling. Here, we decipher the transcriptional regulatory mechanisms that underlie selective effects of MRs and GRs on shared genomic targets in a human renal cellular model. Kinetic, serial, and sequential chromatin immunoprecipitation approaches were performed on the period circadian protein 1 ( PER1) target gene, providing evidence that both receptors dynamically and cyclically interact at the same target promoter in a specific and distinct transcriptional signature. During this process, both receptors regulate PER1 gene by binding as homo- or heterodimers to the same promoter region. Our results suggest a novel level of MR-GR target gene regulation, which should be considered for a better and integrated understanding of corticosteroid-related pathophysiology.-Le Billan, F., Amazit, L., Bleakley, K., Xue, Q.-Y., Pussard, E., Lhadj, C., Kolkhof, P., Viengchareun, S., Fagart, J., Lombès, M. Corticosteroid receptors adopt distinct cyclical transcriptional signatures.

  5. Ets transcription factor GABP controls T cell homeostasis and immunity.

    Science.gov (United States)

    Luo, Chong T; Osmanbeyoglu, Hatice U; Do, Mytrang H; Bivona, Michael R; Toure, Ahmed; Kang, Davina; Xie, Yuchen; Leslie, Christina S; Li, Ming O

    2017-10-20

    Peripheral T cells are maintained in the absence of vigorous stimuli, and respond to antigenic stimulation by initiating cell cycle progression and functional differentiation. Here we show that depletion of the Ets family transcription factor GA-binding protein (GABP) in T cells impairs T-cell homeostasis. In addition, GABP is critically required for antigen-stimulated T-cell responses in vitro and in vivo. Transcriptome and genome-wide GABP-binding site analyses identify GABP direct targets encoding proteins involved in cellular redox balance and DNA replication, including the Mcm replicative helicases. These findings show that GABP has a nonredundant role in the control of T-cell homeostasis and immunity.

  6. AC Initiation System.

    Science.gov (United States)

    An ac initiation system is described which uses three ac transmission signals interlocked for safety by frequency, phase, and power discrimination...The ac initiation system is pre-armed by the application of two ac signals have the proper phases, and activates a load when an ac power signal of the proper frequency and power level is applied. (Author)

  7. Sustainable Agricultural Marketing Initiatives

    Directory of Open Access Journals (Sweden)

    Hakan Adanacıoğlu

    2015-07-01

    Full Text Available Sustainable marketing is a holistic approach that puts equal emphasis on environmental, social equity, and economic concerns in the development of marketing strategies. The purpose of the study is to examine and discuss the sustainable agricultural marketing initiatives practiced throughout the World and Turkey, and to put forth suggestions to further improve the performance of agricultural marketing initiatives in Turkey. Some of the sustainable agricultural marketing initiatives practiced around the world are carried out through civil organizations. Furthermore; some of these initiatives have also launched by farmers, consumers, food processors and retailers. The long-term strategies to increase these initiatives should be determined due to the fact that examples of successful sustainable agricultural marketing initiatives are inadequate and cannot be spread in Turkey. In this context, first of all, the supports provided by the government to improve agricultural marketing systems, such as EU funds for rural development should be compatible with the goals of sustainable marketing. For this purpose, it should be examined whether all proposed projects related to agricultural marketing meet the social, economic, and environmental principles of sustainable marketing. It is important that supporting organizations, especially civil society organisations, should take an active role for faster dissemination and adoption of sustainable agricultural marketing practices in Turkey. These organizations may provide technical assistance in preparing successful project proposals and training to farm groups. In addition, the other organizations, such as local administrations, producers' associations, cooperatives, can contribute to the success of sustainable agricultural marketing initiatives. The use of direct marketing strategies and vertical integration attempts in sustainable agricultural marketing initiatives that will likely be implemented in Turkey is

  8. Eviction of linker histone H1 by NAP-family histone chaperones enhances activated transcription.

    Science.gov (United States)

    Zhang, Qian; Giebler, Holli A; Isaacson, Marisa K; Nyborg, Jennifer K

    2015-01-01

    In the Metazoan nucleus, core histones assemble the genomic DNA to form nucleosome arrays, which are further compacted into dense chromatin structures by the linker histone H1. The extraordinary density of chromatin creates an obstacle for accessing the genetic information. Regulation of chromatin dynamics is therefore critical to cellular homeostasis, and histone chaperones serve as prominent players in these processes. In the current study, we examined the role of specific histone chaperones in negotiating the inherently repressive chromatin structure during transcriptional activation. Using a model promoter, we demonstrate that the human nucleosome assembly protein family members hNap1 and SET/Taf1β stimulate transcription in vitro during pre-initiation complex formation, prior to elongation. This stimulatory effect is dependent upon the presence of activators, p300, and Acetyl-CoA. We show that transcription from our chromatin template is strongly repressed by H1, and that both histone chaperones enhance RNA synthesis by overcoming H1-induced repression. Importantly, both hNap1 and SET/Taf1β directly bind H1, and function to enhance transcription by evicting the linker histone from chromatin reconstituted with H1. In vivo studies demonstrate that SET/Taf1β, but not hNap1, strongly stimulates activated transcription from the chromosomally-integrated model promoter, consistent with the observation that SET/Taf1β is nuclear, whereas hNap1 is primarily cytoplasmic. Together, these observations indicate that SET/Taf1β may serve as a critical regulator of H1 dynamics and gene activation in vivo. These studies uncover a novel function for SET that mechanistically couples transcriptional derepression with H1 dynamics. Furthermore, they underscore the significance of chaperone-dependent H1 displacement as an essential early step in the transition of a promoter from a dense chromatin state into one that is permissive to transcription factor binding and robust

  9. Transcriptional regulation of long-term memory in the marine snail Aplysia

    Directory of Open Access Journals (Sweden)

    Lee Yong-Seok

    2008-06-01

    Full Text Available Abstract Whereas the induction of short-term memory involves only covalent modifications of constitutively expressed preexisting proteins, the formation of long-term memory requires gene expression, new RNA, and new protein synthesis. On the cellular level, transcriptional regulation is thought to be the starting point for a series of molecular steps necessary for both the initiation and maintenance of long-term synaptic facilitation (LTF. The core molecular features of transcriptional regulation involved in the long-term process are evolutionally conserved in Aplysia, Drosophila, and mouse, and indicate that gene regulation by the cyclic AMP response element binding protein (CREB acting in conjunction with different combinations of transcriptional factors is critical for the expression of many forms of long-term memory. In the marine snail Aplysia, the molecular mechanisms that underlie the storage of long-term memory have been extensively studied in the monosynaptic connections between identified sensory neuron and motor neurons of the gill-withdrawal reflex. One tail shock or one pulse of serotonin (5-HT, a modulatory transmitter released by tail shocks, produces a transient facilitation mediated by the cAMP-dependent protein kinase leading to covalent modifications in the sensory neurons that results in an enhancement of transmitter release and a strengthening of synaptic connections lasting minutes. By contrast, repeated pulses of 5-hydroxytryptamine (5-HT induce a transcription- and translation-dependent long-term facilitation (LTF lasting more than 24 h and trigger the activation of a family of transcription factors in the presynaptic sensory neurons including ApCREB1, ApCREB2 and ApC/EBP. In addition, we have recently identified novel transcription factors that modulate the expression of ApC/EBP and also are critically involved in LTF. In this review, we examine the roles of these transcription factors during consolidation of LTF induced

  10. Transcriptional regulation of long-term memory in the marine snail Aplysia.

    Science.gov (United States)

    Lee, Yong-Seok; Bailey, Craig H; Kandel, Eric R; Kaang, Bong-Kiun

    2008-06-17

    Whereas the induction of short-term memory involves only covalent modifications of constitutively expressed preexisting proteins, the formation of long-term memory requires gene expression, new RNA, and new protein synthesis. On the cellular level, transcriptional regulation is thought to be the starting point for a series of molecular steps necessary for both the initiation and maintenance of long-term synaptic facilitation (LTF). The core molecular features of transcriptional regulation involved in the long-term process are evolutionally conserved in Aplysia, Drosophila, and mouse, and indicate that gene regulation by the cyclic AMP response element binding protein (CREB) acting in conjunction with different combinations of transcriptional factors is critical for the expression of many forms of long-term memory. In the marine snail Aplysia, the molecular mechanisms that underlie the storage of long-term memory have been extensively studied in the monosynaptic connections between identified sensory neuron and motor neurons of the gill-withdrawal reflex. One tail shock or one pulse of serotonin (5-HT), a modulatory transmitter released by tail shocks, produces a transient facilitation mediated by the cAMP-dependent protein kinase leading to covalent modifications in the sensory neurons that results in an enhancement of transmitter release and a strengthening of synaptic connections lasting minutes. By contrast, repeated pulses of 5-hydroxytryptamine (5-HT) induce a transcription- and translation-dependent long-term facilitation (LTF) lasting more than 24 h and trigger the activation of a family of transcription factors in the presynaptic sensory neurons including ApCREB1, ApCREB2 and ApC/EBP. In addition, we have recently identified novel transcription factors that modulate the expression of ApC/EBP and also are critically involved in LTF. In this review, we examine the roles of these transcription factors during consolidation of LTF induced by different

  11. Dynamic Effects of Topoisomerase I Inhibition on R-Loops and Short Transcripts at Active Promoters.

    Directory of Open Access Journals (Sweden)

    Jessica Marinello

    Full Text Available Topoisomerase I-DNA-cleavage complexes (Top1cc stabilized by camptothecin (CPT have specific effects at transcriptional levels. We recently reported that Top1cc increase antisense transcript (aRNAs levels at divergent CpG-island promoters and, transiently, DNA/RNA hybrids (R-loop in nuclear and mitochondrial genomes of colon cancer HCT116 cells. However, the relationship between R-loops and aRNAs was not established. Here, we show that aRNAs can form R-loops in N-TERA-2 cells under physiological conditions, and that promoter-associated R-loops are somewhat increased and extended in length immediately upon cell exposure to CPT. In contrast, persistent Top1ccs reduce the majority of R-loops suggesting that CPT-accumulated aRNAs are not commonly involved in R-loops. The enhancement of aRNAs by Top1ccs is present both in human colon cancer HCT116 cells and WI38 fibroblasts suggesting a common response of cancer and normal cells. Although Top1ccs lead to DSB and DDR kinases activation, we do not detect a dependence of aRNA accumulation on ATM or DNA-PK activation. However, we showed that the cell response to persistent Top1ccs can involve an impairment of aRNA turnover rather than a higher synthesis rate. Finally, a genome-wide analysis shows that persistent Top1ccs also determine an accumulation of sense transcripts at 5'-end gene regions suggesting an increased occurrence of truncated transcripts. Taken together, the results indicate that Top1 may regulate transcription initiation by modulating RNA polymerase-generated negative supercoils, which can in turn favor R-loop formation at promoters, and that transcript accumulation at TSS is a response to persistent transcriptional stress by Top1 poisoning.

  12. Neural Progenitors Adopt Specific Identities by Directly Repressing All Alternative Progenitor Transcriptional Programs.

    Science.gov (United States)

    Kutejova, Eva; Sasai, Noriaki; Shah, Ankita; Gouti, Mina; Briscoe, James

    2016-03-21

    In the vertebrate neural tube, a morphogen-induced transcriptional network produces multiple molecularly distinct progenitor domains, each generating different neuronal subtypes. Using an in vitro differentiation system, we defined gene expression signatures of distinct progenitor populations and identified direct gene-regulatory inputs corresponding to locations of specific transcription factor binding. Combined with targeted perturbations of the network, this revealed a mechanism in which a progenitor identity is installed by active repression of the entire transcriptional programs of other neural progenitor fates. In the ventral neural tube, sonic hedgehog (Shh) signaling, together with broadly expressed transcriptional activators, concurrently activates the gene expression programs of several domains. The specific outcome is selected by repressive input provided by Shh-induced transcription factors that act as the key nodes in the network, enabling progenitors to adopt a single definitive identity from several initially permitted options. Together, the data suggest design principles relevant to many developing tissues. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  13. Regulation of cell proliferation by the E2F transcription factors

    DEFF Research Database (Denmark)

    Helin, K

    1998-01-01

    Experimental data generated in the past year have further emphasized the essential role for the E2F transcription factors in the regulation of cell proliferation. Genetic studies have shown that E2F activity is required for normal development in fruitflies, and the generation of E2F-1(-/-) mice h......Fs in the proteasomes. Novel target genes for the E2F transcription factors have been identified that link the E2Fs directly to the initiation of DNA replication.......Experimental data generated in the past year have further emphasized the essential role for the E2F transcription factors in the regulation of cell proliferation. Genetic studies have shown that E2F activity is required for normal development in fruitflies, and the generation of E2F-1(-/-) mice has...... demonstrated that individual members of the E2F transcription factor family are likely to have distinct roles in mammalian development and homeostasis. Additional mechanisms regulating the activity of the E2F transcription factors have been reported, including subcellular localization and proteolysis of the E2...

  14. Neuronal activity-regulated gene transcription: how are distant synaptic signals conveyed to the nucleus?

    Science.gov (United States)

    Matamales, Miriam

    2012-12-19

    Synaptic activity can trigger gene expression programs that are required for the stable change of neuronal properties, a process that is essential for learning and memory. Currently, it is still unclear how the stimulation of dendritic synapses can be coupled to transcription in the nucleus in a timely way given that large distances can separate these two cellular compartments. Although several mechanisms have been proposed to explain long distance communication between synapses and the nucleus, the possible co-existence of these models and their relevance in physiological conditions remain elusive. One model suggests that synaptic activation triggers the translocation to the nucleus of certain transcription regulators localised at postsynaptic sites that function as synapto-nuclear messengers. Alternatively, it has been hypothesised that synaptic activity initiates propagating regenerative intracellular calcium waves that spread through dendrites into the nucleus where nuclear transcription machinery is thereby regulated. It has also been postulated that membrane depolarisation of voltage-gated calcium channels on the somatic membrane is sufficient to increase intracellular calcium concentration and activate transcription without the need for transported signals from distant synapses. Here I provide a critical overview of the suggested mechanisms for coupling synaptic stimulation to transcription, the underlying assumptions behind them and their plausible physiological significance.

  15. Method to determine transcriptional regulation pathways in organisms

    Science.gov (United States)

    Gardner, Timothy S.; Collins, James J.; Hayete, Boris; Faith, Jeremiah

    2012-11-06

    The invention relates to computer-implemented methods and systems for identifying regulatory relationships between expressed regulating polypeptides and targets of the regulatory activities of such regulating polypeptides. More specifically, the invention provides a new method for identifying regulatory dependencies between biochemical species in a cell. In particular embodiments, provided are computer-implemented methods for identifying a regulatory interaction between a transcription factor and a gene target of the transcription factor, or between a transcription factor and a set of gene targets of the transcription factor. Further provided are genome-scale methods for predicting regulatory interactions between a set of transcription factors and a corresponding set of transcriptional target substrates thereof.

  16. 1996 environmental initiatives report

    International Nuclear Information System (INIS)

    1996-01-01

    Progress by Consumers Gas in addressing environmental challenges were reviewed. Proposed environmental initiatives for the next fiscal year and beyond were introduced. Proposed initiatives were placed into three priority categories, high, medium or low, which together with the environmental management framework form the the utility's overall environmental agenda. High on the list of environmental priorities for the company are atmospheric air emissions, planning and construction practices, energy conservation and efficiency, environmental compliance, and methane emissions. The present state of the initiatives by the various company divisions and regions, compiled from the respective business plans, were reported. 21 figs

  17. Interactome analysis of transcriptional coactivator multiprotein bridging factor 1 unveils a yeast AP-1-like transcription factor involved in oxidation tolerance of mycopathogen Beauveria bassiana.

    Science.gov (United States)

    Chu, Xin-Ling; Dong, Wei-Xia; Ding, Jin-Li; Feng, Ming-Guang; Ying, Sheng-Hua

    2018-02-01

    Oxidation tolerance is an important determinant to predict the virulence and biocontrol potential of Beauveria bassiana, a well-known entomopathogenic fungus. As a transcriptional coactivator, multiprotein bridging factor 1 mediates the activity of transcription factor in diverse physiological processes, and its homolog in B. bassiana (BbMBF1) contributes to fungal oxidation tolerance. In this study, the BbMBF1-interactomes under oxidative stress and normal growth condition were deciphered by mass spectrometry integrated with the immunoprecipitation. BbMBF1p factor has a broad interaction with proteins that are involved in various cellular processes, and this interaction is dynamically regulated by oxidative stress. Importantly, a B. bassiana homolog of yeast AP-1-like transcription factor (BbAP-1) was specifically associated with the BbMBF1-interactome under oxidation and significantly contributed to fungal oxidation tolerance. In addition, qPCR analysis revealed that several antioxidant genes are jointly controlled by BbAP-1 and BbMBF1. Conclusively, it is proposed that BbMBF1p protein mediates BbAP-1p factor to transcribe the downstream antioxidant genes in B. bassiana under oxidative stress. This study demonstrates for the first time a proteomic view of the MBF1-interactome in fungi, and presents an initial framework to probe the transcriptional mechanism involved in fungal response to oxidation, which will provide a new strategy to improve the biocontrol efficacy of B. bassiana.

  18. Activation-induced deoxycytidine deaminase (AID) co-transcriptional scanning at single-molecule resolution

    Science.gov (United States)

    Senavirathne, Gayan; Bertram, Jeffrey G.; Jaszczur, Malgorzata; Chaurasiya, Kathy R.; Pham, Phuong; Mak, Chi H.; Goodman, Myron F.; Rueda, David

    2015-12-01

    Activation-induced deoxycytidine deaminase (AID) generates antibody diversity in B cells by initiating somatic hypermutation (SHM) and class-switch recombination (CSR) during transcription of immunoglobulin variable (IgV) and switch region (IgS) DNA. Using single-molecule FRET, we show that AID binds to transcribed dsDNA and translocates unidirectionally in concert with RNA polymerase (RNAP) on moving transcription bubbles, while increasing the fraction of stalled bubbles. AID scans randomly when constrained in an 8 nt model bubble. When unconstrained on single-stranded (ss) DNA, AID moves in random bidirectional short slides/hops over the entire molecule while remaining bound for ~5 min. Our analysis distinguishes dynamic scanning from static ssDNA creasing. That AID alone can track along with RNAP during transcription and scan within stalled transcription bubbles suggests a mechanism by which AID can initiate SHM and CSR when properly regulated, yet when unregulated can access non-Ig genes and cause cancer.

  19. Discrete redox signaling pathways regulate photosynthetic light-harvesting and chloroplast gene transcription.

    Directory of Open Access Journals (Sweden)

    John F Allen

    Full Text Available In photosynthesis in chloroplasts, two related regulatory processes balance the actions of photosystems I and II. These processes are short-term, post-translational redistribution of light-harvesting capacity, and long-term adjustment of photosystem stoichiometry initiated by control of chloroplast DNA transcription. Both responses are initiated by changes in the redox state of the electron carrier, plastoquinone, which connects the two photosystems. Chloroplast Sensor Kinase (CSK is a regulator of transcription of chloroplast genes for reaction centres of the two photosystems, and a sensor of plastoquinone redox state. We asked whether CSK is also involved in regulation of absorbed light energy distribution by phosphorylation of light-harvesting complex II (LHC II. Chloroplast thylakoid membranes isolated from a CSK T-DNA insertion mutant and from wild-type Arabidopsis thaliana exhibit similar light- and redox-induced (32P-labelling of LHC II and changes in 77 K chlorophyll fluorescence emission spectra, while room-temperature chlorophyll fluorescence emission transients from Arabidopsis leaves are perturbed by inactivation of CSK. The results indicate indirect, pleiotropic effects of reaction centre gene transcription on regulation of photosynthetic light-harvesting in vivo. A single, direct redox signal is transmitted separately to discrete transcriptional and post-translational branches of an integrated cytoplasmic regulatory system.

  20. Post-transcription cleavage generates the 3' end of F17R transcripts in vaccinia virus

    International Nuclear Information System (INIS)

    D'Costa, Susan M.; Antczak, James B.; Pickup, David J.; Condit, Richard C.

    2004-01-01

    Most vaccinia virus intermediate and late mRNAs possess 3' ends that are extremely heterogeneous in sequence. However, late mRNAs encoding the cowpox A-type inclusion protein (ATI), the second largest subunit of the RNA polymerase, and the late telomeric transcripts possess homogeneous 3' ends. In the case of the ATI mRNA, it has been shown that the homogeneous 3' end is generated by a post-transcriptional endoribonucleolytic cleavage event. We have determined that the F17R gene also produces homogeneous transcripts generated by a post-transcriptional cleavage event. Mapping of in vivo mRNA shows that the major 3' end of the F17R transcript maps 1262 nt downstream of the F17R translational start site. In vitro transcripts spanning the in vivo 3' end are cleaved in an in vitro reaction using extracts from virus infected cells, and the site of cleavage is the same both in vivo and in vitro. Cleavage is not observed using extract from cells infected in the presence of hydroxyurea; therefore, the cleavage factor is either virus-coded or virus-induced during the post-replicative phase of virus replication. The cis-acting sequence responsible for cleavage is orientation specific and the factor responsible for cleavage activity has biochemical properties similar to the factor required for cleavage of ATI transcripts. Partially purified cleavage factor generates cleavage products of expected size when either the ATI or F17R substrates are used in vitro, strongly suggesting that cleavage of both transcripts is mediated by the same factor

  1. Cyclin D3 interacts with human activating transcription factor 5 and potentiates its transcription activity

    International Nuclear Information System (INIS)

    Liu Wenjin; Sun Maoyun; Jiang Jianhai; Shen Xiaoyun; Sun Qing; Liu Weicheng; Shen Hailian; Gu Jianxin

    2004-01-01

    The Cyclin D3 protein is a member of the D-type cyclins. Besides serving as cell cycle regulators, D-type cyclins have been reported to be able to interact with several transcription factors and modulate their transcriptional activations. Here we report that human activating transcription factor 5 (hATF5) is a new interacting partner of Cyclin D3. The interaction was confirmed by in vivo coimmunoprecipitation and in vitro binding analysis. Neither interaction between Cyclin D1 and hATF5 nor interaction between Cyclin D2 and hATF5 was observed. Confocal microscopy analysis showed that Cyclin D3 could colocalize with hATF5 in the nuclear region. Cyclin D3 could potentiate hATF5 transcriptional activity independently of its Cdk4 partner. But Cyclin D1 and Cyclin D2 had no effect on hATF5 transcriptional activity. These data provide a new clue to understand the new role of Cyclin D3 as a transcriptional regulator

  2. Transcriptional and post-transcriptional regulation of nucleotide excision repair genes in human cells

    Energy Technology Data Exchange (ETDEWEB)

    Lefkofsky, Hailey B. [Translational Oncology Program, University of Michigan Medical School, Ann Arbor, MI (United States); Veloso, Artur [Translational Oncology Program, University of Michigan Medical School, Ann Arbor, MI (United States); Department of Radiation Oncology, University of Michigan Medical School, Ann Arbor, MI (United States); Bioinformatics Program, Department of Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, MI (United States); Ljungman, Mats, E-mail: ljungman@umich.edu [Translational Oncology Program, University of Michigan Medical School, Ann Arbor, MI (United States); Department of Radiation Oncology, University of Michigan Medical School, Ann Arbor, MI (United States); Department of Environmental Health Sciences, School of Public Health, University of Michigan, Ann Arbor, MI (United States)

    2015-06-15

    Nucleotide excision repair (NER) removes DNA helix-distorting lesions induced by UV light and various chemotherapeutic agents such as cisplatin. These lesions efficiently block the elongation of transcription and need to be rapidly removed by transcription-coupled NER (TC-NER) to avoid the induction of apoptosis. Twenty-nine genes have been classified to code for proteins participating in nucleotide excision repair (NER) in human cells. Here we explored the transcriptional and post-transcriptional regulation of these NER genes across 13 human cell lines using Bru-seq and BruChase-seq, respectively. Many NER genes are relatively large in size and therefore will be easily inactivated by UV-induced transcription-blocking lesions. Furthermore, many of these genes produce transcripts that are rather unstable. Thus, these genes are expected to rapidly lose expression leading to a diminished function of NER. One such gene is ERCC6 that codes for the CSB protein critical for TC-NER. Due to its large gene size and high RNA turnover rate, the ERCC6 gene may act as dosimeter of DNA damage so that at high levels of damage, ERCC6 RNA levels would be diminished leading to the loss of CSB expression, inhibition of TC-NER and the promotion of cell death.

  3. Transcription of highly repetitive tandemly organized DNA in amphibians and birds: A historical overview and modern concepts.

    Science.gov (United States)

    Trofimova, Irina; Krasikova, Alla

    2016-12-01

    Tandemly organized highly repetitive DNA sequences are crucial structural and functional elements of eukaryotic genomes. Despite extensive evidence, satellite DNA remains an enigmatic part of the eukaryotic genome, with biological role and significance of tandem repeat transcripts remaining rather obscure. Data on tandem repeats transcription in amphibian and avian model organisms is fragmentary despite their genomes being thoroughly characterized. Review systematically covers historical and modern data on transcription of amphibian and avian satellite DNA in somatic cells and during meiosis when chromosomes acquire special lampbrush form. We highlight how transcription of tandemly repetitive DNA sequences is organized in interphase nucleus and on lampbrush chromosomes. We offer LTR-activation hypotheses of widespread satellite DNA transcription initiation during oogenesis. Recent explanations are provided for the significance of high-yield production of non-coding RNA derived from tandemly organized highly repetitive DNA. In many cases the data on the transcription of satellite DNA can be extrapolated from lampbrush chromosomes to interphase chromosomes. Lampbrush chromosomes with applied novel technical approaches such as superresolution imaging, chromosome microdissection followed by high-throughput sequencing, dynamic observation in life-like conditions provide amazing opportunities for investigation mechanisms of the satellite DNA transcription.

  4. A real-time view of the TAR:Tat:P-TEFb complex at HIV-1 transcription sites

    Directory of Open Access Journals (Sweden)

    Knezevich Anna

    2007-05-01

    Full Text Available Abstract HIV-1 transcription is tightly regulated: silent in long-term latency and highly active in acutely-infected cells. Transcription is activated by the viral protein Tat, which recruits the elongation factor P-TEFb by binding the TAR sequence present in nascent HIV-1 RNAs. In this study, we analyzed the dynamic of the TAR:Tat:P-TEFb complex in living cells, by performing FRAP experiments at HIV-1 transcription sites. Our results indicate that a large fraction of Tat present at these sites is recruited by Cyclin T1. We found that in the presence of Tat, Cdk9 remained bound to nascent HIV-1 RNAs for 71s. In contrast, when transcription was activated by PMA/ionomycin, in the absence of Tat, Cdk9 turned-over rapidly and resided on the HIV-1 promoter for only 11s. Thus, the mechanism of trans-activation determines the residency time of P-TEFb at the HIV-1 gene, possibly explaining why Tat is such a potent transcriptional activator. In addition, we observed that Tat occupied HIV-1 transcription sites for 55s, suggesting that the TAR:Tat:P-TEFb complex dissociates from the polymerase following transcription initiation, and undergoes subsequent cycles of association/dissociation.

  5. Research Programs & Initiatives

    Science.gov (United States)

    CGH develops international initiatives and collaborates with other NCI divisions, NCI-designated Cancer Centers, and other countries to support cancer control planning, encourage capacity building, and support cancer research and research networks.

  6. Nursing Home Quality Initiative

    Data.gov (United States)

    U.S. Department of Health & Human Services — This Nursing Home Quality Initiative (NHQI) website provides consumer and provider information regarding the quality of care in nursing homes. NHQI discusses quality...

  7. Surgical Critical Care Initiative

    Data.gov (United States)

    Federal Laboratory Consortium — The Surgical Critical Care Initiative (SC2i) is a USU research program established in October 2013 to develop, translate, and validate biology-driven critical care....

  8. Global Methane Initiative

    Science.gov (United States)

    The Global Methane Initiative promotes cost-effective, near-term methane recovery through partnerships between developed and developing countries, with participation from the private sector, development banks, and nongovernmental organizations.

  9. Medical Errors Reduction Initiative

    National Research Council Canada - National Science Library

    Mutter, Michael L

    2005-01-01

    The Valley Hospital of Ridgewood, New Jersey, is proposing to extend a limited but highly successful specimen management and medication administration medical errors reduction initiative on a hospital-wide basis...

  10. RAS Initiative - Community Outreach

    Science.gov (United States)

    Through community and technical collaborations, workshops and symposia, and the distribution of reference reagents, the RAS Initiative seeks to increase the sharing of knowledge and resources essential to defeating cancers caused by mutant RAS genes.

  11. RAS Initiative - Events

    Science.gov (United States)

    The NCI RAS Initiative has organized multiple events with outside experts to discuss how the latest scientific and technological breakthroughs can be applied to discover vulnerabilities in RAS-driven cancers.

  12. PESP Landscaping Initiative

    Science.gov (United States)

    Landscaping practices can positively or negatively affect local environments and human health. The Landscaping Initiative seeks to enhance benefits of landscaping while reducing need for pesticides, fertilizers, etc., by working with partners.

  13. About the RAS Initiative

    Science.gov (United States)

    The RAS Initiative, a "hub and spoke" model, connects researchers to better understand and target the more than 30% of cancers driven by mutations in RAS genes. Includes oversight and contact information.

  14. The Yekaterinburg headache initiative

    DEFF Research Database (Denmark)

    Lebedeva, Elena R; Olesen, Jes; Osipova, Vera V

    2013-01-01

    for a demonstrational interventional project in Russia, undertaken within the Global Campaign against Headache. The initiative proposes three actions: 1) raise awareness of need for improvement; 2) design and implement a three-tier model (from primary care to a single highly specialized centre with academic affiliation......) for efficient and equitable delivery of headache-related health care; 3) develop a range of educational initiatives aimed at primary-care physicians, non-specialist neurologists, pharmacists and the general public to support the second action. RESULTS AND CONCLUSION: We set these proposals in a context...... of a health-care needs assessment, and as a model for all Russia. We present and discuss early progress of the initiative, justify the investment of resources required for implementation and call for the political support that full implementation requires. The more that the Yekaterinburg headache initiative...

  15. The RAS Initiative

    Science.gov (United States)

    NCI established the RAS Initiative to explore innovative approaches for attacking the proteins encoded by mutant forms of RAS genes and to ultimately create effective, new therapies for RAS-related cancers.

  16. Piezoelectrically Initiated Pyrotechnic Igniter

    Science.gov (United States)

    Quince, Asia; Dutton, Maureen; Hicks, Robert; Burnham, Karen

    2013-01-01

    This innovation consists of a pyrotechnic initiator and piezoelectric initiation system. The device will be capable of being initiated mechanically; resisting initiation by EMF, RF, and EMI (electromagnetic field, radio frequency, and electromagnetic interference, respectively); and initiating in water environments and space environments. Current devices of this nature are initiated by the mechanical action of a firing pin against a primer. Primers historically are prone to failure. These failures are commonly known as misfires or hang-fires. In many cases, the primer shows the dent where the firing pin struck the primer, but the primer failed to fire. In devices such as "T" handles, which are commonly used to initiate the blowout of canopies, loss of function of the device may result in loss of crew. In devices such as flares or smoke generators, failure can result in failure to spot a downed pilot. The piezoelectrically initiated ignition system consists of a pyrotechnic device that plugs into a mechanical system (activator), which on activation, generates a high-voltage spark. The activator, when released, will strike a stack of electrically linked piezo crystals, generating a high-voltage, low-amperage current that is then conducted to the pyro-initiator. Within the initiator, an electrode releases a spark that passes through a pyrotechnic first-fire mixture, causing it to combust. The combustion of the first-fire initiates a primary pyrotechnic or explosive powder. If used in a "T" handle, the primary would ramp the speed of burn up to the speed of sound, generating a shock wave that would cause a high explosive to go "high order." In a flare or smoke generator, the secondary would produce the heat necessary to ignite the pyrotechnic mixture. The piezo activator subsystem is redundant in that a second stack of crystals would be struck at the same time with the same activation force, doubling the probability of a first strike spark generation. If the first

  17. Supply Chain Initiatives Database

    Energy Technology Data Exchange (ETDEWEB)

    None

    2012-11-01

    The Supply Chain Initiatives Database (SCID) presents innovative approaches to engaging industrial suppliers in efforts to save energy, increase productivity and improve environmental performance. This comprehensive and freely-accessible database was developed by the Institute for Industrial Productivity (IIP). IIP acknowledges Ecofys for their valuable contributions. The database contains case studies searchable according to the types of activities buyers are undertaking to motivate suppliers, target sector, organization leading the initiative, and program or partnership linkages.

  18. The "fourth dimension" of gene transcription.

    Science.gov (United States)

    O'Malley, Bert W

    2009-05-01

    The three dimensions of space provide our relationship to position on the earth, but the fourth dimension of time has an equally profound influence on our lives. Everything from light and sound to weather and biology operate on the principle of measurable temporal periodicity. Consequently, a wide variety of time clocks affect all aspects of our existence. The annual (and biannual) cycles of activity, metabolism, and mating, the monthly physiological clocks of women and men, and the 24-h diurnal rhythms of humans are prime examples. Should it be surprising to us that the fourth dimension also impinges upon gene expression and that the genome itself is regulated by the fastest running of all biological clocks? Recent evidence substantiates the existence of such a ubiquitin-dependent transcriptional clock that is based upon the activation and destruction of transcriptional coactivators.

  19. Deconstructing transcriptional heterogeneity in pluripotent stem cells

    Science.gov (United States)

    Shalek, Alex K.; Satija, Rahul; DaleyKeyser, AJay; Li, Hu; Zhang, Jin; Pardee, Keith; Gennert, David; Trombetta, John J.; Ferrante, Thomas C.; Regev, Aviv; Daley, George Q.; Collins, James J.

    2014-01-01

    SUMMARY Pluripotent stem cells (PSCs) are capable of dynamic interconversion between distinct substates, but the regulatory circuits specifying these states and enabling transitions between them are not well understood. We set out to characterize transcriptional heterogeneity in PSCs by single-cell expression profiling under different chemical and genetic perturbations. Signaling factors and developmental regulators show highly variable expression, with expression states for some variable genes heritable through multiple cell divisions. Expression variability and population heterogeneity can be influenced by perturbation of signaling pathways and chromatin regulators. Strikingly, either removal of mature miRNAs or pharmacologic blockage of signaling pathways drives PSCs into a low-noise ground state characterized by a reconfigured pluripotency network, enhanced self-renewal, and a distinct chromatin state, an effect mediated by opposing miRNA families acting on the c-myc / Lin28 / let-7 axis. These data illuminate the nature of transcriptional heterogeneity in PSCs. PMID:25471879

  20. Computational Investigations of Post-Transcriptional Regulation

    DEFF Research Database (Denmark)

    Rasmussen, Simon Horskjær

    and miRNA regulation was studied by cross-linking immunoprecipitation (CLIP) and RBP double knockdown experiments. A comprehensive analysis of 107 CLIP datasets of 49 RBPs demonstrated that RBPs modulate miRNA regulation. Results suggest it is mediated by RBP-binding hotspots that likely...... investigated using high-throughput data. Analysis of IMP RIP-seq, iCLIP and RNA-seq datasets identified transcripts associated with cytoplasmic IMP ribonucleoproteins. Many of these transcripts were functionally involved in actin cytoskeletal remodeling. Further analyses of this data permitted estimation...... of a bipartite motif, composed of an AU-rich and a CA-rich domain. In addition, a regulatory motif discovery method was developed and applied to identify motifs using differential expression data and CLIP-data in the above investigations. This thesis increased the understanding of the role of RBPs in mi...

  1. A Herpesviral Immediate Early Protein Promotes Transcription Elongation of Viral Transcripts.

    Science.gov (United States)

    Fox, Hannah L; Dembowski, Jill A; DeLuca, Neal A

    2017-06-13

    Herpes simplex virus 1 (HSV-1) genes are transcribed by cellular RNA polymerase II (RNA Pol II). While four viral immediate early proteins (ICP4, ICP0, ICP27, and ICP22) function in some capacity in viral transcription, the mechanism by which ICP22 functions remains unclear. We observed that the FACT complex (comprised of SSRP1 and Spt16) was relocalized in infected cells as a function of ICP22. ICP22 was also required for the association of FACT and the transcription elongation factors SPT5 and SPT6 with viral genomes. We further demonstrated that the FACT complex interacts with ICP22 throughout infection. We therefore hypothesized that ICP22 recruits cellular transcription elongation factors to viral genomes for efficient transcription elongation of viral genes. We reevaluated the phenotype of an ICP22 mutant virus by determining the abundance of all viral mRNAs throughout infection by transcriptome sequencing (RNA-seq). The accumulation of almost all viral mRNAs late in infection was reduced compared to the wild type, regardless of kinetic class. Using chromatin immunoprecipitation sequencing (ChIP-seq), we mapped the location of RNA Pol II on viral genes and found that RNA Pol II levels on the bodies of viral genes were reduced in the ICP22 mutant compared to wild-type virus. In contrast, the association of RNA Pol II with transcription start sites in the mutant was not reduced. Taken together, our results indicate that ICP22 plays a role in recruiting elongation factors like the FACT complex to the HSV-1 genome to allow for efficient viral transcription elongation late in viral infection and ultimately infectious virion production. IMPORTANCE HSV-1 interacts with many cellular proteins throughout productive infection. Here, we demonstrate the interaction of a viral protein, ICP22, with a subset of cellular proteins known to be involved in transcription elongation. We determined that ICP22 is required to recruit the FACT complex and other transcription

  2. Reconstructing transcriptional regulatory networks through genomics data

    OpenAIRE

    Sun, Ning; Zhao, Hongyu

    2009-01-01

    One central problem in biology is to understand how gene expression is regulated under different conditions. Microarray gene expression data and other high throughput data have made it possible to dissect transcriptional regulatory networks at the genomics level. Owing to the very large number of genes that need to be studied, the relatively small number of data sets available, the noise in the data and the different natures of the distinct data types, network inference presents great challen...

  3. NUR TRANSCRIPTION FACTORS IN STRESS AND ADDICTION

    Directory of Open Access Journals (Sweden)

    Danae eCampos-Melo

    2013-12-01

    Full Text Available The Nur transcription factors Nur77 (NGFI-B, NR4A1, Nurr1 (NR4A2 and Nor-1 (NR4A3 are a sub-family of orphan members of the nuclear receptor superfamily. These transcription factors are products of immediate early genes, whose expression is rapidly and transiently induced in the central nervous system by several types of stimuli. Nur factors are present throughout the hypothalamus-pituitary-adrenal axis where are prominently induced in response to stress. Drugs of abuse and stress also induce the expression of Nur factors in nuclei of the motivation/reward circuit of the brain, indicating their participation in the process of drug addiction and in non-hypothalamic responses to stress. Repeated use of addictive drugs and chronic stress induce long-lasting dysregulation of the brain motivation/reward circuit, due to reprogramming of gene expression and enduring alterations in neuronal function. Here, we review the data supporting that Nur transcription factors are key players in the molecular basis of the dysregulation of neuronal circuits involved in chronic stress and addiction.

  4. The transcriptional regulatory network of Mycobacterium tuberculosis.

    Directory of Open Access Journals (Sweden)

    Joaquín Sanz

    Full Text Available Under the perspectives of network science and systems biology, the characterization of transcriptional regulatory (TR networks beyond the context of model organisms offers a versatile tool whose potential remains yet mainly unexplored. In this work, we present an updated version of the TR network of Mycobacterium tuberculosis (M.tb, which incorporates newly characterized transcriptional regulations coming from 31 recent, different experimental works available in the literature. As a result of the incorporation of these data, the new network doubles the size of previous data collections, incorporating more than a third of the entire genome of the bacterium. We also present an exhaustive topological analysis of the new assembled network, focusing on the statistical characterization of motifs significances and the comparison with other model organisms. The expanded M.tb transcriptional regulatory network, considering its volume and completeness, constitutes an important resource for diverse tasks such as dynamic modeling of gene expression and signaling processes, computational reliability determination or protein function prediction, being the latter of particular relevance, given that the function of only a small percent of the proteins of M.tb is known.

  5. Evolution of transcriptional enhancers and animal diversity

    Science.gov (United States)

    Rubinstein, Marcelo; de Souza, Flávio S. J.

    2013-01-01

    Deciphering the genetic bases that drive animal diversity is one of the major challenges of modern biology. Although four decades ago it was proposed that animal evolution was mainly driven by changes in cis-regulatory DNA elements controlling gene expression rather than in protein-coding sequences, only now are powerful bioinformatics and experimental approaches available to accelerate studies into how the evolution of transcriptional enhancers contributes to novel forms and functions. In the introduction to this Theme Issue, we start by defining the general properties of transcriptional enhancers, such as modularity and the coexistence of tight sequence conservation with transcription factor-binding site shuffling as different mechanisms that maintain the enhancer grammar over evolutionary time. We discuss past and current methods used to identify cell-type-specific enhancers and provide examples of how enhancers originate de novo, change and are lost in particular lineages. We then focus in the central part of this Theme Issue on analysing examples of how the molecular evolution of enhancers may change form and function. Throughout this introduction, we present the main findings of the articles, reviews and perspectives contributed to this Theme Issue that together illustrate some of the great advances and current frontiers in the field. PMID:24218630

  6. Curated compendium of human transcriptional biomarker data.

    Science.gov (United States)

    Golightly, Nathan P; Bell, Avery; Bischoff, Anna I; Hollingsworth, Parker D; Piccolo, Stephen R

    2018-04-17

    One important use of genome-wide transcriptional profiles is to identify relationships between transcription levels and patient outcomes. These translational insights can guide the development of biomarkers for clinical application. Data from thousands of translational-biomarker studies have been deposited in public repositories, enabling reuse. However, data-reuse efforts require considerable time and expertise because transcriptional data are generated using heterogeneous profiling technologies, preprocessed using diverse normalization procedures, and annotated in non-standard ways. To address this problem, we curated 45 publicly available, translational-biomarker datasets from a variety of human diseases. To increase the data's utility, we reprocessed the raw expression data using a uniform computational pipeline, addressed quality-control problems, mapped the clinical annotations to a controlled vocabulary, and prepared consistently structured, analysis-ready data files. These data, along with scripts we used to prepare the data, are available in a public repository. We believe these data will be particularly useful to researchers seeking to perform benchmarking studies-for example, to compare and optimize machine-learning algorithms' ability to predict biomedical outcomes.

  7. Adaptive evolution of transcription factor binding sites

    Directory of Open Access Journals (Sweden)

    Berg Johannes

    2004-10-01

    Full Text Available Abstract Background The regulation of a gene depends on the binding of transcription factors to specific sites located in the regulatory region of the gene. The generation of these binding sites and of cooperativity between them are essential building blocks in the evolution of complex regulatory networks. We study a theoretical model for the sequence evolution of binding sites by point mutations. The approach is based on biophysical models for the binding of transcription factors to DNA. Hence we derive empirically grounded fitness landscapes, which enter a population genetics model including mutations, genetic drift, and selection. Results We show that the selection for factor binding generically leads to specific correlations between nucleotide frequencies at different positions of a binding site. We demonstrate the possibility of rapid adaptive evolution generating a new binding site for a given transcription factor by point mutations. The evolutionary time required is estimated in terms of the neutral (background mutation rate, the selection coefficient, and the effective population size. Conclusions The efficiency of binding site formation is seen to depend on two joint conditions: the binding site motif must be short enough and the promoter region must be long enough. These constraints on promoter architecture are indeed seen in eukaryotic systems. Furthermore, we analyse the adaptive evolution of genetic switches and of signal integration through binding cooperativity between different sites. Experimental tests of this picture involving the statistics of polymorphisms and phylogenies of sites are discussed.

  8. Transcription of tandemly repetitive DNA: functional roles.

    Science.gov (United States)

    Biscotti, Maria Assunta; Canapa, Adriana; Forconi, Mariko; Olmo, Ettore; Barucca, Marco

    2015-09-01

    A considerable fraction of the eukaryotic genome is made up of satellite DNA constituted of tandemly repeated sequences. These elements are mainly located at centromeres, pericentromeres, and telomeres and are major components of constitutive heterochromatin. Although originally satellite DNA was thought silent and inert, an increasing number of studies are providing evidence on its transcriptional activity supporting, on the contrary, an unexpected dynamicity. This review summarizes the multiple structural roles of satellite noncoding RNAs at chromosome level. Indeed, satellite noncoding RNAs play a role in the establishment of a heterochromatic state at centromere and telomere. These highly condensed structures are indispensable to preserve chromosome integrity and genome stability, preventing recombination events, and ensuring the correct chromosome pairing and segregation. Moreover, these RNA molecules seem to be involved also in maintaining centromere identity and in elongation, capping, and replication of telomere. Finally, the abnormal variation of centromeric and pericentromeric DNA transcription across major eukaryotic lineages in stress condition and disease has evidenced the critical role that these transcripts may play and the potentially dire consequences for the organism.

  9. Repetitive Elements in Mycoplasma hyopneumoniae Transcriptional Regulation.

    Directory of Open Access Journals (Sweden)

    Amanda Malvessi Cattani

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

  10. Repetitive Elements in Mycoplasma hyopneumoniae Transcriptional Regulation.

    Science.gov (United States)

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

    2016-01-01

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

  11. Modulation of transcription factors by curcumin.

    Science.gov (United States)

    Shishodia, Shishir; Singh, Tulika; Chaturvedi, Madan M

    2007-01-01

    Curcumin is the active ingredient of turmeric that has been consumed as a dietary spice for ages. Turmeric is widely used in traditional Indian medicine to cure biliary disorders, anorexia, cough, diabetic wounds, hepatic disorders, rheumatism, and sinusitis. Extensive investigation over the last five decades has indicated that curcumin reduces blood cholesterol, prevents low-density lipoprotein oxidation, inhibits platelet aggregation, suppresses thrombosis and myocardial infarction, suppresses symptoms associated with type II diabetes, rheumatoid arthritis, multiple sclerosis, and Alzheimer's disease, inhibits HIV replication, enhances wound healing, protects from liver injury, increases bile secretion, protects from cataract formation, and protects from pulmonary toxicity and fibrosis. Evidence indicates that the divergent effects of curcumin are dependent on its pleiotropic molecular effects. These include the regulation of signal transduction pathways and direct modulation of several enzymatic activities. Most of these signaling cascades lead to the activation of transcription factors. Curcumin has been found to modulate the activity of several key transcription factors and, in turn, the cellular expression profiles. Curcumin has been shown to elicit vital cellular responses such as cell cycle arrest, apoptosis, and differentiation by activating a cascade of molecular events. In this chapter, we briefly review the effects of curcumin on transcription factors NF-KB, AP-1, Egr-1, STATs, PPAR-gamma, beta-catenin, nrf2, EpRE, p53, CBP, and androgen receptor (AR) and AR-related cofactors giving major emphasis to the molecular mechanisms of its action.

  12. Fatty Acid–Regulated Transcription Factors in the Liver

    Science.gov (United States)

    Jump, Donald B.; Tripathy, Sasmita; Depner, Christopher M.

    2014-01-01

    Fatty acid regulation of hepatic gene transcription was first reported in the early 1990s. Several transcription factors have been identified as targets of fatty acid regulation. This regulation is achieved by direct fatty acid binding to the transcription factor or by indirect mechanisms where fatty acids regulate signaling pathways controlling the expression of transcription factors or the phosphorylation, ubiquitination, or proteolytic cleavage of the transcription factor. Although dietary fatty acids are well-established regulators of hepatic transcription factors, emerging evidence indicates that endogenously generated fatty acids are equally important in controlling transcription factors in the context of glucose and lipid homeostasis. Our first goal in this review is to provide an up-to-date examination of the molecular and metabolic bases of fatty acid regulation of key transcription factors controlling hepatic metabolism. Our second goal is to link these mechanisms to nonalcoholic fatty liver disease (NAFLD), a growing health concern in the obese population. PMID:23528177

  13. To Your Health: NLM update transcript - Gun safety strategies

    Science.gov (United States)

    ... transcript040918.html To Your Health: NLM update Transcript Gun safety strategies : 04/09/2018 To use the ... on weekly topics. An evidence-based, public health gun safety strategy that is consistent with second amendment ...

  14. Determination of specificity influencing residues for key transcription factor families

    DEFF Research Database (Denmark)

    Patel, Ronak Y.; Garde, Christian; Stormo, Gary D.

    2015-01-01

    Transcription factors (TFs) are major modulators of transcription and subsequent cellular processes. The binding of TFs to specific regulatory elements is governed by their specificity. Considering the gap between known TFs sequence and specificity, specificity prediction frameworks are highly de...

  15. Theory of site-specific interactions of the combinatorial transcription factors with DNA

    International Nuclear Information System (INIS)

    Murugan, R

    2010-01-01

    We derive a functional relationship between the mean first passage time associated with the concurrent binding of multiple transcription factors (TFs) at their respective combinatorial cis-regulatory module sites (CRMs) and the number n of TFs involved in the regulation of the initiation of transcription of a gene of interest. Our results suggest that the overall search time τ s that is required by the n TFs to locate their CRMs which are all located on the same DNA chain scales with n as τ s ∼n α where α ∼ (2/5). When the jump size k that is associated with the dynamics of all the n TFs along DNA is higher than that of the critical jump size k c that scales with the size of DNA N as k c ∼ N 2/3 , we observe a similar power law scaling relationship and also the exponent α. When k c , α shows a strong dependence on both n and k. Apparently there is a critical number of combinatorial TFs n c ∼ 20 that is required to efficiently regulate the initiation of transcription of a given gene below which (2/5) 1. These results seem to be independent of the initial distances between the TFs and their corresponding CRMs and also suggest that the maximum number of TFs involved in a given combinatorial regulation of the initiation of transcription of a gene of interest seems to be restricted by the degree of condensation of the genomic DNA. The optimum number m opt of roadblock protein molecules per genome at which the search time associated with these n TFs to locate their binding sites is a minimum seems to scale as m opt ∼Ln α/2 where L is the sliding length of TFs whose maximum value seems to be such that L ≤ 10 4 bps for the E. coli bacterial genome.

  16. DNA Binding by the Ribosomal DNA Transcription Factor Rrn3 Is Essential for Ribosomal DNA Transcription*

    Science.gov (United States)

    Stepanchick, Ann; Zhi, Huijun; Cavanaugh, Alice H.; Rothblum, Katrina; Schneider, David A.; Rothblum, Lawrence I.

    2013-01-01

    The human homologue of yeast Rrn3 is an RNA polymerase I-associated transcription factor that is essential for ribosomal DNA (rDNA) transcription. The generally accepted model is that Rrn3 functions as a bridge between RNA polymerase I and the transcription factors bound to the committed template. In this model Rrn3 would mediate an interaction between the mammalian Rrn3-polymerase I complex and SL1, the rDNA transcription factor that binds to the core promoter element of the rDNA. In the course of studying the role of Rrn3 in recruitment, we found that Rrn3 was in fact a DNA-binding protein. Analysis of the sequence of Rrn3 identified a domain with sequence similarity to the DNA binding domain of heat shock transcription factor 2. Randomization, or deletion, of the amino acids in this region in Rrn3, amino acids 382–400, abrogated its ability to bind DNA, indicating that this domain was an important contributor to DNA binding by Rrn3. Control experiments demonstrated that these mutant Rrn3 constructs were capable of interacting with both rpa43 and SL1, two other activities demonstrated to be essential for Rrn3 function. However, neither of these Rrn3 mutants was capable of functioning in transcription in vitro. Moreover, although wild-type human Rrn3 complemented a yeast rrn3-ts mutant, the DNA-binding site mutant did not. These results demonstrate that DNA binding by Rrn3 is essential for transcription by RNA polymerase I. PMID:23393135

  17. DNA binding by the ribosomal DNA transcription factor rrn3 is essential for ribosomal DNA transcription.

    Science.gov (United States)

    Stepanchick, Ann; Zhi, Huijun; Cavanaugh, Alice H; Rothblum, Katrina; Schneider, David A; Rothblum, Lawrence I

    2013-03-29

    The human homologue of yeast Rrn3 is an RNA polymerase I-associated transcription factor that is essential for ribosomal DNA (rDNA) transcription. The generally accepted model is that Rrn3 functions as a bridge between RNA polymerase I and the transcription factors bound to the committed template. In this model Rrn3 would mediate an interaction between the mammalian Rrn3-polymerase I complex and SL1, the rDNA transcription factor that binds to the core promoter element of the rDNA. In the course of studying the role of Rrn3 in recruitment, we found that Rrn3 was in fact a DNA-binding protein. Analysis of the sequence of Rrn3 identified a domain with sequence similarity to the DNA binding domain of heat shock transcription factor 2. Randomization, or deletion, of the amino acids in this region in Rrn3, amino acids 382-400, abrogated its ability to bind DNA, indicating that this domain was an important contributor to DNA binding by Rrn3. Control experiments demonstrated that these mutant Rrn3 constructs were capable of interacting with both rpa43 and SL1, two other activities demonstrated to be essential for Rrn3 function. However, neither of these Rrn3 mutants was capable of functioning in transcription in vitro. Moreover, although wild-type human Rrn3 complemented a yeast rrn3-ts mutant, the DNA-binding site mutant did not. These results demonstrate that DNA binding by Rrn3 is essential for transcription by RNA polymerase I.

  18. Validation, automatic generation and use of broad phonetic transcriptions

    NARCIS (Netherlands)

    Bael, Cristophe Patrick Jan Van

    2007-01-01

    Broad phonetic transcriptions represent the pronunciation of words as strings of characters from specifically designed symbol sets. In everyday life, broad phonetic transcriptions are often used as aids to pronounce (foreign) words. In addition, broad phonetic transcriptions are often used for

  19. DNA residence time is a regulatory factor of transcription repression

    Science.gov (United States)

    Clauß, Karen; Popp, Achim P.; Schulze, Lena; Hettich, Johannes; Reisser, Matthias; Escoter Torres, Laura; Uhlenhaut, N. Henriette

    2017-01-01

    Abstract Transcription comprises a highly regulated sequence of intrinsically stochastic processes, resulting in bursts of transcription intermitted by quiescence. In transcription activation or repression, a transcription factor binds dynamically to DNA, with a residence time unique to each factor. Whether the DNA residence time is important in the transcription process is unclear. Here, we designed a series of transcription repressors differing in their DNA residence time by utilizing the modular DNA binding domain of transcription activator-like effectors (TALEs) and varying the number of nucleotide-recognizing repeat domains. We characterized the DNA residence times of our repressors in living cells using single molecule tracking. The residence times depended non-linearly on the number of repeat domains and differed by more than a factor of six. The factors provoked a residence time-dependent decrease in transcript level of the glucocorticoid receptor-activated gene SGK1. Down regulation of transcription was due to a lower burst frequency in the presence of long binding repressors and is in accordance with a model of competitive inhibition of endogenous activator binding. Our single molecule experiments reveal transcription factor DNA residence time as a regulatory factor controlling transcription repression and establish TALE-DNA binding domains as tools for the temporal dissection of transcription regulation. PMID:28977492

  20. DNA to DNA transcription might exist in eukaryotic cells

    OpenAIRE

    Li, Gao-De

    2016-01-01

    Till now, in biological sciences, the term, transcription, mainly refers to DNA to RNA transcription. But our recently published experimental findings obtained from Plasmodium falciparum strongly suggest the existence of DNA to DNA transcription in the genome of eukaryotic cells, which could shed some light on the functions of certain noncoding DNA in the human and other eukaryotic genomes.

  1. Transcription-associated quality control of mRNP

    DEFF Research Database (Denmark)

    Schmid, Manfred; Jensen, Torben Heick

    2013-01-01

    Although a prime purpose of transcription is to produce RNA, a substantial amount of transcript is nevertheless turned over very early in its lifetime. During transcription RNAs are matured by nucleases from longer precursors and activities are also employed to exert quality control over the RNA...

  2. International EUREKA: Initialization Segment

    International Nuclear Information System (INIS)

    1982-02-01

    The Initialization Segment creates the starting description of the uranium market. The starting description includes the international boundaries of trade, the geologic provinces, resources, reserves, production, uranium demand forecasts, and existing market transactions. The Initialization Segment is designed to accept information of various degrees of detail, depending on what is known about each region. It must transform this information into a specific data structure required by the Market Segment of the model, filling in gaps in the information through a predetermined sequence of defaults and built in assumptions. A principal function of the Initialization Segment is to create diagnostic messages indicating any inconsistencies in data and explaining which assumptions were used to organize the data base. This permits the user to manipulate the data base until such time the user is satisfied that all the assumptions used are reasonable and that any inconsistencies are resolved in a satisfactory manner

  3. Initiating events frequency determination

    International Nuclear Information System (INIS)

    Simic, Z.; Mikulicic, V.; Vukovic, I.

    2004-01-01

    The paper describes work performed for the Nuclear Power Station (NPS). Work is related to the periodic initiating events frequency update for the Probabilistic Safety Assessment (PSA). Data for all relevant NPS initiating events (IE) were reviewed. The main focus was on events occurring during most recent operating history (i.e., last four years). The final IE frequencies were estimated by incorporating both NPS experience and nuclear industry experience. Each event was categorized according to NPS individual plant examination (IPE) initiating events grouping approach. For the majority of the IE groups, few, or no events have occurred at the NPS. For those IE groups with few or no NPS events, the final estimate was made by means of a Bayesian update with general nuclear industry values. Exceptions are rare loss-of-coolant-accidents (LOCA) events, where evaluation of engineering aspects is used in order to determine frequency.(author)

  4. RCRA facility stabilization initiative

    International Nuclear Information System (INIS)

    1995-02-01

    The RCRA Facility Stabilization Initiative was developed as a means of implementing the Corrective Action Program's management goals recommended by the RIS for stabilizing actual or imminent releases from solid waste management units that threaten human health and the environment. The overall goal of stabilization is to, as situations warrant, control or abate threats to human health and/or the environment from releases at RCRA facilities, and/or to prevent or minimize the further spread of contamination while long-term remedies are pursued. The Stabilization initiative is a management philosophy and should not be confused with stabilization technologies

  5. Elucidating MicroRNA Regulatory Networks Using Transcriptional, Post-transcriptional, and Histone Modification Measurements

    Directory of Open Access Journals (Sweden)

    Sara J.C. Gosline

    2016-01-01

    Full Text Available MicroRNAs (miRNAs regulate diverse biological processes by repressing mRNAs, but their modest effects on direct targets, together with their participation in larger regulatory networks, make it challenging to delineate miRNA-mediated effects. Here, we describe an approach to characterizing miRNA-regulatory networks by systematically profiling transcriptional, post-transcriptional and epigenetic activity in a pair of isogenic murine fibroblast cell lines with and without Dicer expression. By RNA sequencing (RNA-seq and CLIP (crosslinking followed by immunoprecipitation sequencing (CLIP-seq, we found that most of the changes induced by global miRNA loss occur at the level of transcription. We then introduced a network modeling approach that integrated these data with epigenetic data to identify specific miRNA-regulated transcription factors that explain the impact of miRNA perturbation on gene expression. In total, we demonstrate that combining multiple genome-wide datasets spanning diverse regulatory modes enables accurate delineation of the downstream miRNA-regulated transcriptional network and establishes a model for studying similar networks in other systems.

  6. RNA-guided transcriptional regulation in planta via synthetic dCas9-based transcription factors

    KAUST Repository

    Piatek, Agnieszka Anna

    2014-11-14

    Targeted genomic regulation is a powerful approach to accelerate trait discovery and development in agricultural biotechnology. Bacteria and archaea use clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) regulatory systems for adaptive molecular immunity against foreign nucleic acids introduced by invading phages and conjugative plasmids. The type II CRISPR/Cas system has been adapted for genome editing in many cell types and organisms. A recent study used the catalytically inactive Cas9 (dCas9) protein combined with guide-RNAs (gRNAs) as a DNA-targeting platform to modulate gene expression in bacterial, yeast, and human cells. Here, we modified this DNA-targeting platform for targeted transcriptional regulation in planta by developing chimeric dCas9-based transcriptional activators and repressors. To generate transcriptional activators, we fused the dCas9 C-terminus with the activation domains of EDLL and TAL effectors. To generate a transcriptional repressor, we fused the dCas9 C-terminus with the SRDX repression domain. Our data demonstrate that dCas9 fusion with the EDLL activation domain (dCas9:EDLL) and the TAL activation domain (dCas9:TAD), guided by gRNAs complementary to selected promoter elements, induce strong transcriptional activation on Bs3

  7. RNA-guided transcriptional regulation in planta via synthetic dCas9-based transcription factors

    KAUST Repository

    Piatek, Agnieszka Anna; Ali, Zahir; Baazim, Hatoon; Li, Lixin; Abulfaraj, Aala A.; Alshareef, Sahar; Aouida, Mustapha; Mahfouz, Magdy M.

    2014-01-01

    Targeted genomic regulation is a powerful approach to accelerate trait discovery and development in agricultural biotechnology. Bacteria and archaea use clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) regulatory systems for adaptive molecular immunity against foreign nucleic acids introduced by invading phages and conjugative plasmids. The type II CRISPR/Cas system has been adapted for genome editing in many cell types and organisms. A recent study used the catalytically inactive Cas9 (dCas9) protein combined with guide-RNAs (gRNAs) as a DNA-targeting platform to modulate gene expression in bacterial, yeast, and human cells. Here, we modified this DNA-targeting platform for targeted transcriptional regulation in planta by developing chimeric dCas9-based transcriptional activators and repressors. To generate transcriptional activators, we fused the dCas9 C-terminus with the activation domains of EDLL and TAL effectors. To generate a transcriptional repressor, we fused the dCas9 C-terminus with the SRDX repression domain. Our data demonstrate that dCas9 fusion with the EDLL activation domain (dCas9:EDLL) and the TAL activation domain (dCas9:TAD), guided by gRNAs complementary to selected promoter elements, induce strong transcriptional activation on Bs3

  8. A test of the transcription model for biased inheritance of yeast mitochondrial DNA.

    Science.gov (United States)

    Lorimer, H E; Brewer, B J; Fangman, W L

    1995-09-01

    Two strand-specific origins of replication appear to be required for mammalian mitochondrial DNA (mtDNA) replication. Structural equivalents of these origins are found in the rep sequences of Saccharomyces cerevisiae mtDNA. These striking similarities have contributed to a universal model for the initiation of mtDNA replication in which a primer is created by cleavage of an origin region transcript. Consistent with this model are the properties of deletion mutants of yeast mtDNA ([rho-]) with a high density of reps (HS [rho-]). These mutant mtDNAs are preferentially inherited by the progeny resulting from the mating of HS [rho-] cells with cells containing wild-type mtDNA ([rho+]). This bias is presumed to result from a replication advantage conferred on HS [rho-] mtDNA by the high density of rep sequences acting as origins. To test whether transcription is indeed required for the preferential inheritance of HS [rho-] mtDNA, we deleted the nuclear gene (RPO41) for the mitochondrial RNA polymerase, reducing transcripts by at least 1000-fold. Since [rho-] genomes, but not [rho+] genomes, are stable when RPO41 is deleted, we examined matings between HS [rho-] and neutral [rho-] cells. Neutral [rho-] mtDNAs lack rep sequences and are not preferentially inherited in [rho-] x [rho+] crosses. In HS [rho-] x neutral [rho-] matings, the HS [rho-] mtDNA was preferentially inherited whether both parents were wild type or both were deleted for RPO41. Thus, transcription from the rep promoter does not appear to be necessary for biased inheritance. Our results, and analysis of the literature, suggest that priming by transcription is not a universal mechanism for mtDNA replication initiation.

  9. The spatial biology of transcription and translation in rapidly growing Escherichia coli

    Directory of Open Access Journals (Sweden)

    Somenath eBakshi

    2015-07-01

    Full Text Available Single-molecule fluorescence provides high resolution spatial distributions of ribosomes and RNA polymerase (RNAP in live, rapidly growing E. coli. Ribosomes are more strongly segregated from the nucleoids (chromosomal DNA than previous widefield fluorescence studies suggested. While most transcription may be co-translational, the evidence indicates that most translation occurs on free mRNA copies that have diffused from the nucleoids to a ribosome-rich region. Analysis of time-resolved images of the nucleoid spatial distribution after treatment with the transcription-halting drug rifampicin and the translation-halting drug chloramphenicol shows that both drugs cause nucleoid contraction on the 0-3 min timescale. This is consistent with the transertion hypothesis. We suggest that the longer-term (20-30 min nucleoid expansion after Rif treatment arises from conversion of 70S-polysomes to 30S and 50S subunits, which readily penetrate the nucleoids. Monte Carlo simulations of a polymer bead model built to mimic the chromosomal DNA and ribosomes (either 70S-polysomes or 30S and 50S subunits explain spatial segregation or mixing of ribosomes and nucleoids in terms of excluded volume and entropic effects alone. A comprehensive model of the transcription-translation-transertion system incorporates this new information about the spatial organization of the E. coli cytoplasm. We propose that transertion, which radially expands the nucleoids, is essential for recycling of 30S and 50S subunits from ribosome-rich regions back into the nucleoids. There they initiate co-transcriptional translation, which is an important mechanism for maintaining RNAP forward progress and protecting the nascent mRNA chain. Segregation of 70S-polysomes from the nucleoid may facilitate rapid growth by shortening the search time for ribosomes to find free mRNA concentrated outside the nucleoid and the search time for RNAP concentrated within the nucleoid to find transcription

  10. Directional gene expression and antisense transcripts in sexual and asexual stages of Plasmodium falciparum

    Directory of Open Access Journals (Sweden)

    López-Barragán María J

    2011-11-01

    Full Text Available Abstract Background It has been shown that nearly a quarter of the initial predicted gene models in the Plasmodium falciparum genome contain errors. Although there have been efforts to obtain complete cDNA sequences to correct the errors, the coverage of cDNA sequences on the predicted genes is still incomplete, and many gene models for those expressed in sexual or mosquito stages have not been validated. Antisense transcripts have widely been reported in P. falciparum; however, the extent and pattern of antisense transcripts in different developmental stages remain largely unknown. Results We have sequenced seven bidirectional libraries from ring, early and late trophozoite, schizont, gametocyte II, gametocyte V, and ookinete, and four strand-specific libraries from late trophozoite, schizont, gametocyte II, and gametocyte V of the 3D7 parasites. Alignment of the cDNA sequences to the 3D7 reference genome revealed stage-specific antisense transcripts and novel intron-exon splicing junctions. Sequencing of strand-specific cDNA libraries suggested that more genes are expressed in one direction in gametocyte than in schizont. Alternatively spliced genes, antisense transcripts, and stage-specific expressed genes were also characterized. Conclusions It is necessary to continue to sequence cDNA from different developmental stages, particularly those of non-erythrocytic stages. The presence of antisense transcripts in some gametocyte and ookinete genes suggests that these antisense RNA may play an important role in gene expression regulation and parasite development. Future gene expression studies should make use of directional cDNA libraries. Antisense transcripts may partly explain the observed discrepancy between levels of mRNA and protein expression.

  11. Palmitoylation regulates 17β-estradiol-induced estrogen receptor-α degradation and transcriptional activity.

    Science.gov (United States)

    La Rosa, Piergiorgio; Pesiri, Valeria; Leclercq, Guy; Marino, Maria; Acconcia, Filippo

    2012-05-01

    The estrogen receptor-α (ERα) is a transcription factor that regulates gene expression through the binding to its cognate hormone 17β-estradiol (E2). ERα transcriptional activity is regulated by E2-evoked 26S proteasome-mediated ERα degradation and ERα serine (S) residue 118 phosphorylation. Furthermore, ERα mediates fast cell responses to E2 through the activation of signaling cascades such as the MAPK/ERK and phosphoinositide-3-kinase/v-akt murine thymoma viral oncogene homolog 1 pathways. These E2 rapid effects require a population of the ERα located at the cell plasma membrane through palmitoylation, a dynamic enzymatic modification mediated by palmitoyl-acyl-transferases. However, whether membrane-initiated and transcriptional ERα activities integrate in a unique picture or represent parallel pathways still remains to be firmly clarified. Hence, we evaluated here the impact of ERα palmitoylation on E2-induced ERα degradation and S118 phosphorylation. The lack of palmitoylation renders ERα more susceptible to E2-dependent degradation, blocks ERα S118 phosphorylation and prevents E2-induced ERα estrogen-responsive element-containing promoter occupancy. Consequently, ERα transcriptional activity is prevented and the receptor addressed to the nuclear matrix subnuclear compartment. These data uncover a circuitry in which receptor palmitoylation links E2-dependent ERα degradation, S118 phosphorylation, and transcriptional activity in a unique molecular mechanism. We propose that rapid E2-dependent signaling could be considered as a prerequisite for ERα transcriptional activity and suggest an integrated model of ERα intracellular signaling where E2-dependent early extranuclear effects control late receptor-dependent nuclear actions.

  12. Bacillus subtilis δ Factor Functions as a Transcriptional Regulator by Facilitating the Open Complex Formation.

    Science.gov (United States)

    Prajapati, Ranjit Kumar; Sengupta, Shreya; Rudra, Paulami; Mukhopadhyay, Jayanta

    2016-01-15

    Most bacterial RNA polymerases (RNAP) contain five conserved subunits, viz. 2α, β, β', and ω. However, in many Gram-positive bacteria, especially in fermicutes, RNAP is associated with an additional factor, called δ. For over three decades since its identification, it had been thought that δ functioned as a subunit of RNAP to enhance the level of transcripts by recycling RNAP. In support of the previous observations, we also find that δ is involved in recycling of RNAP by releasing the RNA from the ternary complex. We further show that δ binds to RNA and is able to recycle RNAP when the length of the nascent RNA reaches a critical length. However, in this work we decipher a new function of δ. Performing biochemical and mutational analysis, we show that Bacillus subtilis δ binds to DNA immediately upstream of the promoter element at A-rich sequences on the abrB and rrnB1 promoters and facilitates open complex formation. As a result, δ facilitates RNAP to initiate transcription in the second scale, compared with minute scale in the absence of δ. Using transcription assay, we show that δ-mediated recycling of RNAP cannot be the sole reason for the enhancement of transcript yield. Our observation that δ does not bind to RNAP holo enzyme but is required to bind to DNA upstream of the -35 promoter element for transcription activation suggests that δ functions as a transcriptional regulator. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. Inhibition of enterovirus 71 entry by transcription factor XBP1

    Energy Technology Data Exchange (ETDEWEB)

    Jheng, Jia-Rong; Lin, Chiou-Yan [Department of Biochemistry and Research Center for Emerging Viral Infections, Chang Gung University, 259 Wen-Hwa First Road, Kweishan, Taoyuan 333, Taiwan (China); Horng, Jim-Tong, E-mail: jimtong@mail.cgu.edu.tw [Department of Biochemistry and Research Center for Emerging Viral Infections, Chang Gung University, 259 Wen-Hwa First Road, Kweishan, Taoyuan 333, Taiwan (China); Lau, Kean Seng [Department of Biochemistry and Research Center for Emerging Viral Infections, Chang Gung University, 259 Wen-Hwa First Road, Kweishan, Taoyuan 333, Taiwan (China)

    2012-04-20

    Highlights: Black-Right-Pointing-Pointer IRE1 was activated but no XBP1 splicing was detected during enterovirus 71 infection. Black-Right-Pointing-Pointer XBP1 was subject to translational shutoff by enterovirus 71-induced eIF4G cleavage. Black-Right-Pointing-Pointer The uptake of UV-irradiated virus was decreased in XBP1-overexpressing cells. -- Abstract: Inositol-requiring enzyme 1 (IRE1) plays an important role in the endoplasmic reticulum (ER), or unfolded protein, stress response by activating its downstream transcription factor X-box-binding protein 1 (XBP1). We demonstrated previously that enterovirus 71 (EV71) upregulated XBP1 mRNA levels but did not activate spliced XBP1 (XBP1s) mRNA or its downstream target genes, EDEM and chaperones. In this study, we investigated further this regulatory mechanism and found that IRE1 was phosphorylated and activated after EV71 infection, whereas its downstream XBP1s protein level decreased. We also found that XBP1s was not cleaved directly by 2A{sup pro}, but that cleavage of eukaryotic translation initiation factor 4G by the EV71 2A{sup pro} protein may contribute to the decrease in XBP1s expression. Knockdown of XBP1 increased viral protein expression, and the synthesis of EV71 viral protein and the production of EV71 viral particles were inhibited in XBP1-overexpressing RD cells. When incubated with replication-deficient and UV-irradiated EV71, XBP1-overexpressing RD cells exhibited reduced viral RNA levels, suggesting that the inhibition of XBP1s by viral infection may underlie viral entry, which is required for viral replication. Our findings are the first indication of the ability of XBP1 to inhibit viral entry, possibly via its transcriptional activity in regulating molecules in the endocytic machinery.

  14. Inhibition of enterovirus 71 entry by transcription factor XBP1

    International Nuclear Information System (INIS)

    Jheng, Jia-Rong; Lin, Chiou-Yan; Horng, Jim-Tong; Lau, Kean Seng

    2012-01-01

    Highlights: ► IRE1 was activated but no XBP1 splicing was detected during enterovirus 71 infection. ► XBP1 was subject to translational shutoff by enterovirus 71-induced eIF4G cleavage. ► The uptake of UV-irradiated virus was decreased in XBP1-overexpressing cells. -- Abstract: Inositol-requiring enzyme 1 (IRE1) plays an important role in the endoplasmic reticulum (ER), or unfolded protein, stress response by activating its downstream transcription factor X-box-binding protein 1 (XBP1). We demonstrated previously that enterovirus 71 (EV71) upregulated XBP1 mRNA levels but did not activate spliced XBP1 (XBP1s) mRNA or its downstream target genes, EDEM and chaperones. In this study, we investigated further this regulatory mechanism and found that IRE1 was phosphorylated and activated after EV71 infection, whereas its downstream XBP1s protein level decreased. We also found that XBP1s was not cleaved directly by 2A pro , but that cleavage of eukaryotic translation initiation factor 4G by the EV71 2A pro protein may contribute to the decrease in XBP1s expression. Knockdown of XBP1 increased viral protein expression, and the synthesis of EV71 viral protein and the production of EV71 viral particles were inhibited in XBP1-overexpressing RD cells. When incubated with replication-deficient and UV-irradiated EV71, XBP1-overexpressing RD cells exhibited reduced viral RNA levels, suggesting that the inhibition of XBP1s by viral infection may underlie viral entry, which is required for viral replication. Our findings are the first indication of the ability of XBP1 to inhibit viral entry, possibly via its transcriptional activity in regulating molecules in the endocytic machinery.

  15. Activating transcription factor 6 polymorphisms and haplotypes are associated with impaired glucose homeostasis and type 2 diabetes in dutch Caucasians

    NARCIS (Netherlands)

    Meex, Steven J. R.; van Greevenbroek, Marleen M. J.; Ayoubi, Torik A.; Vlietinck, Robert; van Vliet-Ostaptchouk, Jana V.; Hofker, Martin H.; Vermeulen, Vicky M. M. -J.; Schalkwijk, Casper G.; Feskens, Edith J. M.; Boer, Jolanda M. A.; Stehouwer, Coen D. A.; van der Kallen, Carla J. H.; de Bruin, Tjerk W. A.

    Context: Activating transcription factor 6 (ATF6) is critical for initiation and full activation of the unfolded protein response. An association between genetic variation in ATF6 and type 2 diabetes (DM2) was recently reported in Pima Indians. Objectives: To investigate the broader significance of

  16. Cdc7-Dbf4 Regulates NDT80 Transcription as Well as Reductional Segregation during Budding Yeast Meiosis

    OpenAIRE

    Lo, Hsiao-Chi; Wan, Lihong; Rosebrock, Adam; Futcher, Bruce; Hollingsworth, Nancy M.

    2008-01-01

    In budding yeast, as in other eukaryotes, the Cdc7 protein kinase is important for initiation of DNA synthesis in vegetative cells. In addition, Cdc7 has crucial meiotic functions: it facilitates premeiotic DNA replication, and it is essential for the initiation of recombination. This work uses a chemical genetic approach to demonstrate that Cdc7 kinase has additional roles in meiosis. First, Cdc7 allows expression of NDT80, a meiosis-specific transcriptional activator required for the induct...

  17. The SEED Initiative

    Science.gov (United States)

    Teich, Carolyn R.

    2011-01-01

    Committed to fulfilling the promise of the green economy, the American Association of Community Colleges (AACC) launched the Sustainability Education and Economic Development (SEED) initiative (www.theseedcenter.org) in October 2010. The project advances sustainability and clean energy workforce development practices at community colleges by…

  18. Major New Initiatives

    Indian Academy of Sciences (India)

    First page Back Continue Last page Overview Graphics. Major New Initiatives. Multi-party multi-rate video conferencing OOPS. Live Lecture OOPS. Rural ATM Machine Vortex. Finger print detection HP-IITM. Medical Diagnostic kit NeuroSynaptic. LCD projection system TeNeT. Web Terminal MeTeL Midas. Entertainment ...

  19. Mixed-Initiative Clustering

    Science.gov (United States)

    Huang, Yifen

    2010-01-01

    Mixed-initiative clustering is a task where a user and a machine work collaboratively to analyze a large set of documents. We hypothesize that a user and a machine can both learn better clustering models through enriched communication and interactive learning from each other. The first contribution or this thesis is providing a framework of…

  20. Monolithic exploding foil initiator

    Science.gov (United States)

    Welle, Eric J; Vianco, Paul T; Headley, Paul S; Jarrell, Jason A; Garrity, J. Emmett; Shelton, Keegan P; Marley, Stephen K

    2012-10-23

    A monolithic exploding foil initiator (EFI) or slapper detonator and the method for making the monolithic EFI wherein the exploding bridge and the dielectric from which the flyer will be generated are integrated directly onto the header. In some embodiments, the barrel is directly integrated directly onto the header.

  1. Next generation initiation techniques

    Science.gov (United States)

    Warner, Tom; Derber, John; Zupanski, Milija; Cohn, Steve; Verlinde, Hans

    1993-01-01

    Four-dimensional data assimilation strategies can generally be classified as either current or next generation, depending upon whether they are used operationally or not. Current-generation data-assimilation techniques are those that are presently used routinely in operational-forecasting or research applications. They can be classified into the following categories: intermittent assimilation, Newtonian relaxation, and physical initialization. It should be noted that these techniques are the subject of continued research, and their improvement will parallel the development of next generation techniques described by the other speakers. Next generation assimilation techniques are those that are under development but are not yet used operationally. Most of these procedures are derived from control theory or variational methods and primarily represent continuous assimilation approaches, in which the data and model dynamics are 'fitted' to each other in an optimal way. Another 'next generation' category is the initialization of convective-scale models. Intermittent assimilation systems use an objective analysis to combine all observations within a time window that is centered on the analysis time. Continuous first-generation assimilation systems are usually based on the Newtonian-relaxation or 'nudging' techniques. Physical initialization procedures generally involve the use of standard or nonstandard data to force some physical process in the model during an assimilation period. Under the topic of next-generation assimilation techniques, variational approaches are currently being actively developed. Variational approaches seek to minimize a cost or penalty function which measures a model's fit to observations, background fields and other imposed constraints. Alternatively, the Kalman filter technique, which is also under investigation as a data assimilation procedure for numerical weather prediction, can yield acceptable initial conditions for mesoscale models. The

  2. A Herpesviral Immediate Early Protein Promotes Transcription Elongation of Viral Transcripts

    Directory of Open Access Journals (Sweden)

    Hannah L. Fox

    2017-06-01

    Full Text Available Herpes simplex virus 1 (HSV-1 genes are transcribed by cellular RNA polymerase II (RNA Pol II. While four viral immediate early proteins (ICP4, ICP0, ICP27, and ICP22 function in some capacity in viral transcription, the mechanism by which ICP22 functions remains unclear. We observed that the FACT complex (comprised of SSRP1 and Spt16 was relocalized in infected cells as a function of ICP22. ICP22 was also required for the association of FACT and the transcription elongation factors SPT5 and SPT6 with viral genomes. We further demonstrated that the FACT complex interacts with ICP22 throughout infection. We therefore hypothesized that ICP22 recruits cellular transcription elongation factors to viral genomes for efficient transcription elongation of viral genes. We reevaluated the phenotype of an ICP22 mutant virus by determining the abundance of all viral mRNAs throughout infection by transcriptome sequencing (RNA-seq. The accumulation of almost all viral mRNAs late in infection was reduced compared to the wild type, regardless of kinetic class. Using chromatin immunoprecipitation sequencing (ChIP-seq, we mapped the location of RNA Pol II on viral genes and found that RNA Pol II levels on the bodies of viral genes were reduced in the ICP22 mutant compared to wild-type virus. In contrast, the association of RNA Pol II with transcription start sites in the mutant was not reduced. Taken together, our results indicate that ICP22 plays a role in recruiting elongation factors like the FACT complex to the HSV-1 genome to allow for efficient viral transcription elongation late in viral infection and ultimately infectious virion production.

  3. Processivity and coupling in messenger RNA transcription.

    Directory of Open Access Journals (Sweden)

    Stuart Aitken

    2010-01-01

    Full Text Available The complexity of messenger RNA processing is now being uncovered by experimental techniques that are capable of detecting individual copies of mRNA in cells, and by quantitative real-time observations that reveal the kinetics. This processing is commonly modelled by permitting mRNA to be transcribed only when the promoter is in the on state. In this simple on/off model, the many processes involved in active transcription are represented by a single reaction. These processes include elongation, which has a minimum time for completion and processing that is not captured in the model.In this paper, we explore the impact on the mRNA distribution of representing the elongation process in more detail. Consideration of the mechanisms of elongation leads to two alternative models of the coupling between the elongating polymerase and the state of the promoter: Processivity allows polymerases to complete elongation irrespective of the promoter state, whereas coupling requires the promoter to be active to produce a full-length transcript. We demonstrate that these alternatives have a significant impact on the predicted distributions. Models are simulated by the Gillespie algorithm, and the third and fourth moments of the resulting distribution are computed in order to characterise the length of the tail, and sharpness of the peak. By this methodology, we show that the moments provide a concise summary of the distribution, showing statistically-significant differences across much of the feasible parameter range.We conclude that processivity is not fully consistent with the on/off model unless the probability of successfully completing elongation is low--as has been observed. The results also suggest that some form of coupling between the promoter and a rate-limiting step in transcription may explain the cell's inability to maintain high mRNA levels at low noise--a prediction of the on/off model that has no supporting evidence.

  4. Rickettsia conorii transcriptional response within inoculation eschar.

    Directory of Open Access Journals (Sweden)

    Patricia Renesto

    Full Text Available BACKGROUND: Rickettsia conorii, the causative agent of the Mediterranean spotted fever, is transmitted to humans by the bite of infected ticks Rhipicephalus sanguineus. The skin thus constitutes an important barrier for the entry and propagation of R. conorii. Given this, analysis of the survival strategies used by the bacterium within infected skin is critical for our understanding of rickettsiosis. METHODOLOGY/PRINCIPAL FINDINGS: Here, we report the first genome-wide analysis of R. conorii gene expression from infected human skin biopsies. Our data showed that R. conorii exhibited a striking transcript signature that is remarkably conserved across patients, regardless of genotype. The expression profiles obtained using custom Agilent microarrays were validated by quantitative RT-PCR. Within eschars, the amount of detected R. conorii transcripts was of 55%, this value being of 74% for bacteria grown in Vero cells. In such infected host tissues, approximately 15% (n = 211 of the total predicted R. conorii ORFs appeared differentially expressed compared to bacteria grown in standard laboratory conditions. These genes are mostly down-regulated and encode proteins essential for bacterial replication. Some of the strategies displayed by rickettsiae to overcome the host defense barriers, thus avoiding killing, were also pointed out. The observed up-regulation of rickettsial genes associated with DNA repair is likely to correspond to a DNA-damaging agent enriched environment generated by the host cells to eradicate the pathogens. Survival of R. conorii within eschars also involves adaptation to osmotic stress, changes in cell surface proteins and up-regulation of some virulence factors. Interestingly, in contrast to down-regulated transcripts, we noticed that up-regulated ones rather exhibit a small nucleotide size, most of them being exclusive for the spotted fever group rickettsiae. CONCLUSION/SIGNIFICANCE: Because eschar is a site for rickettsial

  5. Extensive polycistronism and antisense transcription in the mammalian Hox clusters.

    Directory of Open Access Journals (Sweden)

    Gaëll Mainguy

    Full Text Available The Hox clusters play a crucial role in body patterning during animal development. They encode both Hox transcription factor and micro-RNA genes that are activated in a precise temporal and spatial sequence that follows their chromosomal order. These remarkable collinear properties confer functional unit status for Hox clusters. We developed the TranscriptView platform to establish high resolution transcriptional profiling and report here that transcription in the Hox clusters is far more complex than previously described in both human and mouse. Unannotated transcripts can represent up to 60% of the total transcriptional output of a cluster. In particular, we identified 14 non-coding Transcriptional Units antisense to Hox genes, 10 of which (70% have a detectable mouse homolog. Most of these Transcriptional Units in both human and mouse present conserved sizeable sequences (>40 bp overlapping Hox transcripts, suggesting that these Hox antisense transcripts are functional. Hox clusters also display at least seven polycistronic clusters, i.e., different genes being co-transcribed on long isoforms (up to 30 kb. This work provides a reevaluated framework for understanding Hox gene function and dys-function. Such extensive transcriptions may provide a structural explanation for Hox clustering.

  6. Transcription arrest caused by long nascent RNA chains

    DEFF Research Database (Denmark)

    Bentin, Thomas; Cherny, Dmitry; Larsen, H Jakob

    2004-01-01

    on transcription. Using phage T3 RNA polymerase (T3 RNAP) and covalently closed circular (cccDNA) DNA templates that did not contain any strong termination signal, transcription was severely inhibited after a short period of time. Less than approximately 10% residual transcriptional activity remained after 10 min......The transcription process is highly processive. However, specific sequence elements encoded in the nascent RNA may signal transcription pausing and/or termination. We find that under certain conditions nascent RNA chains can have a strong and apparently sequence-independent inhibitory effect...... of incubation. The addition of RNase A almost fully restored transcription in a dose dependent manner. Throughout RNase A rescue, an elongation rate of approximately 170 nt/s was maintained and this velocity was independent of RNA transcript length, at least up to 6 kb. Instead, RNase A rescue increased...

  7. Transcriptional activation of ribosomal RNA genes during compensatory renal hypertrophy

    International Nuclear Information System (INIS)

    Ouellette, A.J.; Moonka, R.; Zelenetz, A.; Malt, R.A.

    1986-01-01

    The overall rate of rDNA transcription increases by 50% during the first 24 hours of compensatory renal hypertrophy in the mouse. To study mechanisms of ribosome accumulation after uninephrectomy, transcription rates were measured in isolated kidneys by transcriptional runoff. 32 P-labeled nascent transcripts were hybridized to blots containing linearized, denatured cloned rDNA, and hybridization was quantitated autoradiographically and by direct counting. Overall transcriptional activity of rDNA was increased by 30% above control levels at 6 hrs after nephrectomy and by 50% at 12, 18, and 24 hrs after operation. Hybridizing RNA was insensitive to inhibiby alpha-amanitin, and no hybridization was detected to vector DNA. Thus, accelerated rDNA transcription is one regulatory element in the accretion of ribosomes in renal growth, and the regulatory event is an early event. Mechanisms of activation may include enhanced transcription of active genes or induction of inactive DNA

  8. Controllability analysis of transcriptional regulatory networks reveals circular control patterns among transcription factors

    DEFF Research Database (Denmark)

    Österlund, Tobias; Bordel, Sergio; Nielsen, Jens

    2015-01-01

    % for the human network. The high controllability (low number of drivers needed to control the system) in yeast, mouse and human is due to the presence of internal loops in their regulatory networks where the TFs regulate each other in a circular fashion. We refer to these internal loops as circular control...... motifs (CCM). The E. coli transcriptional regulatory network, which does not have any CCMs, shows a hierarchical structure of the transcriptional regulatory network in contrast to the eukaryal networks. The presence of CCMs also has influence on the stability of these networks, as the presence of cycles...

  9. The evolution of WRKY transcription factors.

    Science.gov (United States)

    Rinerson, Charles I; Rabara, Roel C; Tripathi, Prateek; Shen, Qingxi J; Rushton, Paul J

    2015-02-27

    The availability of increasing numbers of sequenced genomes has necessitated a re-evaluation of the evolution of the WRKY transcription factor family. Modern day plants descended from a charophyte green alga that colonized the land between 430 and 470 million years ago. The first charophyte genome sequence from Klebsormidium flaccidum filled a gap in the available genome sequences in the plant kingdom between unicellular green algae that typically have 1-3 WRKY genes and mosses that contain 30-40. WRKY genes have been previously found in non-plant species but their occurrence has been difficult to explain. Only two WRKY genes are present in the Klebsormidium flaccidum genome and the presence of a Group IIb gene was unexpected because it had previously been thought that Group IIb WRKY genes first appeared in mosses. We found WRKY transcription factor genes outside of the plant lineage in some diplomonads, social amoebae, fungi incertae sedis, and amoebozoa. This patchy distribution suggests that lateral gene transfer is responsible. These lateral gene transfer events appear to pre-date the formation of the WRKY groups in flowering plants. Flowering plants contain proteins with domains typical for both resistance (R) proteins and WRKY transcription factors. R protein-WRKY genes have evolved numerous times in flowering plants, each type being restricted to specific flowering plant lineages. These chimeric proteins contain not only novel combinations of protein domains but also novel combinations and numbers of WRKY domains. Once formed, R protein WRKY genes may combine different components of signalling pathways that may either create new diversity in signalling or accelerate signalling by short circuiting signalling pathways. We propose that the evolution of WRKY transcription factors includes early lateral gene transfers to non-plant organisms and the occurrence of algal WRKY genes that have no counterparts in flowering plants. We propose two alternative hypotheses

  10. Transcriptional regulation by Polycomb group proteins

    DEFF Research Database (Denmark)

    Di Croce, Luciano; Helin, Kristian

    2013-01-01

    Polycomb group (PcG) proteins are epigenetic regulators of transcription that have key roles in stem-cell identity, differentiation and disease. Mechanistically, they function within multiprotein complexes, called Polycomb repressive complexes (PRCs), which modify histones (and other proteins......) and silence target genes. The dynamics of PRC1 and PRC2 components has been the focus of recent research. Here we discuss our current knowledge of the PRC complexes, how they are targeted to chromatin and how the high diversity of the PcG proteins allows these complexes to influence cell identity....

  11. Transcriptional landscape of glomerular parietal epithelial cells.

    Directory of Open Access Journals (Sweden)

    Sina A Gharib

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

  12. Transcriptional delay stabilizes bistable gene networks.

    Science.gov (United States)

    Gupta, Chinmaya; López, José Manuel; Ott, William; Josić, Krešimir; Bennett, Matthew R

    2013-08-02

    Transcriptional delay can significantly impact the dynamics of gene networks. Here we examine how such delay affects bistable systems. We investigate several stochastic models of bistable gene networks and find that increasing delay dramatically increases the mean residence times near stable states. To explain this, we introduce a non-Markovian, analytically tractable reduced model. The model shows that stabilization is the consequence of an increased number of failed transitions between stable states. Each of the bistable systems that we simulate behaves in this manner.

  13. Transcription of repetitive DNA in Neurospora crassa

    Energy Technology Data Exchange (ETDEWEB)

    Dutta, S K; Chaudhuri, R K

    1975-01-01

    Repeated DNA sequences of Neurospora crassa were isolated and characterized. Approximately 10 to 12 percent of N. crassa DNA sequence were repeated, of which 7.3 percent were found to be transcribed in mid-log phase of mycelial growth as measured by DNA:RNA hybridization. It is suggested that part of repetitive DNA transcripts in N. crassa were mitochondrial and part were nuclear DNA. Most of the nuclear repeated DNAs, however, code for rRNA and tRNA in N. crassa. (auth)

  14. 'Crisis' and 'everyday' initiators: A qualitative study of coercion and agency in the context of methadone maintenance treatment initiation.

    Science.gov (United States)

    Damon, Will; Small, Will; Anderson, Solanna; Maher, Lisa; Wood, Evan; Kerr, Thomas; McNeil, Ryan

    2017-03-01

    Patient attrition is common among people enrolled in methadone maintenance treatment (MMT) programs and most pronounced during the first year of treatment. However, the experiences of patients initiating MMT have been overlooked in the literature. This study explores experiences of MMT initiation among MMT patients, focusing on contextual influences on MMT initiation and perceptions of MMT and their subsequent influence on treatment retention. Semi-structured qualitative interviews were conducted with 39 MMT patients in Vancouver, Canada. Individuals reporting enrolment in MMT were recruited from within two ongoing cohort studies comprised of people who use drugs. Interview transcripts were analysed using an inductive and iterative approach. Two groups of MMT initiators were identified: (i) 'crisis initiators' prescribed methadone following critical transition events, such as incarceration or pregnancy; and (ii) 'everyday initiators' enrolled in MMT as part of routine healthcare utilisation. While most 'crisis initiators' and some 'everyday initiators' described experiencing coercion during MMT initiation, 'crisis initiators' were further subjected to the coercive leveraging of their vulnerability to motivate 'consent' for MMT. 'Crisis initiators' developed negative views towards MMT and were more likely to discontinue treatment. Long-standing patient-provider relationships and open dialogue were associated with more positive views regarding MMT, regardless of the circumstances of initiation. Findings underscore the need for clear and effective communication regarding treatment regimens and expectations during MMT initiation. Furthermore, training in trauma-informed care may help reduce perceptions of coercion and rates of early treatment termination. [Damon W, Small W, Anderson S, Maher L, Wood E, Kerr T, McNeil R. Crisis' and 'everyday' initiators: A qualitative study of coercion and agency in the context of methadone maintenance treatment initiation. Drug

  15. Funding Initiatives | Women in Science | Initiatives | Indian Academy ...

    Indian Academy of Sciences (India)

    Home; Initiatives; Women in Science; Funding Initiatives ... The Fellowship Scheme for Women Scientists for societal programmes is initiative of the ... at a young age of 52, after a valiant battle with cancer, today on 29th March 2016 in Delhi.

  16. Sustaining Participatory Design Initiatives

    DEFF Research Database (Denmark)

    Iversen, Ole Sejer; Dindler, Christian

    2014-01-01

    While many participatory design (PD) projects succeed in establishing new organisational initiatives or creating technology that is attuned to the people affected, the issue of how such results are sustained after the project ends remains an important challenge. We explore the challenge...... these various forms of sustainability may be pursued in PD practice and how they can become a resource in reflecting on PD activities. Finally, we discuss implications for PD practice, suggesting that a nuanced conception of sustainability and how it may relate to PD practice are useful resources for designers...... of sustaining PD initiatives beyond the individual project and discuss implications for PD practice. First, based on current PD literature, we distinguish between four ideal typical forms of sustainability: maintaining, scaling, replicating and evolving. Second, we demonstrate from a case study how...

  17. Initial management of breastfeeding.

    Science.gov (United States)

    Sinusas, K; Gagliardi, A

    2001-09-15

    Breast milk is widely accepted as the ideal source of nutrition for infants. In order to ensure success in breastfeeding, it is important that it be initiated as early as possible during the neonatal period. This is facilitated by skin-to-skin contact between the mother and infant immediately following birth. When possible, the infant should be allowed to root and latch on spontaneously within the first hour of life. Many common nursery routines such as weighing the infant, administration of vitamin K and application of ocular antibiotics can be safely delayed until after the initial breastfeeding. Postpartum care practices that improve breastfeeding rates include rooming-in, anticipatory guidance about breastfeeding problems and the avoidance of formula supplementation and pacifiers.

  18. Self-initiated expatriates

    DEFF Research Database (Denmark)

    Selmer, Jan; Lauring, Jakob

    2014-01-01

    Purpose – As it has been suggested that adult third-culture kids may be more culturally adaptable than others, they have been labelled “the ideal” expatriates. In this article, we explore the adjustment of self-initiated expatriate academics in Hong Kong, comparing adult third-culture kids...... with adult mono-culture kids. Design/methodology/approach – We use survey results from 267 self-initiated expatriate academics in Hong Kong. Findings – Exploratory results show that adult third-culture kids had a higher extent of general adjustment. No significant results were found in relation...... to interaction adjustment and job adjustment. We also found that recent expatriate experiences generally had a positive association with the adjustment of adult mono-culture kids, but this association only existed in terms of general adjustment for adult third-culture kids. Originality/value – Once corroborated...

  19. INITIAL TRAINING OF RESEARCHERS

    Directory of Open Access Journals (Sweden)

    Karina Alejandra Cruz-Pallares

    2015-07-01

    Full Text Available The document presents results of a research that used as strategy a complementary training project with thirty-three students of a Bachelors Degree in Primary School 1997(DPS,1997 of an Education Faculty for the initial training of investigators, applied by four teachers members of the academic research group in Mexico; that develops through process of action research methodology. Highlighted in results is the strengthening of the competition of reading, understanding and writing scientific texts, which is analogous to the first feature of the graduate profile called intellectual skills. Among the conclusions it is emphasized that the initial training of teachers in a task that is quite interesting, challenging and complex, as is the educational complex phenomenon.

  20. Hyperosmotic stress regulates the distribution and stability of myocardin-related transcription factor, a key modulator of the cytoskeleton

    DEFF Research Database (Denmark)

    Ly, Donald L.; Waheed, Faiza; Lodyga, Monika

    2013-01-01

    Hyperosmotic stress initiates several adaptive responses, including the remodeling of the cytoskeleton. Besides maintaining structural integrity, the cytoskeleton has emerged as an important regulator of gene transcription. Myocardin-related transcription factor (MRTF), an actin-regulated coactiv......Hyperosmotic stress initiates several adaptive responses, including the remodeling of the cytoskeleton. Besides maintaining structural integrity, the cytoskeleton has emerged as an important regulator of gene transcription. Myocardin-related transcription factor (MRTF), an actin......-regulated coactivator of serum response factor, is a major link between the actin skeleton and transcriptional control. We therefore investigated whether MRTF is regulated by hyperosmotic stress. Here we show that hypertonicity induces robust, rapid, and transient translocation of MRTF from the cytosol to the nucleus...... in kidney tubular cells. We found that the hyperosmolarity-triggered MRTF translocation is mediated by the RhoA/Rho kinase (ROK) pathway. Moreover, the Rho guanine nucleotide exchange factor GEF-H1 is activated by hyperosmotic stress, and it is a key contributor to the ensuing RhoA activation and MRTF...

  1. Transcription factor Oct1 is a somatic and cancer stem cell determinant.

    Directory of Open Access Journals (Sweden)

    Jessica Maddox

    Full Text Available Defining master transcription factors governing somatic and cancer stem cell identity is an important goal. Here we show that the Oct4 paralog Oct1, a transcription factor implicated in stress responses, metabolic control, and poised transcription states, regulates normal and pathologic stem cell function. Oct1(HI cells in the colon and small intestine co-express known stem cell markers. In primary malignant tissue, high Oct1 protein but not mRNA levels strongly correlate with the frequency of CD24(LOCD44(HI cancer-initiating cells. Reducing Oct1 expression via RNAi reduces the proportion of ALDH(HI and dye efflux(HI cells, and increasing Oct1 increases the proportion of ALDH(HI cells. Normal ALDH(HI cells harbor elevated Oct1 protein but not mRNA levels. Functionally, we show that Oct1 promotes tumor engraftment frequency and promotes hematopoietic stem cell engraftment potential in competitive and serial transplants. In addition to previously described Oct1 transcriptional targets, we identify four Oct1 targets associated with the stem cell phenotype. Cumulatively, the data indicate that Oct1 regulates normal and cancer stem cell function.

  2. The flavonoid pathway in tomato seedlings: transcript abundance and the modeling of metabolite dynamics.

    Directory of Open Access Journals (Sweden)

    Marian Groenenboom

    Full Text Available Flavonoids are secondary metabolites present in all terrestrial plants. The flavonoid pathway has been extensively studied, and many of the involved genes and metabolites have been described in the literature. Despite this extensive knowledge, the functioning of the pathway in vivo is still poorly understood. Here, we study the flavonoid pathway using both experiments and mathematical models. We measured flavonoid metabolite dynamics in two tissues, hypocotyls and cotyledons, during tomato seedling development. Interestingly, the same backbone of interactions leads to very different accumulation patterns in the different tissues. Initially, we developed a mathematical model with constant enzyme concentrations that described the metabolic networks separately in both tissues. This model was unable to fit the measured flavonoid dynamics in the hypocotyls, even if we allowed unrealistic parameter values. This suggested us to investigate the effect of transcript abundance on flavonoid accumulation. We found that the expression of candidate flavonoid genes varies considerably with time. Variation in transcript abundance results in enzymatic variation, which could have a large effect on metabolite accumulation. Candidate transcript abundance was included in the mathematical model as representative for enzyme concentration. We fitted the resulting model to the flavonoid dynamics in the cotyledons, and tested it by applying it to the data from hypocotyls. When transcript abundance is included, we are indeed able to explain flavonoid dynamics in both tissues. Importantly, this is possible under the biologically relevant restriction that the enzymatic properties estimated by the model are conserved between the tissues.

  3. In Silico discovery of transcription factors as potential diagnostic biomarkers of ovarian cancer

    KAUST Repository

    Kaur, Mandeep

    2011-09-19

    Background: Our study focuses on identifying potential biomarkers for diagnosis and early detection of ovarian cancer (OC) through the study of transcription regulation of genes affected by estrogen hormone.Results: The results are based on a set of 323 experimentally validated OC-associated genes compiled from several databases, and their subset controlled by estrogen. For these two gene sets we computationally determined transcription factors (TFs) that putatively regulate transcription initiation. We ranked these TFs based on the number of genes they are likely to control. In this way, we selected 17 top-ranked TFs as potential key regulators and thus possible biomarkers for a set of 323 OC-associated genes. For 77 estrogen controlled genes from this set we identified three unique TFs as potential biomarkers.Conclusions: We introduced a new methodology to identify potential diagnostic biomarkers for OC. This report is the first bioinformatics study that explores multiple transcriptional regulators of OC-associated genes as potential diagnostic biomarkers in connection with estrogen responsiveness. We show that 64% of TF biomarkers identified in our study are validated based on real-time data from microarray expression studies. As an illustration, our method could identify CP2 that in combination with CA125 has been reported to be sensitive in diagnosing ovarian tumors. 2011 Kaur et al; licensee BioMed Central Ltd.

  4. In Silico discovery of transcription factors as potential diagnostic biomarkers of ovarian cancer

    KAUST Repository

    Kaur, Mandeep; MacPherson, Cameron R; Schmeier, Sebastian; Narasimhan, Kothandaraman; Choolani, Mahesh; Bajic, Vladimir B.

    2011-01-01

    Background: Our study focuses on identifying potential biomarkers for diagnosis and early detection of ovarian cancer (OC) through the study of transcription regulation of genes affected by estrogen hormone.Results: The results are based on a set of 323 experimentally validated OC-associated genes compiled from several databases, and their subset controlled by estrogen. For these two gene sets we computationally determined transcription factors (TFs) that putatively regulate transcription initiation. We ranked these TFs based on the number of genes they are likely to control. In this way, we selected 17 top-ranked TFs as potential key regulators and thus possible biomarkers for a set of 323 OC-associated genes. For 77 estrogen controlled genes from this set we identified three unique TFs as potential biomarkers.Conclusions: We introduced a new methodology to identify potential diagnostic biomarkers for OC. This report is the first bioinformatics study that explores multiple transcriptional regulators of OC-associated genes as potential diagnostic biomarkers in connection with estrogen responsiveness. We show that 64% of TF biomarkers identified in our study are validated based on real-time data from microarray expression studies. As an illustration, our method could identify CP2 that in combination with CA125 has been reported to be sensitive in diagnosing ovarian tumors. 2011 Kaur et al; licensee BioMed Central Ltd.

  5. saRNA-guided Ago2 targets the RITA complex to promoters to stimulate transcription.

    Science.gov (United States)

    Portnoy, Victoria; Lin, Szu Hua Sharon; Li, Kathy H; Burlingame, Alma; Hu, Zheng-Hui; Li, Hao; Li, Long-Cheng

    2016-03-01

    Small activating RNAs (saRNAs) targeting specific promoter regions are able to stimulate gene expression at the transcriptional level, a phenomenon known as RNA activation (RNAa). It is known that RNAa depends on Ago2 and is associated with epigenetic changes at the target promoters. However, the precise molecular mechanism of RNAa remains elusive. Using human CDKN1A (p21) as a model gene, we characterized the molecular nature of RNAa. We show that saRNAs guide Ago2 to and associate with target promoters. saRNA-loaded Ago2 facilitates the assembly of an RNA-induced transcriptional activation (RITA) complex, which, in addition to saRNA-Ago2 complex, includes RHA and CTR9, the latter being a component of the PAF1 complex. RITA interacts with RNA polymerase II to stimulate transcription initiation and productive elongation, accompanied by monoubiquitination of histone 2B. Our results establish the existence of a cellular RNA-guided genome-targeting and transcriptional activation mechanism and provide important new mechanistic insights into the RNAa process.

  6. Influence of major-groove chemical modifications of DNA on transcription by bacterial RNA polymerases.

    Science.gov (United States)

    Raindlová, Veronika; Janoušková, Martina; Slavíčková, Michaela; Perlíková, Pavla; Boháčová, Soňa; Milisavljevič, Nemanja; Šanderová, Hana; Benda, Martin; Barvík, Ivan; Krásný, Libor; Hocek, Michal

    2016-04-20

    DNA templates containing a set of base modifications in the major groove (5-substituted pyrimidines or 7-substituted 7-deazapurines bearing H, methyl, vinyl, ethynyl or phenyl groups) were prepared by PCR using the corresponding base-modified 2'-deoxyribonucleoside triphosphates (dNTPs). The modified templates were used in an in vitro transcription assay using RNA polymerase from Bacillus subtilis and Escherichia coli Some modified nucleobases bearing smaller modifications (H, Me in 7-deazapurines) were perfectly tolerated by both enzymes, whereas bulky modifications (Ph at any nucleobase) and, surprisingly, uracil blocked transcription. Some middle-sized modifications (vinyl or ethynyl) were partly tolerated mostly by the E. colienzyme. In all cases where the transcription proceeded, full length RNA product with correct sequence was obtained indicating that the modifications of the template are not mutagenic and the inhibition is probably at the stage of initiation. The results are promising for the development of bioorthogonal reactions for artificial chemical switching of the transcription. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  7. Embryonic maturation of epidermal Merkel cells is controlled by a redundant transcription factor network.

    Science.gov (United States)

    Perdigoto, Carolina N; Bardot, Evan S; Valdes, Victor J; Santoriello, Francis J; Ezhkova, Elena

    2014-12-01

    Merkel cell-neurite complexes are located in touch-sensitive areas of the mammalian skin and are involved in recognition of the texture and shape of objects. Merkel cells are essential for these tactile discriminations, as they generate action potentials in response to touch stimuli and induce the firing of innervating afferent nerves. It has been shown that Merkel cells originate from epidermal stem cells, but the cellular and molecular mechanisms of their development are largely unknown. In this study, we analyzed Merkel cell differentiation during development and found that it is a temporally regulated maturation process characterized by a sequential activation of Merkel cell-specific genes. We uncovered key transcription factors controlling this process and showed that the transcription factor Atoh1 is required for initial Merkel cell specification. The subsequent maturation steps of Merkel cell differentiation are controlled by cooperative function of the transcription factors Sox2 and Isl1, which physically interact and work to sustain Atoh1 expression. These findings reveal the presence of a robust transcriptional network required to produce functional Merkel cells that are required for tactile discrimination. © 2014. Published by The Company of Biologists Ltd.

  8. Transcriptional regulation of the Hansenula polymorpha GSH2 gene in the response to cadmium ion treatment

    Directory of Open Access Journals (Sweden)

    O. V. Blazhenko

    2014-02-01

    Full Text Available In a previous study we cloned GSH2 gene, encoding γ-glutamylcysteine synthetase (γGCS in the yeast Hansenula рolymorpha. In this study an analysis of molecular organisation of the H. рolymorpha GSH2 gene promoter was conducted and the potential binding sites of Yap1, Skn7, Creb/Atf1, and Cbf1 transcription factors were detected. It was established that full regulation of GSH2 gene expression in the response to cadmium and oxidative stress requires the length of GSH2 promoter to be longer than 450 bp from the start of translation initiation. To study the transcriptional regulation of H. polymorpha GSH2 gene recombinant strain, harbouring­ a reporter system, in which 1.832 kb regulatory region of GSH2 gene was fused to structural and terminatory regions of alcohol oxidase gene, was constructed. It was shown that maximum increase in H. polymorpha GSH2 gene transcription by 33% occurs in the rich medium under four-hour incubation with 1 μM concentration of cadmium ions. In the minimal medium the GSH2 gene expression does not correlate with the increased total cellular glutathione levels under cadmium ion treatment. We assume that the increased content of total cellular glutathione under cadmium stress in the yeast H. polymorpha probably is not controlled on the level of GSH2 gene transcription.

  9. Exosome proteomics reveals transcriptional regulator proteins with potential to mediate downstream pathways.

    Science.gov (United States)

    Ung, Timothy H; Madsen, Helen J; Hellwinkel, Justin E; Lencioni, Alex M; Graner, Michael W

    2014-11-01

    Exosomes are virus-sized, membrane-enclosed vesicles with origins in the cellular endosomal system, but are released extracellularly. As a population, these tiny vesicles carry relatively enormous amounts of information in their protein, lipid and nucleic acid content, and the vesicles can have profound impacts on recipient cells. This review employs publically-available data combined with gene ontology applications to propose a novel concept, that exosomes transport transcriptional and translational machinery that may have direct impacts on gene expression in recipient cells. Here, we examine the previously published proteomic contents of medulloblastoma-derived exosomes, focusing on transcriptional regulators; we found that there are numerous proteins that may have potential roles in transcriptional and translational regulation with putative influence on downstream, cancer-related pathways. We expanded this search to all of the proteins in the Vesiclepedia database; using gene ontology approaches, we see that these regulatory factors are implicated in many of the processes involved in cancer initiation and progression. This information suggests that some of the effects of exosomes on recipient cells may be due to the delivery of protein factors that can directly and fundamentally change the transcriptional landscape of the cells. Within a tumor environment, this has potential to tilt the advantage towards the cancer. © 2014 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

  10. Extended region of nodulation genes in Rhizobium meliloti 1021. II. Nucleotide sequence, transcription start sites and protein products

    International Nuclear Information System (INIS)

    Fisher, R.F.; Swanson, J.A.; Mulligan, J.T.; Long, S.R.

    1987-01-01

    The authors have established the DNA sequence and analyzed the transcription and translation products of a series of putative nodulation (nod) genes in Rhizobium meliloti strain 1021. Four loci have been designated nodF, nodE, nodG and nodH. The correlation of transposon insertion positions with phenotypes and open reading frames was confirmed by sequencing the insertion junctions of the transposons. The protein products of these nod genes were visualized by in vitro expression of cloned DNA segments in a R. meliloti transcription-translation system. In addition, the sequence for nodG was substantiated by creating translational fusions in all three reading frames at several points in the sequence; the resulting fusions were expressed in vitro in both E. coli and R. meliloti transcription-translation systems. A DNA segment bearing several open reading frames downstream of nodG corresponds to the putative nod gene mutated in strain nod-216. The transcription start sites of nodF and nodH were mapped by primer extension of RNA from cells induced with the plant flavone, luteolin. Initiation of transcription occurs approximately 25 bp downstream from the conserved sequence designated the nod box, suggesting that this conserved sequence acts as an upstream regulator of inducible nod gene expression. Its distance from the transcription start site is more suggestive of an activator binding site rather than an RNA polymerase binding site

  11. Stimulation of ribosomal RNA gene promoter by transcription factor Sp1 involves active DNA demethylation by Gadd45-NER pathway.

    Science.gov (United States)

    Rajput, Pallavi; Pandey, Vijaya; Kumar, Vijay

    2016-08-01

    The well-studied Pol II transcription factor Sp1 has not been investigated for its regulatory role in rDNA transcription. Here, we show that Sp1 bound to specific sites on rDNA and localized into the nucleoli during the G1 phase of cell cycle to activate rDNA transcription. It facilitated the recruitment of Pol I pre-initiation complex and impeded the binding of nucleolar remodeling complex (NoRC) to rDNA resulting in the formation of euchromatin active state. More importantly, Sp1 also orchestrated the site-specific binding of Gadd45a-nucleotide excision repair (NER) complex resulting in active demethylation and transcriptional activation of rDNA. Interestingly, knockdown of Sp1 impaired rDNA transcription due to reduced engagement of the Gadd45a-NER complex and hypermethylation of rDNA. Thus, the present study unveils a novel role of Sp1 in rDNA transcription involving promoter demethylation. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Mechanism and manipulation of DNA:RNA hybrid G-quadruplex formation in transcription of G-rich DNA.

    Science.gov (United States)

    Zhang, Jia-yu; Zheng, Ke-wei; Xiao, Shan; Hao, Yu-hua; Tan, Zheng

    2014-01-29

    We recently reported that a DNA:RNA hybrid G-quadruplex (HQ) forms during transcription of DNA that bears two or more tandem guanine tracts (G-tract) on the nontemplate strand. Putative HQ-forming sequences are enriched in the nearby 1000 nt region right downstream of transcription start sites in the nontemplate strand of warm-blooded animals, and HQ regulates transcription under both in vitro and in vivo conditions. Therefore, knowledge of the mechanism of HQ formation is important for understanding the biological function of HQ as well as for manipulating gene expression by targeting HQ. In this work, we studied the mechanism of HQ formation using an in vitro T7 transcription model. We show that RNA synthesis initially produces an R-loop, a DNA:RNA heteroduplex formed by a nascent RNA transcript and the template DNA strand. In the following round of transcription, the RNA in the R-loop is displaced, releasing the RNA in single-stranded form (ssRNA). Then the G-tracts in the RNA can jointly form HQ with those in the nontemplate DNA strand. We demonstrate that the structural cascade R-loop → ssRNA → HQ offers opportunities to intercept HQ formation, which may provide a potential method to manipulate gene expression.

  13. A human transcription factor in search mode.

    Science.gov (United States)

    Hauser, Kevin; Essuman, Bernard; He, Yiqing; Coutsias, Evangelos; Garcia-Diaz, Miguel; Simmerling, Carlos

    2016-01-08

    Transcription factors (TF) can change shape to bind and recognize DNA, shifting the energy landscape from a weak binding, rapid search mode to a higher affinity recognition mode. However, the mechanism(s) driving this conformational change remains unresolved and in most cases high-resolution structures of the non-specific complexes are unavailable. Here, we investigate the conformational switch of the human mitochondrial transcription termination factor MTERF1, which has a modular, superhelical topology complementary to DNA. Our goal was to characterize the details of the non-specific search mode to complement the crystal structure of the specific binding complex, providing a basis for understanding the recognition mechanism. In the specific complex, MTERF1 binds a significantly distorted and unwound DNA structure, exhibiting a protein conformation incompatible with binding to B-form DNA. In contrast, our simulations of apo MTERF1 revealed significant flexibility, sampling structures with superhelical pitch and radius complementary to the major groove of B-DNA. Docking these structures to B-DNA followed by unrestrained MD simulations led to a stable complex in which MTERF1 was observed to undergo spontaneous diffusion on the DNA. Overall, the data support an MTERF1-DNA binding and recognition mechanism driven by intrinsic dynamics of the MTERF1 superhelical topology. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  14. Transcriptional and posttranscriptional regulation of cyanobacterial photosynthesis.

    Science.gov (United States)

    Wilde, Annegret; Hihara, Yukako

    2016-03-01

    Cyanobacteria are well established model organisms for the study of oxygenic photosynthesis, nitrogen metabolism, toxin biosynthesis, and salt acclimation. However, in comparison to other model bacteria little is known about regulatory networks, which allow cyanobacteria to acclimate to changing environmental conditions. The current work has begun to illuminate how transcription factors modulate expression of different photosynthetic regulons. During the past few years, the research on other regulatory principles like RNA-based regulation showed the importance of non-protein regulators for bacterial lifestyle. Investigations on modulation of photosynthetic components should elucidate the contributions of all factors within the context of a larger regulatory network. Here, we focus on regulation of photosynthetic processes including transcriptional and posttranscriptional mechanisms, citing examples from a limited number of cyanobacterial species. Though, the general idea holds true for most species, important differences exist between various organisms, illustrating diversity of acclimation strategies in the very heterogeneous cyanobacterial clade. This article is part of a Special Issue entitled Organization and dynamics of bioenergetic systems in bacteria, edited by Prof Conrad Mullineaux. Copyright © 2015 Elsevier B.V. All rights reserved.

  15. Transcriptional regulation of Drosophila gonad formation.

    Science.gov (United States)

    Tripathy, Ratna; Kunwar, Prabhat S; Sano, Hiroko; Renault, Andrew D

    2014-08-15

    The formation of the Drosophila embryonic gonad, involving the fusion of clusters of somatic gonadal precursor cells (SGPs) and their ensheathment of germ cells, provides a simple and genetically tractable model for the interplay between cells during organ formation. In a screen for mutants affecting gonad formation we identified a SGP cell autonomous role for Midline (Mid) and Longitudinals lacking (Lola). These transcriptional factors are required for multiple aspects of SGP behaviour including SGP cluster fusion, germ cell ensheathment and gonad compaction. The lola locus encodes more than 25 differentially spliced isoforms and we have identified an isoform specific requirement for lola in the gonad which is distinct from that in nervous system development. Mid and Lola work in parallel in gonad formation and surprisingly Mid overexpression in a lola background leads to additional SGPs at the expense of fat body cells. Our findings support the idea that although the transcription factors required by SGPs can ostensibly be assigned to those being required for either SGP specification or behaviour, they can also interact to impinge on both processes. Copyright © 2014 Elsevier Inc. All rights reserved.

  16. Inferring Molecular Processes Heterogeneity from Transcriptional Data.

    Science.gov (United States)

    Gogolewski, Krzysztof; Wronowska, Weronika; Lech, Agnieszka; Lesyng, Bogdan; Gambin, Anna

    2017-01-01

    RNA microarrays and RNA-seq are nowadays standard technologies to study the transcriptional activity of cells. Most studies focus on tracking transcriptional changes caused by specific experimental conditions. Information referring to genes up- and downregulation is evaluated analyzing the behaviour of relatively large population of cells by averaging its properties. However, even assuming perfect sample homogeneity, different subpopulations of cells can exhibit diverse transcriptomic profiles, as they may follow different regulatory/signaling pathways. The purpose of this study is to provide a novel methodological scheme to account for possible internal, functional heterogeneity in homogeneous cell lines, including cancer ones. We propose a novel computational method to infer the proportion between subpopulations of cells that manifest various functional behaviour in a given sample. Our method was validated using two datasets from RNA microarray experiments. Both experiments aimed to examine cell viability in specific experimental conditions. The presented methodology can be easily extended to RNA-seq data as well as other molecular processes. Moreover, it complements standard tools to indicate most important networks from transcriptomic data and in particular could be useful in the analysis of cancer cell lines affected by biologically active compounds or drugs.

  17. Transcriptional regulatory programs underlying barley germination and regulatory functions of Gibberellin and abscisic acid

    Science.gov (United States)

    2011-01-01

    Background Seed germination is a complex multi-stage developmental process, and mainly accomplished through concerted activities of many gene products and biological pathways that are often subjected to strict developmental regulation. Gibberellins (GA) and abscisic acid (ABA) are two key phytohormones regulating seed germination and seedling growth. However, transcriptional regulatory networks underlying seed germination and its associated biological pathways are largely unknown. Results The studies examined transcriptomes of barley representing six distinct and well characterized germination stages and revealed that the transcriptional regulatory program underlying barley germination was composed of early, late, and post-germination phases. Each phase was accompanied with transcriptional up-regulation of distinct biological pathways. Cell wall synthesis and regulatory components including transcription factors, signaling and post-translational modification components were specifically and transiently up-regulated in early germination phase while histone families and many metabolic pathways were up-regulated in late germination phase. Photosynthesis and seed reserve mobilization pathways were up-regulated in post-germination phase. However, stress related pathways and seed storage proteins were suppressed through the entire course of germination. A set of genes were transiently up-regulated within three hours of imbibition, and might play roles in initiating biological pathways involved in seed germination. However, highly abundant transcripts in dry barley and Arabidopsis seeds were significantly conserved. Comparison with transcriptomes of barley aleurone in response to GA and ABA identified three sets of germination responsive genes that were regulated coordinately by GA, antagonistically by ABA, and coordinately by GA but antagonistically by ABA. Major CHO metabolism, cell wall degradation and protein degradation pathways were up-regulated by both GA and seed

  18. Transcriptional regulatory programs underlying barley germination and regulatory functions of Gibberellin and abscisic acid

    Directory of Open Access Journals (Sweden)

    Lin Li

    2011-06-01

    Full Text Available Abstract Background Seed germination is a complex multi-stage developmental process, and mainly accomplished through concerted activities of many gene products and biological pathways that are often subjected to strict developmental regulation. Gibberellins (GA and abscisic acid (ABA are two key phytohormones regulating seed germination and seedling growth. However, transcriptional regulatory networks underlying seed germination and its associated biological pathways are largely unknown. Results The studies examined transcriptomes of barley representing six distinct and well characterized germination stages and revealed that the transcriptional regulatory program underlying barley germination was composed of early, late, and post-germination phases. Each phase was accompanied with transcriptional up-regulation of distinct biological pathways. Cell wall synthesis and regulatory components including transcription factors, signaling and post-translational modification components were specifically and transiently up-regulated in early germination phase while histone families and many metabolic pathways were up-regulated in late germination phase. Photosynthesis and seed reserve mobilization pathways were up-regulated in post-germination phase. However, stress related pathways and seed storage proteins were suppressed through the entire course of germination. A set of genes were transiently up-regulated within three hours of imbibition, and might play roles in initiating biological pathways involved in seed germination. However, highly abundant transcripts in dry barley and Arabidopsis seeds were significantly conserved. Comparison with transcriptomes of barley aleurone in response to GA and ABA identified three sets of germination responsive genes that were regulated coordinately by GA, antagonistically by ABA, and coordinately by GA but antagonistically by ABA. Major CHO metabolism, cell wall degradation and protein degradation pathways were up

  19. Global transcriptional regulatory network for Escherichia coli robustly connects gene expression to transcription factor activities

    DEFF Research Database (Denmark)

    Fang, Xin; Sastry, Anand; Mih, Nathan

    2017-01-01

    Transcriptional regulatory networks (TRNs) have been studied intensely for >25 y. Yet, even for the Escherichia coli TRN-probably the best characterized TRN-several questions remain. Here, we address three questions: (i) How complete is our knowledge of the E. coli TRN; (ii) how well can we predi...

  20. Transcriptional and Post-Transcriptional Mechanisms of the Development of Neocortical Lamination

    Directory of Open Access Journals (Sweden)

    Tatiana Popovitchenko

    2017-11-01

    Full Text Available The neocortex is a laminated brain structure that is the seat of higher cognitive capacity and responses, long-term memory, sensory and emotional functions, and voluntary motor behavior. Proper lamination requires that progenitor cells give rise to a neuron, that the immature neuron can migrate away from its mother cell and past other cells, and finally that the immature neuron can take its place and adopt a mature identity characterized by connectivity and gene expression; thus lamination proceeds through three steps: genesis, migration, and maturation. Each neocortical layer contains pyramidal neurons that share specific morphological and molecular characteristics that stem from their prenatal birth date. Transcription factors are dynamic proteins because of the cohort of downstream factors that they regulate. RNA-binding proteins are no less dynamic, and play important roles in every step of mRNA processing. Indeed, recent screens have uncovered post-transcriptional mechanisms as being integral regulatory mechanisms to neocortical development. Here, we summarize major aspects of neocortical laminar development, emphasizing transcriptional and post-transcriptional mechanisms, with the aim of spurring increased understanding and study of its intricacies.

  1. Heregulin Co-opts PR Transcriptional Action Via Stat3 Role As a Coregulator to Drive Cancer Growth.

    Science.gov (United States)

    Proietti, Cecilia J; Izzo, Franco; Díaz Flaqué, María Celeste; Cordo Russo, Rosalía; Venturutti, Leandro; Mercogliano, María Florencia; De Martino, Mara; Pineda, Viviana; Muñoz, Sergio; Guzmán, Pablo; Roa, Juan C; Schillaci, Roxana; Elizalde, Patricia V

    2015-10-01

    Accumulated findings have demonstrated the presence of bidirectional interactions between progesterone receptor (PR) and the ErbB family of receptor tyrosine kinases signaling pathways in breast cancer. We previously revealed signal transducer and activator of transcription 3 (Stat3) as a nodal convergence point between said signaling pathways proving that Stat3 is activated by one of the ErbBs' ligands, heregulin (HRG)β1 via ErbB2 and through the co-option of PR as a signaling molecule. Here, we found that HRGβ1 induced Stat3 recruitment to the promoters of the progestin-regulated cell cycle modulators Bcl-XL and p21(CIP1) and also stimulated Stat3 binding to the mouse mammary tumor virus promoter, which carries consensus progesterone response elements. Interestingly, HRGβ1-activated Stat3 displayed differential functions on PR activity depending on the promoter bound. Indeed, Stat3 was required for PR binding in bcl-X, p21(CIP1), and c-myc promoters while exerting a PR coactivator function on the mouse mammary tumor virus promoter. Stat3 also proved to be necessary for HRGβ1-induced in vivo tumor growth. Our results endow Stat3 a novel function as a coregulator of HRGβ1-activated PR to promote breast cancer growth. These findings underscore the importance of understanding the complex interactions between PR and other regulatory factors, such as Stat3, that contribute to determine the context-dependent transcriptional actions of PR.

  2. Heregulin Co-opts PR Transcriptional Action Via Stat3 Role As a Coregulator to Drive Cancer Growth

    Science.gov (United States)

    Izzo, Franco; Díaz Flaqué, María Celeste; Cordo Russo, Rosalía; Venturutti, Leandro; Mercogliano, María Florencia; De Martino, Mara; Pineda, Viviana; Muñoz, Sergio; Guzmán, Pablo; Roa, Juan C.; Schillaci, Roxana

    2015-01-01

    Accumulated findings have demonstrated the presence of bidirectional interactions between progesterone receptor (PR) and the ErbB family of receptor tyrosine kinases signaling pathways in breast cancer. We previously revealed signal transducer and activator of transcription 3 (Stat3) as a nodal convergence point between said signaling pathways proving that Stat3 is activated by one of the ErbBs' ligands, heregulin (HRG)β1 via ErbB2 and through the co-option of PR as a signaling molecule. Here, we found that HRGβ1 induced Stat3 recruitment to the promoters of the progestin-regulated cell cycle modulators Bcl-XL and p21CIP1 and also stimulated Stat3 binding to the mouse mammary tumor virus promoter, which carries consensus progesterone response elements. Interestingly, HRGβ1-activated Stat3 displayed differential functions on PR activity depending on the promoter bound. Indeed, Stat3 was required for PR binding in bcl-X, p21CIP1, and c-myc promoters while exerting a PR coactivator function on the mouse mammary tumor virus promoter. Stat3 also proved to be necessary for HRGβ1-induced in vivo tumor growth. Our results endow Stat3 a novel function as a coregulator of HRGβ1-activated PR to promote breast cancer growth. These findings underscore the importance of understanding the complex interactions between PR and other regulatory factors, such as Stat3, that contribute to determine the context-dependent transcriptional actions of PR. PMID:26340407

  3. A nuclear factor I-like activity and a liver-specific repressor govern estrogen-regulated in vitro transcription from the Xenopus laevis vitellogenin B1 promoter.

    OpenAIRE

    Corthésy, B; Cardinaux, J R; Claret, F X; Wahli, W

    1989-01-01

    A hormone-controlled in vitro transcription system derived from Xenopus liver nuclear extracts was exploited to identify novel cis-acting elements within the vitellogenin gene B1 promoter region. In addition to the already well-documented estrogen-responsive element (ERE), two elements were found within the 140 base pairs upstream of the transcription initiation site. One of them, a negative regulatory element, is responsible for the lack of promoter activity in the absence of the hormone and...

  4. Transcriptional interference networks coordinate the expression of functionally-related genes clustered in the same genomic loci

    Directory of Open Access Journals (Sweden)

    Zsolt eBoldogkoi

    2012-07-01

    Full Text Available The regulation of gene expression is essential for normal functioning of biological systems in every form of life. Gene expression is primarily controlled at the level of transcription, especially at the phase of initiation. Non-coding RNAs are one of the major players at every level of genetic regulation, including the control of chromatin organisation, transcription, various post-transcriptional processes and translation. In this study, the Transcriptional Interference Network (TIN hypothesis was put forward in an attempt to explain the global expression of antisense RNAs and the overall occurrence of tandem gene clusters in the genomes of various biological systems ranging from viruses to mammalian cells. The TIN hypothesis suggests the existence of a novel layer of genetic regulation, based on the interactions between the transcriptional machineries of neighbouring genes at their overlapping regions, which are assumed to play a fundamental role in coordinating gene expression within a cluster of functionally-linked genes. It is claimed that the transcriptional overlaps between adjacent genes are much more widespread in genomes than is thought today. The Waterfall model of the TIN hypothesis postulates a unidirectional effect of upstream genes on the transcription of downstream genes within a cluster of tandemly-arrayed genes, while the Seesaw model proposes a mutual interdependence of gene expression between the oppositely-oriented genes. The TIN represents an auto-regulatory system with an exquisitely timed and highly synchronised cascade of gene expression in functionally-linked genes located in close physical proximity to each other. In this study, we focused on herpesviruses. The reason for this lies in the compressed nature of viral genes, which allows a tight regulation and an easier investigation of the transcriptional interactions between genes. However, I believe that the same or similar principles can be applied to cellular

  5. Coordinated Evolution of Transcriptional and Post-Transcriptional Regulation for Mitochondrial Functions in Yeast Strains.

    Directory of Open Access Journals (Sweden)

    Xuepeng Sun

    Full Text Available Evolution of gene regulation has been proposed to play an important role in environmental adaptation. Exploring mechanisms underlying coordinated evolutionary changes at various levels of gene regulation could shed new light on how organism adapt in nature. In this study, we focused on regulatory differences between a laboratory Saccharomyces cerevisiae strain BY4742 and a pathogenic S. cerevisiae strain, YJM789. The two strains diverge in many features, including growth rate, morphology, high temperature tolerance, and pathogenicity. Our RNA-Seq and ribosomal footprint profiling data showed that gene expression differences are pervasive, and genes functioning in mitochondria are mostly divergent between the two strains at both transcriptional and translational levels. Combining functional genomics data from other yeast strains, we further demonstrated that significant divergence of expression for genes functioning in the electron transport chain (ETC was likely caused by differential expression of a transcriptional factor, HAP4, and that post-transcriptional regulation mediated by an RNA-binding protein, PUF3, likely led to expression divergence for genes involved in mitochondrial translation. We also explored mito-nuclear interactions via mitochondrial DNA replacement between strains. Although the two mitochondrial genomes harbor substantial sequence divergence, neither growth nor gene expression were affected by mitochondrial DNA replacement in both fermentative and respiratory growth media, indicating compatible mitochondrial and nuclear genomes between these two strains in the tested conditions. Collectively, we used mitochondrial functions as an example to demonstrate for the first time that evolution at both transcriptional and post-transcriptional levels could lead to coordinated regulatory changes underlying strain specific functional variations.

  6. Transcription Factors Expressed in Lateral Organ Boundaries: Identification of Downstream Targets

    Energy Technology Data Exchange (ETDEWEB)

    Springer, Patricia S

    2010-07-12

    The processes of lateral organ initiation and patterning are central to the generation of mature plant form. Characterization of the molecular mechanisms underlying these processes is essential to our understanding of plant development. Communication between the shoot apical meristem and initiating organ primordia is important both for functioning of the meristem and for proper organ patterning, and very little is known about this process. In particular, the boundary between meristem and leaf is emerging as a critical region that is important for SAM maintenance and regulation of organogenesis. The goal of this project was to characterize three boundary-expressed genes that encode predicted transcription factors. Specifically, we have studied LATERAL ORGAN BOUNDARIES (LOB), LATERAL ORGAN FUSION1 (LOF1), and LATERAL ORGAN FUSION2 (LOF2). LOB encodes the founding member of the LOB-DOMAIN (LBD) plant-specific DNA binding transcription factor family and LOF1 and LOF2 encode paralogous MYB-domain transcription factors. We characterized the genetic relationship between these three genes and other boundary and meristem genes. We also used an ectopic inducible expression system to identify direct targets of LOB.

  7. The Role of Epigenetic Regulation in Transcriptional Memory in the Immune System.

    Science.gov (United States)

    Woodworth, A M; Holloway, A F

    The immune system is exquisitely poised to identify, respond to, and eradicate pathogens from the body, as well as to produce a more rapid and augmented response to a subsequent encounter with the pathogen. These cellular responses rely on the highly coordinated and rapid activation of gene expression programs as well as the ability of the cell to retain a memory of the initial gene response. It is clear that chromatin structure and epigenetic mechanisms play a crucial role in determining these gene responses, and in fact the immune system has proved an instructive model for investigating the multifaceted mechanisms through which the chromatin landscape contributes to gene expression programs. These mechanisms include modifications to the DNA and histone proteins, the positioning, composition, and remodeling of nucleosomes, as well as the formation of higher-order chromatin structures. Moreover, it is now apparent that epigenetic mechanisms also provide an instrument by which cells can retain memory of the initial transcriptional response, "priming" the genome so that it can respond more quickly to subsequent exposure to the signal. Here, we use the immune system as a model to demonstrate the complex interplay between transcription factors and the chromatin landscape required to orchestrate precise gene responses to external stimuli and further to demonstrate how these interactions can establish memory of past transcriptional events. We focus on what we have learnt from the immune system and how this can inform our understanding of other cellular systems. © 2017 Elsevier Inc. All rights reserved.

  8. Preliminary crystallographic analysis of a possible transcription factor encoded by the mimivirus L544 gene

    International Nuclear Information System (INIS)

    Ciaccafava, Alexandre; Lartigue, Audrey; Mansuelle, Pascal; Jeudy, Sandra; Abergel, Chantal

    2011-01-01

    The mimivirus L544 gene product was expressed in E. coli and crystallized; preliminary phasing of a MAD data set was performed using the selenium signal present in a crystal of recombinant selenomethionine-substituted protein. Mimivirus is the prototype of a new family (the Mimiviridae) of nucleocytoplasmic large DNA viruses (NCLDVs), which already include the Poxviridae, Iridoviridae, Phycodnaviridae and Asfarviridae. Mimivirus specifically replicates in cells from the genus Acanthamoeba. Proteomic analysis of purified mimivirus particles revealed the presence of many subunits of the DNA-directed RNA polymerase II complex. A fully functional pre-transcriptional complex appears to be loaded in the virions, allowing mimivirus to initiate transcription within the host cytoplasm immediately upon infection independently of the host nuclear apparatus. To fully understand this process, a systematic study of mimivirus proteins that are predicted (by bioinformatics) or suspected (by proteomic analysis) to be involved in transcription was initiated by cloning and expressing them in Escherichia coli in order to determine their three-dimensional structures. Here, preliminary crystallographic analysis of the recombinant L544 protein is reported. The crystals belonged to the orthorhombic space group C222 1 with one monomer per asymmetric unit. A MAD data set was used for preliminary phasing using the selenium signal present in a selenomethionine-substituted protein crystal

  9. The effect of U.V.-irradiation on lambda DNA transcription

    International Nuclear Information System (INIS)

    Ranade, S.S.

    1977-01-01

    The effect of U.V.-irradiation of template DNA has been studied in vitro in the E.coli RNA polymerase system with native and U.V.-treated lambda DNA. Lambda DNA was more susceptible to U.V. than was calf-thymus DNA, yet a residual activity was observed at a U.V. dose of 0.5 x 10 4 erg/mm 2 . From the kinetic analysis of the reaction and the incorporation of lambda 32 P-labelled nucleoside triphosphates, it seems reasonable to conclude that U.V.-irradiation probably did not affect the DNA initiation sites, recognizable by RNA polymerase. The transcription products made with U.V.-irradiated lambda DNA were asymmetrical, and hybridized to the right half (R) and the left half (L) of lambda DNA with the ratio of R/L=4/1, and they showed a lower hybridizability than the transcripts with native lambda DNA. The initiation sites recognizable by RNA polymerase seemed to be the same on both native and U.V.-irradiated lambda DNA, though the transcription of U.V.-treated lambda DNA appeared to terminate with rather short RNA chains. (author)

  10. A regulatory transcriptional loop controls proliferation and differentiation in Drosophila neural stem cells.

    Directory of Open Access Journals (Sweden)

    Tetsuo Yasugi

    Full Text Available Neurogenesis is initiated by a set of basic Helix-Loop-Helix (bHLH transcription factors that specify neural progenitors and allow them to generate neurons in multiple rounds of asymmetric cell division. The Drosophila Daughterless (Da protein and its mammalian counterparts (E12/E47 act as heterodimerization factors for proneural genes and are therefore critically required for neurogenesis. Here, we demonstrate that Da can also be an inhibitor of the neural progenitor fate whose absence leads to stem cell overproliferation and tumor formation. We explain this paradox by demonstrating that Da induces the differentiation factor Prospero (Pros whose asymmetric segregation is essential for differentiation in one of the two daughter cells. Da co-operates with the bHLH transcription factor Asense, whereas the other proneural genes are dispensible. After mitosis, Pros terminates Asense expression in one of the two daughter cells. In da mutants, pros is not expressed, leading to the formation of lethal transplantable brain tumors. Our results define a transcriptional feedback loop that regulates the balance between self-renewal and differentiation in Drosophila optic lobe neuroblasts. They indicate that initiation of a neural differentiation program in stem cells is essential to prevent tumorigenesis.

  11. Hanford tanks initiative plan

    International Nuclear Information System (INIS)

    McKinney, K.E.

    1997-01-01

    Abstract: The Hanford Tanks Initiative (HTI) is a five-year project resulting from the technical and financial partnership of the U.S. Department of Energy's Office of Waste Management (EM-30) and Office of Science and Technology Development (EM-50). The HTI project accelerates activities to gain key technical, cost performance, and regulatory information on two high-level waste tanks. The HTI will provide a basis for design and regulatory decisions affecting the remainder of the Tank Waste Remediation System's tank waste retrieval Program

  12. UNLV Nuclear Hydrogen Initiative

    Energy Technology Data Exchange (ETDEWEB)

    Hechanova, Anthony E.; Johnson, Allen; O' Toole, Brendan; Trabia, Mohamed; Peterson, Per

    2012-10-25

    Evaluation of the Crack growth rate (CGR) of Alloy 617 and Alloy 276 under constant K at ambient temperature has been completed. Creep deformation of Alloy 230 at different temperature range and load level has been completed and heat to heat variation has been noticed. Creep deformation study of Alloy 276 has been completed under an applied initial stress level of 10% of yield stress at 950ºC. The grain size evaluation of the tested creep specimens of Alloy 276 has been completed.

  13. Feedback stabilization initiative

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1997-06-01

    Much progress has been made in attaining high confinement regimes in magnetic confinement devices. These operating modes tend to be transient, however, due to the onset of MHD instabilities, and their stabilization is critical for improved performance at steady state. This report describes the Feedback Stabilization Initiative (FSI), a broad-based, multi-institutional effort to develop and implement methods for raising the achievable plasma betas through active MHD feedback stabilization. A key element in this proposed effort is the Feedback Stabilization Experiment (FSX), a medium-sized, national facility that would be specifically dedicated to demonstrating beta improvement in reactor relevant plasmas by using a variety of MHD feedback stabilization schemes.

  14. Feedback stabilization initiative

    International Nuclear Information System (INIS)

    1997-06-01

    Much progress has been made in attaining high confinement regimes in magnetic confinement devices. These operating modes tend to be transient, however, due to the onset of MHD instabilities, and their stabilization is critical for improved performance at steady state. This report describes the Feedback Stabilization Initiative (FSI), a broad-based, multi-institutional effort to develop and implement methods for raising the achievable plasma betas through active MHD feedback stabilization. A key element in this proposed effort is the Feedback Stabilization Experiment (FSX), a medium-sized, national facility that would be specifically dedicated to demonstrating beta improvement in reactor relevant plasmas by using a variety of MHD feedback stabilization schemes

  15. Initiation of slug flow

    Energy Technology Data Exchange (ETDEWEB)

    Hanratty, T.J.; Woods, B.D. [Univ. of Illinois, Urbana, IL (United States)

    1995-12-31

    The initiation of slug flow in a horizontal pipe can be predicted either by considering the stability of a slug or by considering the stability of a stratified flow. Measurements of the shedding rate of slugs are used to define necessary conditions for the existence of a slug. Recent results show that slugs develop from an unstable stratified flow through the evolution of small wavelength waves into large wavelength waves that have the possibility of growing to form a slug. The mechanism appears to be quite different for fluids with viscosities close to water than for fluids with large viscosities (20 centipoise).

  16. Stirling to Flight Initiative

    Science.gov (United States)

    Hibbard, Kenneth E.; Mason, Lee S.; Ndu, Obi; Smith, Clayton; Withrow, James P.

    2016-01-01

    Flight (S2F) initiative with the objective of developing a 100-500 We Stirling generator system. Additionally, a different approach is being devised for this initiative to avoid pitfalls of the past, and apply lessons learned from the recent ASRG experience. Two key aspects of this initiative are a Stirling System Technology Maturation Effort, and a Surrogate Mission Team (SMT) intended to provide clear mission pull and requirements context. The S2F project seeks to lead directly into a DOE flight system development of a new SRG. This paper will detail the proposed S2F initiative, and provide specifics on the key efforts designed to pave a forward path for bringing Stirling technology to flight.

  17. Initiating statistical maintenance optimization

    International Nuclear Information System (INIS)

    Doyle, E. Kevin; Tuomi, Vesa; Rowley, Ian

    2007-01-01

    Since the 1980 s maintenance optimization has been centered around various formulations of Reliability Centered Maintenance (RCM). Several such optimization techniques have been implemented at the Bruce Nuclear Station. Further cost refinement of the Station preventive maintenance strategy includes evaluation of statistical optimization techniques. A review of successful pilot efforts in this direction is provided as well as initial work with graphical analysis. The present situation reguarding data sourcing, the principle impediment to use of stochastic methods in previous years, is discussed. The use of Crowe/AMSAA (Army Materials Systems Analysis Activity) plots is demonstrated from the point of view of justifying expenditures in optimization efforts. (author)

  18. Transcriptional Programs Controlling Perinatal Lung Maturation

    Science.gov (United States)

    Xu, Yan; Wang, Yanhua; Besnard, Valérie; Ikegami, Machiko; Wert, Susan E.; Heffner, Caleb; Murray, Stephen A.; Donahue, Leah Rae; Whitsett, Jeffrey A.

    2012-01-01

    The timing of lung maturation is controlled precisely by complex genetic and cellular programs. Lung immaturity following preterm birth frequently results in Respiratory Distress Syndrome (RDS) and Broncho-Pulmonary Dysplasia (BPD), which are leading causes of mortality and morbidity in preterm infants. Mechanisms synchronizing gestational length and lung maturation remain to be elucidated. In this study, we designed a genome-wide mRNA expression time-course study from E15.5 to Postnatal Day 0 (PN0) using lung RNAs from C57BL/6J (B6) and A/J mice that differ in gestational length by ∼30 hr (B6controlling lung maturation. We identified both temporal and strain dependent gene expression patterns during lung maturation. For time dependent changes, cell adhesion, vasculature development, and lipid metabolism/transport were major bioprocesses induced during the saccular stage of lung development at E16.5–E17.5. CEBPA, PPARG, VEGFA, CAV1 and CDH1 were found to be key signaling and transcriptional regulators of these processes. Innate defense/immune responses were induced at later gestational ages (E18.5–20.5), STAT1, AP1, and EGFR being important regulators of these responses. Expression of RNAs associated with the cell cycle and chromatin assembly was repressed during prenatal lung maturation and was regulated by FOXM1, PLK1, chromobox, and high mobility group families of transcription factors. Strain dependent lung mRNA expression differences peaked at E18.5. At this time, mRNAs regulating surfactant and innate immunity were more abundantly expressed in lungs of B6 (short gestation) than in A/J (long gestation) mice, while expression of genes involved in chromatin assembly and histone modification were expressed at lower levels in B6 than in A/J mice. The present study systemically mapped key regulators, bioprocesses, and transcriptional networks controlling lung maturation, providing the basis for new therapeutic strategies to enhance lung function in preterm

  19. Initiating Service Encounter-based Innovation by Word-of-Business

    DEFF Research Database (Denmark)

    Mattsson, Jan

    2015-01-01

    Purpose – This paper aims to set up a natural experiment as action research and to develop a framework of cognitive distance of informants to improve the initiation of service encounter-based innovation. Design/methodology/approach – Natural experiment as action research in one Scandinavian case...... transcriptions of interviews and transcriptions. Research limitations/implications – Only one Scandinavian company and a limited number of informants were activated. Also, the time period only included the initiation phase of service encounter-based innovation. Practical implications – Three different strategies...... in an emerging innovation field, open/user-driven innovation. Theory from business marketing, service encounter and innovation is also used....

  20. Amino acid-dependent signaling via S6K1 and MYC is essential for regulation of rDNA transcription

    Science.gov (United States)

    Kang, Jian; Kusnadi, Eric P.; Ogden, Allison J.; Hicks, Rodney J.; Bammert, Lukas; Kutay, Ulrike; Hung, Sandy; Sanij, Elaine; Hannan, Ross D.; Hannan, Katherine M.; Pearson, Richard B.

    2016-01-01

    Dysregulation of RNA polymerase I (Pol I)-dependent ribosomal DNA (rDNA) transcription is a consistent feature of malignant transformation that can be targeted to treat cancer. Understanding how rDNA transcription is coupled to the availability of growth factors and nutrients will provide insight into how ribosome biogenesis is maintained in a tumour environment characterised by limiting nutrients. We demonstrate that modulation of rDNA transcription initiation, elongation and rRNA processing is an immediate, co-regulated response to altered amino acid abundance, dependent on both mTORC1 activation of S6K1 and MYC activity. Growth factors regulate rDNA transcription initiation while amino acids modulate growth factor-dependent rDNA transcription by primarily regulating S6K1-dependent rDNA transcription elongation and processing. Thus, we show for the first time amino acids regulate rRNA synthesis by a distinct, post-initiation mechanism, providing a novel model for integrated control of ribosome biogenesis that has implications for understanding how this process is dysregulated in cancer. PMID:27385002

  1. Detecting novel low-abundant transcripts in Drosophila

    DEFF Research Database (Denmark)

    Lee, Sanggyu; Bao, Jingyue; Zhou, Guolin

    2005-01-01

    Increasing evidence suggests that low-abundant transcripts may play fundamental roles in biological processes. In an attempt to estimate the prevalence of low-abundant transcripts in eukaryotic genomes, we performed a transcriptome analysis in Drosophila using the SAGE technique. We collected 244......,313 SAGE tags from transcripts expressed in Drosophila embryonic, larval, pupae, adult, and testicular tissue. From these SAGE tags, we identified 40,823 unique SAGE tags. Our analysis showed that 55% of the 40,823 unique SAGE tags are novel without matches in currently known Drosophila transcripts...... in the Drosophila genome. Our study reveals the presence of a significant number of novel low-abundant transcripts in Drosophila, and highlights the need to isolate these novel low-abundant transcripts for further biological studies. Udgivelsesdato: 2005-Jun...

  2. The LHCb Starterkit initiative

    CERN Document Server

    Puig Navarro, Albert

    2017-01-01

    The vast majority of high-energy physicists use and produce software every day. Software skills are usually acquired on the go and dedicated training courses are rare. The LHCb Starterkit is a new training format for getting LHCb collaborators started in effectively using software to perform their research. The initiative, combining courses and online tutorials, focuses on teaching basic skills for research computing, as well as LHCb software specifics. Unlike traditional tutorials we focus on starting with basics, performing all the material live, with a high degree of interactivity, giving priority to understanding the tools as opposed to handing out recipes that work “as if by magic”. The LHCb Starterkit was started by young members of the collaboration inspired by the principles of Software Carpentry, and the material is created in a collaborative fashion using the tools we teach. Three successful entry-level workshops, as well as two advanced ones, have taken place since the start of the initiative i...

  3. Combat Wound Initiative program.

    Science.gov (United States)

    Stojadinovic, Alexander; Elster, Eric; Potter, Benjamin K; Davis, Thomas A; Tadaki, Doug K; Brown, Trevor S; Ahlers, Stephen; Attinger, Christopher E; Andersen, Romney C; Burris, David; Centeno, Jose; Champion, Hunter; Crumbley, David R; Denobile, John; Duga, Michael; Dunne, James R; Eberhardt, John; Ennis, William J; Forsberg, Jonathan A; Hawksworth, Jason; Helling, Thomas S; Lazarus, Gerald S; Milner, Stephen M; Mullick, Florabel G; Owner, Christopher R; Pasquina, Paul F; Patel, Chirag R; Peoples, George E; Nissan, Aviram; Ring, Michael; Sandberg, Glenn D; Schaden, Wolfgang; Schultz, Gregory S; Scofield, Tom; Shawen, Scott B; Sheppard, Forest R; Stannard, James P; Weina, Peter J; Zenilman, Jonathan M

    2010-07-01

    The Combat Wound Initiative (CWI) program is a collaborative, multidisciplinary, and interservice public-private partnership that provides personalized, state-of-the-art, and complex wound care via targeted clinical and translational research. The CWI uses a bench-to-bedside approach to translational research, including the rapid development of a human extracorporeal shock wave therapy (ESWT) study in complex wounds after establishing the potential efficacy, biologic mechanisms, and safety of this treatment modality in a murine model. Additional clinical trials include the prospective use of clinical data, serum and wound biomarkers, and wound gene expression profiles to predict wound healing/failure and additional clinical patient outcomes following combat-related trauma. These clinical research data are analyzed using machine-based learning algorithms to develop predictive treatment models to guide clinical decision-making. Future CWI directions include additional clinical trials and study centers and the refinement and deployment of our genetically driven, personalized medicine initiative to provide patient-specific care across multiple medical disciplines, with an emphasis on combat casualty care.

  4. Initiatives for proliferation prevention

    International Nuclear Information System (INIS)

    1997-04-01

    Preventing the proliferation of weapons of mass destruction is a central part of US national security policy. A principal instrument of the Department of Energy's (DOE's) program for securing weapons of mass destruction technology and expertise and removing incentives for scientists, engineers and technicians in the newly independent states (NIS) of the former Soviet Union to go to rogue countries or assist terrorist groups is the Initiatives for Proliferation Prevention (IPP). IPP was initiated pursuant to the 1994 Foreign Operations Appropriations Act. IPP is a nonproliferation program with a commercialization strategy. IPP seeks to enhance US national security and to achieve nonproliferation objectives by engaging scientists, engineers and technicians from former NIS weapons institutes; redirecting their activities in cooperatively-developed, commercially viable non-weapons related projects. These projects lead to commercial and economic benefits for both the NIS and the US IPP projects are funded in Russian, Ukraine, Kazakhstan and Belarus. This booklet offers an overview of the IPP program as well as a sampling of some of the projects which are currently underway

  5. Automatic Phonetic Transcription for Danish Speech Recognition

    DEFF Research Database (Denmark)

    Kirkedal, Andreas Søeborg

    , like Danish, the graphemic and phonetic representations are very dissimilar and more complex rewriting rules must be applied to create the correct phonetic representation. Automatic phonetic transcribers use different strategies, from deep analysis to shallow rewriting rules, to produce phonetic......, syllabication, stød and several other suprasegmental features (Kirkedal, 2013). Simplifying the transcriptions by filtering out the symbols for suprasegmental features in a post-processing step produces a format that is suitable for ASR purposes. eSpeak is an open source speech synthesizer originally created...... for particular words and word classes in addition. In comparison, English has 5,852 spelling-tophoneme rules and 4,133 additional rules and 8,278 rules and 3,829 additional rules. Phonix applies deep morphological analysis as a preprocessing step. Should the analysis fail, several fallback strategies...

  6. A transcription factor for cold sensation!

    Directory of Open Access Journals (Sweden)

    Milbrandt Jeffrey

    2005-03-01

    Full Text Available Abstract The ability to feel hot and cold is critical for animals and human beings to survive in the natural environment. Unlike other sensations, the physiology of cold sensation is mostly unknown. In the present study, we use genetically modified mice that do not express nerve growth factor-inducible B (NGFIB to investigate the possible role of NGFIB in cold sensation. We found that genetic deletion of NGFIB selectively affected behavioral responses to cold stimuli while behavioral responses to noxious heat or mechanical stimuli were normal. Furthermore, behavioral responses remained reduced or blocked in NGFIB knockout mice even after repetitive application of cold stimuli. Our results provide strong evidence that the first transcription factor NGFIB determines the ability of animals to respond to cold stimulation.

  7. Transcriptional regulation of c-fos

    International Nuclear Information System (INIS)

    Prywes, R.; Fisch, T.M.; Roeder, R.G.

    1988-01-01

    Expression of the c-fos proto-oncogene is induced rapidly and transiently by serum and other mitogenic agents. This rapid induction is therefore likely to involve posttranslational modifications and provides an excellent model for an early nuclear target of the signal transduction process, growth factors that bind to tyrosine kinase receptors. The authors have sought to understand the mechanism of transcriptional induction by each of these agents. The first step in this process was to identify the sequence elements in the c-fos gene responsible for induction by each of these agents. A specific element, termed serum response element (SRE), has been identified by transfection experiments of c-fos promoter constructs. To study regulation via SRE, a nuclear factor that binds to the SRE, termed serum response factor (SRF), has been identified with the gel mobility shift assay

  8. An anatomic transcriptional atlas of human glioblastoma.

    Science.gov (United States)

    Puchalski, Ralph B; Shah, Nameeta; Miller, Jeremy; Dalley, Rachel; Nomura, Steve R; Yoon, Jae-Guen; Smith, Kimberly A; Lankerovich, Michael; Bertagnolli, Darren; Bickley, Kris; Boe, Andrew F; Brouner, Krissy; Butler, Stephanie; Caldejon, Shiella; Chapin, Mike; Datta, Suvro; Dee, Nick; Desta, Tsega; Dolbeare, Tim; Dotson, Nadezhda; Ebbert, Amanda; Feng, David; Feng, Xu; Fisher, Michael; Gee, Garrett; Goldy, Jeff; Gourley, Lindsey; Gregor, Benjamin W; Gu, Guangyu; Hejazinia, Nika; Hohmann, John; Hothi, Parvinder; Howard, Robert; Joines, Kevin; Kriedberg, Ali; Kuan, Leonard; Lau, Chris; Lee, Felix; Lee, Hwahyung; Lemon, Tracy; Long, Fuhui; Mastan, Naveed; Mott, Erika; Murthy, Chantal; Ngo, Kiet; Olson, Eric; Reding, Melissa; Riley, Zack; Rosen, David; Sandman, David; Shapovalova, Nadiya; Slaughterbeck, Clifford R; Sodt, Andrew; Stockdale, Graham; Szafer, Aaron; Wakeman, Wayne; Wohnoutka, Paul E; White, Steven J; Marsh, Don; Rostomily, Robert C; Ng, Lydia; Dang, Chinh; Jones, Allan; Keogh, Bart; Gittleman, Haley R; Barnholtz-Sloan, Jill S; Cimino, Patrick J; Uppin, Megha S; Keene, C Dirk; Farrokhi, Farrokh R; Lathia, Justin D; Berens, Michael E; Iavarone, Antonio; Bernard, Amy; Lein, Ed; Phillips, John W; Rostad, Steven W; Cobbs, Charles; Hawrylycz, Michael J; Foltz, Greg D

    2018-05-11

    Glioblastoma is an aggressive brain tumor that carries a poor prognosis. The tumor's molecular and cellular landscapes are complex, and their relationships to histologic features routinely used for diagnosis are unclear. We present the Ivy Glioblastoma Atlas, an anatomically based transcriptional atlas of human glioblastoma that aligns individual histologic features with genomic alterations and gene expression patterns, thus assigning molecular information to the most important morphologic hallmarks of the tumor. The atlas and its clinical and genomic database are freely accessible online data resources that will serve as a valuable platform for future investigations of glioblastoma pathogenesis, diagnosis, and treatment. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  9. Single molecule transcription profiling with AFM

    International Nuclear Information System (INIS)

    Reed, Jason; Mishra, Bud; Pittenger, Bede; Magonov, Sergei; Troke, Joshua; Teitell, Michael A; Gimzewski, James K

    2007-01-01

    Established techniques for global gene expression profiling, such as microarrays, face fundamental sensitivity constraints. Due to greatly increasing interest in examining minute samples from micro-dissected tissues, including single cells, unorthodox approaches, including molecular nanotechnologies, are being explored in this application. Here, we examine the use of single molecule, ordered restriction mapping, combined with AFM, to measure gene transcription levels from very low abundance samples. We frame the problem mathematically, using coding theory, and present an analysis of the critical error sources that may serve as a guide to designing future studies. We follow with experiments detailing the construction of high density, single molecule, ordered restriction maps from plasmids and from cDNA molecules, using two different enzymes, a result not previously reported. We discuss these results in the context of our calculations

  10. High-mobility group (HMG) protein HMG-1 and TATA-binding protein-associated factor TAF(II)30 affect estrogen receptor-mediated transcriptional activation.

    Science.gov (United States)

    Verrier, C S; Roodi, N; Yee, C J; Bailey, L R; Jensen, R A; Bustin, M; Parl, F F

    1997-07-01

    The estrogen receptor (ER) belongs to a family of ligand-inducible nuclear receptors that exert their effects by binding to cis-acting DNA elements in the regulatory region of target genes. The detailed mechanisms by which ER interacts with the estrogen response element (ERE) and affects transcription still remain to be elucidated. To study the ER-ERE interaction and transcription initiation, we employed purified recombinant ER expressed in both the baculovirus-Sf9 and his-tagged bacterial systems. The effect of high-mobility group (HMG) protein HMG-1 and purified recombinant TATA-binding protein-associated factor TAF(II)30 on ER-ERE binding and transcription initiation were assessed by electrophoretic mobility shift assay and in vitro transcription from an ERE-containing template (pERE2LovTATA), respectively. We find that purified, recombinant ER fails to bind to ERE in spite of high ligand-binding activity and electrophoretic and immunological properties identical to ER in MCF-7 breast cancer cells. HMG-1 interacts with ER and promotes ER-ERE binding in a concentration- and time-dependent manner. The effectiveness of HMG-1 to stimulate ER-ERE binding in the electrophoretic mobility shift assay depends on the sequence flanking the ERE consensus as well as the position of the latter in the oligonucleotide. We find that TAF(II)30 has no effect on ER-ERE binding either alone or in combination with ER and HMG-1. Although HMG-1 promotes ER-ERE binding, it fails to stimulate transcription initiation either in the presence or absence of hormone. In contrast, TAF(II)30, while not affecting ER-ERE binding, stimulates transcription initiation 20-fold in the presence of HMG-1. These results indicate that HMG-1 and TAF(II)30 act in sequence, the former acting to promote ER-ERE binding followed by the latter to stimulate transcription initiation.

  11. Global Analysis of Photosynthesis Transcriptional Regulatory Networks

    Science.gov (United States)

    Imam, Saheed; Noguera, Daniel R.; Donohue, Timothy J.

    2014-01-01

    Photosynthesis is a crucial biological process that depends on the interplay of many components. This work analyzed the gene targets for 4 transcription factors: FnrL, PrrA, CrpK and MppG (RSP_2888), which are known or predicted to control photosynthesis in Rhodobacter sphaeroides. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) identified 52 operons under direct control of FnrL, illustrating its regulatory role in photosynthesis, iron homeostasis, nitrogen metabolism and regulation of sRNA synthesis. Using global gene expression analysis combined with ChIP-seq, we mapped the regulons of PrrA, CrpK and MppG. PrrA regulates ∼34 operons encoding mainly photosynthesis and electron transport functions, while CrpK, a previously uncharacterized Crp-family protein, regulates genes involved in photosynthesis and maintenance of iron homeostasis. Furthermore, CrpK and FnrL share similar DNA binding determinants, possibly explaining our observation of the ability of CrpK to partially compensate for the growth defects of a ΔFnrL mutant. We show that the Rrf2 family protein, MppG, plays an important role in photopigment biosynthesis, as part of an incoherent feed-forward loop with PrrA. Our results reveal a previously unrealized, high degree of combinatorial regulation of photosynthetic genes and significant cross-talk between their transcriptional regulators, while illustrating previously unidentified links between photosynthesis and the maintenance of iron homeostasis. PMID:25503406

  12. Global analysis of photosynthesis transcriptional regulatory networks.

    Directory of Open Access Journals (Sweden)

    Saheed Imam

    2014-12-01

    Full Text Available Photosynthesis is a crucial biological process that depends on the interplay of many components. This work analyzed the gene targets for 4 transcription factors: FnrL, PrrA, CrpK and MppG (RSP_2888, which are known or predicted to control photosynthesis in Rhodobacter sphaeroides. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq identified 52 operons under direct control of FnrL, illustrating its regulatory role in photosynthesis, iron homeostasis, nitrogen metabolism and regulation of sRNA synthesis. Using global gene expression analysis combined with ChIP-seq, we mapped the regulons of PrrA, CrpK and MppG. PrrA regulates ∼34 operons encoding mainly photosynthesis and electron transport functions, while CrpK, a previously uncharacterized Crp-family protein, regulates genes involved in photosynthesis and maintenance of iron homeostasis. Furthermore, CrpK and FnrL share similar DNA binding determinants, possibly explaining our observation of the ability of CrpK to partially compensate for the growth defects of a ΔFnrL mutant. We show that the Rrf2 family protein, MppG, plays an important role in photopigment biosynthesis, as part of an incoherent feed-forward loop with PrrA. Our results reveal a previously unrealized, high degree of combinatorial regulation of photosynthetic genes and significant cross-talk between their transcriptional regulators, while illustrating previously unidentified links between photosynthesis and the maintenance of iron homeostasis.

  13. Automatic generation of pictorial transcripts of video programs

    Science.gov (United States)

    Shahraray, Behzad; Gibbon, David C.

    1995-03-01

    An automatic authoring system for the generation of pictorial transcripts of video programs which are accompanied by closed caption information is presented. A number of key frames, each of which represents the visual information in a segment of the video (i.e., a scene), are selected automatically by performing a content-based sampling of the video program. The textual information is recovered from the closed caption signal and is initially segmented based on its implied temporal relationship with the video segments. The text segmentation boundaries are then adjusted, based on lexical analysis and/or caption control information, to account for synchronization errors due to possible delays in the detection of scene boundaries or the transmission of the caption information. The closed caption text is further refined through linguistic processing for conversion to lower- case with correct capitalization. The key frames and the related text generate a compact multimedia presentation of the contents of the video program which lends itself to efficient storage and transmission. This compact representation can be viewed on a computer screen, or used to generate the input to a commercial text processing package to generate a printed version of the program.

  14. Dynamic zebrafish interactome reveals transcriptional mechanisms of dioxin toxicity.

    Directory of Open Access Journals (Sweden)

    Andrey Alexeyenko

    2010-05-01

    Full Text Available In order to generate hypotheses regarding the mechanisms by which 2,3,7,8-tetrachlorodibenzo-p-dioxin (dioxin causes toxicity, we analyzed global gene expression changes in developing zebrafish embryos exposed to this potent toxicant in the context of a dynamic gene network. For this purpose, we also computationally inferred a zebrafish (Danio rerio interactome based on orthologs and interaction data from other eukaryotes.Using novel computational tools to analyze this interactome, we distinguished between dioxin-dependent and dioxin-independent interactions between proteins, and tracked the temporal propagation of dioxin-dependent transcriptional changes from a few genes that were altered initially, to large groups of biologically coherent genes at later times. The most notable processes altered at later developmental stages were calcium and iron metabolism, embryonic morphogenesis including neuronal and retinal development, a variety of mitochondria-related functions, and generalized stress response (not including induction of antioxidant genes. Within the interactome, many of these responses were connected to cytochrome P4501A (cyp1a as well as other genes that were dioxin-regulated one day after exposure. This suggests that cyp1a may play a key role initiating the toxic dysregulation of those processes, rather than serving simply as a passive marker of dioxin exposure, as suggested by earlier research.Thus, a powerful microarray experiment coupled with a flexible interactome and multi-pronged interactome tools (which are now made publicly available for microarray analysis and related work suggest the hypothesis that dioxin, best known in fish as a potent cardioteratogen, has many other targets. Many of these types of toxicity have been observed in mammalian species and are potentially caused by alterations to cyp1a.

  15. How salicylic acid takes transcriptional control over jasmonic acid signaling

    Directory of Open Access Journals (Sweden)

    Lotte eCaarls

    2015-03-01

    Full Text Available Transcriptional regulation is a central process in plant immunity. The induction or repression of defense genes is orchestrated by signaling networks that are directed by plant hormones of which salicylic acid (SA and jasmonic acid (JA are the major players. Extensive cross-communication between the hormone signaling pathways allows for fine tuning of transcriptional programs, determining resistance to invaders and trade-offs with plant development. Here, we give an overview of how SA can control transcriptional reprogramming of JA-induced genes in Arabidopsis thaliana. SA can influence activity and/or localization of transcriptional regulators by post-translational modifications of transcription factors and co-regulators. SA-induced redox changes, mediated by thioredoxins and glutaredoxins, modify transcriptional regulators that are involved in suppression of JA-dependent genes, such as NPR1 and TGA transcription factors, which affects their localization or DNA binding activity. Furthermore, SA can mediate sequestering of JA-responsive transcription factors away from their target genes by stalling them in the cytosol or in complexes with repressor proteins in the nucleus. SA also affects JA-induced transcription by inducing degradation of transcription factors with an activating role in JA signaling, as was shown for the ERF transcription factor ORA59. Additionally, SA can induce negative regulators, among which WRKY transcription factors, that can directly or indirectly inhibit JA-responsive gene expression. Finally, at the DNA level, modification of histones by SA-dependent factors can result in repression of JA-responsive genes. These diverse and complex regulatory mechanisms affect important signaling hubs in the integration of hormone signaling networks. Some pathogens have evolved effectors that highjack hormone crosstalk mechanisms for their own good, which are described in this review as well.

  16. Transcriptional mutagenesis: causes and involvement in tumor development

    Science.gov (United States)

    Brégeon, Damien; Doetsch, Paul W.

    2013-01-01

    The majority of normal cells in a human do not multiply continuously but are quiescent and devote most of their energy to gene transcription. When DNA damages in the transcribed strand of an active gene are bypassed by an RNA polymerase, they can miscode at the damaged site and produce mutant transcripts. This process known as transcriptional mutagenesis can lead to the production of mutant proteins that could be important in tumor development. PMID:21346784

  17. Characterization of human mesothelin transcripts in ovarian and pancreatic cancer

    International Nuclear Information System (INIS)

    Muminova, Zhanat E; Strong, Theresa V; Shaw, Denise R

    2004-01-01

    Mesothelin is an attractive target for cancer immunotherapy due to its restricted expression in normal tissues and high level expression in several tumor types including ovarian and pancreatic adenocarcinomas. Three mesothelin transcript variants have been reported, but their relative expression in normal tissues and tumors has been poorly characterized. The goal of the present study was to clarify which mesothelin transcript variants are commonly expressed in human tumors. Human genomic and EST nucleotide sequences in the public databases were used to evaluate sequences reported for the three mesothelin transcript variants in silico. Subsequently, RNA samples from normal ovary, ovarian and pancreatic carcinoma cell lines, and primary ovarian tumors were analyzed by reverse transcription-polymerase chain reaction (RT-PCR) and nucleotide sequencing to directly identify expressed transcripts. In silico comparisons of genomic DNA sequences with available EST sequences supported expression of mesothelin transcript variants 1 and 3, but there were no sequence matches for transcript variant 2. Newly-derived nucleotide sequences of RT-PCR products from tissues and cell lines corresponded to mesothelin transcript variant 1. Mesothelin transcript variant 2 was not detected. Transcript variant 3 was observed as a small percentage of total mesothelin amplification products from all studied cell lines and tissues. Fractionation of nuclear and cytoplasmic RNA indicated that variant 3 was present primarily in the nuclear fraction. Thus, mesothelin transcript variant 3 may represent incompletely processed hnRNA. Mesothelin transcript variant 1 represents the predominant mature mRNA species expressed by both normal and tumor cells. This conclusion should be important for future development of cancer immunotherapies, diagnostic tests, and gene microarray studies targeting mesothelin

  18. Plastic downregulation of the transcriptional repressor BCL6 during maturation of human dendritic cells

    International Nuclear Information System (INIS)

    Pantano, Serafino; Jarrossay, David; Saccani, Simona; Bosisio, Daniela; Natoli, Gioacchino

    2006-01-01

    Dendritic cell (DC) maturation links peripheral events initiated by the encounter with pathogens to the activation and expansion of antigen-specific T lymphocytes in secondary lymphoid organs. Here, we describe an as yet unrecognized modulator of human DC maturation, the transcriptional repressor BCL6. We found that both myeloid and plasmacytoid DCs constitutively express BCL6, which is rapidly downregulated following maturation triggered by selected stimuli. Both in unstimulated and maturing DCs, control of BCL6 protein levels reflects the convergence of several mechanisms regulating BCL6 stability, mRNA transcription and nuclear export. By regulating the induction of several genes implicated in the immune response, including inflammatory cytokines, chemokines and survival genes, BCL6 may represent a pivotal modulator of the afferent branch of the immune response

  19. Transcriptionally Driven DNA Replication Program of the Human Parasite Leishmania major

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    Rodrigo Lombraña

    2016-08-01

    Full Text Available Faithful inheritance of eukaryotic genomes requires the orchestrated activation of multiple DNA replication origins (ORIs. Although origin firing is mechanistically conserved, how origins are specified and selected for activation varies across different model systems. Here, we provide a complete analysis of the nucleosomal landscape and replication program of the human parasite Leishmania major, building on a better evolutionary understanding of replication organization in Eukarya. We found that active transcription is a driving force for the nucleosomal organization of the L. major genome and that both the spatial and the temporal program of DNA replication can be explained as associated to RNA polymerase kinetics. This simple scenario likely provides flexibility and robustness to deal with the environmental changes that impose alterations in the genetic programs during parasitic life cycle stages. Our findings also suggest that coupling replication initiation to transcription elongation could be an ancient solution used by eukaryotic cells for origin maintenance.

  20. Acetylation-mediated suppression of transcription-independent memory: bidirectional modulation of memory by acetylation.

    Directory of Open Access Journals (Sweden)

    Katja Merschbaecher

    Full Text Available Learning induced changes in protein acetylation, mediated by histone acetyl transferases (HATs, and the antagonistic histone deacetylases (HDACs play a critical role in memory formation. The status of histone acetylation affects the interaction between the transcription-complex and DNA and thus regulates transcription-dependent processes required for long-term memory (LTM. While the majority of studies report on the role of elevated acetylation in memory facilitation, we address the impact of both, increased and decreased acetylation on formation of appetitive olfactory memory in honeybees. We show that learning-induced changes in the acetylation of histone H3 at aminoacid-positions H3K9 and H3K18 exhibit distinct and different dynamics depending on the training strength. A strong training that induces LTM leads to an immediate increase in acetylation at H3K18 that stays elevated for hours. A weak training, not sufficient to trigger LTM, causes an initial increase in acetylation at H3K18, followed by a strong reduction in acetylation at H3K18 below the control group level. Acetylation at position H3K9 is not affected by associative conditioning, indicating specific learning-induced actions on the acetylation machinery. Elevating acetylation levels by blocking HDACs after conditioning leads to an improved memory. While memory after strong training is enhanced for at least 2 days, the enhancement after weak training is restricted to 1 day. Reducing acetylation levels by blocking HAT activity after strong training leads to a suppression of transcription-dependent LTM. The memory suppression is also observed in case of weak training, which does not require transcription processes. Thus, our findings demonstrate that acetylation-mediated processes act as bidirectional regulators of memory formation that facilitate or suppress memory independent of its transcription-requirement.