Genes uniquely expressed in human growth plate chondrocytes uncover a distinct regulatory network.

PubMed

Li, Bing; Balasubramanian, Karthika; Krakow, Deborah; Cohn, Daniel H

2017-12-20

Chondrogenesis is the earliest stage of skeletal development and is a highly dynamic process, integrating the activities and functions of transcription factors, cell signaling molecules and extracellular matrix proteins. The molecular mechanisms underlying chondrogenesis have been extensively studied and multiple key regulators of this process have been identified. However, a genome-wide overview of the gene regulatory network in chondrogenesis has not been achieved. In this study, employing RNA sequencing, we identified 332 protein coding genes and 34 long non-coding RNA (lncRNA) genes that are highly selectively expressed in human fetal growth plate chondrocytes. Among the protein coding genes, 32 genes were associated with 62 distinct human skeletal disorders and 153 genes were associated with skeletal defects in knockout mice, confirming their essential roles in skeletal formation. These gene products formed a comprehensive physical interaction network and participated in multiple cellular processes regulating skeletal development. The data also revealed 34 transcription factors and 11,334 distal enhancers that were uniquely active in chondrocytes, functioning as transcriptional regulators for the cartilage-selective genes. Our findings revealed a complex gene regulatory network controlling skeletal development whereby transcription factors, enhancers and lncRNAs participate in chondrogenesis by transcriptional regulation of key genes. Additionally, the cartilage-selective genes represent candidate genes for unsolved human skeletal disorders.

ChIPBase: a database for decoding the transcriptional regulation of long non-coding RNA and microRNA genes from ChIP-Seq data.

PubMed

Yang, Jian-Hua; Li, Jun-Hao; Jiang, Shan; Zhou, Hui; Qu, Liang-Hu

2013-01-01

Long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) represent two classes of important non-coding RNAs in eukaryotes. Although these non-coding RNAs have been implicated in organismal development and in various human diseases, surprisingly little is known about their transcriptional regulation. Recent advances in chromatin immunoprecipitation with next-generation DNA sequencing (ChIP-Seq) have provided methods of detecting transcription factor binding sites (TFBSs) with unprecedented sensitivity. In this study, we describe ChIPBase (http://deepbase.sysu.edu.cn/chipbase/), a novel database that we have developed to facilitate the comprehensive annotation and discovery of transcription factor binding maps and transcriptional regulatory relationships of lncRNAs and miRNAs from ChIP-Seq data. The current release of ChIPBase includes high-throughput sequencing data that were generated by 543 ChIP-Seq experiments in diverse tissues and cell lines from six organisms. By analysing millions of TFBSs, we identified tens of thousands of TF-lncRNA and TF-miRNA regulatory relationships. Furthermore, two web-based servers were developed to annotate and discover transcriptional regulatory relationships of lncRNAs and miRNAs from ChIP-Seq data. In addition, we developed two genome browsers, deepView and genomeView, to provide integrated views of multidimensional data. Moreover, our web implementation supports diverse query types and the exploration of TFs, lncRNAs, miRNAs, gene ontologies and pathways.

The PPAR{gamma} coding region and its role in visceral obesity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Boon Yin, Khoo; Institute for Research in Molecular Medicine; Najimudin, Nazalan

Peroxisome proliferator-activated receptor gamma (PPAR{gamma}) is a ligand activated transcription factor, plays many essential roles of biological function in higher organisms. The PPAR{gamma} is mainly expressed in adipose tissue. It regulates the transcriptional activity of genes by binding with other transcription factor. The PPAR{gamma} coding region has been found to be closest to that of monkey in ours and other research groups. Thus, monkey is a more suitable animal model for future PPAR{gamma} studying, although mice and rat are frequently being used. The PPAR{gamma} is involved in regulating alterations of adipose tissue masses result from changes in mature adipocyte sizemore » and/or number through a complex interplay process called adipogenesis. However, the role of PPAR{gamma} in negatively regulating the process of adipogenesis remains unclear. This review may help we investigate the differential expression of key transcription factor in adipose tissue in response to visceral obesity-induced diet in vivo. The study may also provide valuable information to define a more appropriate physiological condition in adipogenesis which may help to prevent diseases cause by negative regulation of the transcription factors in adipose tissue.« less

Regulatory coding of lymphoid lineage choice by hematopoietic transcription factors

NASA Technical Reports Server (NTRS)

Warren, Luigi A.; Rothenberg, Ellen V.

2003-01-01

During lymphopoiesis, precursor cells negotiate a complex regulatory space, defined by the levels of several competing and cross-regulating transcription factors, before arriving at stable states of commitment to the B-, T- and NK-specific developmental programs. Recent perturbation experiments provide evidence that this space has three major axes, corresponding to the PU.1 versus GATA-1 balance, the intensity of Notch signaling through the CSL pathway, and the ratio of E-box transcription factors to their Id protein antagonists.

Control of transcription of the Bacillus subtilis spoIIIG gene, which codes for the forespore-specific transcription factor sigma G.

PubMed

Sun, D X; Cabrera-Martinez, R M; Setlow, P

1991-05-01

The Bacillus subtilis spoIIIG gene codes for a sigma factor termed sigma G which directs transcription of genes expressed only in the forespore compartment of the sporulating cell. Use of spoIIIG-lacZ transcriptional fusions showed that spoIIIG is cotranscribed with the spoIIG operon beginning at t0.5-1 of sporulation. However, this large mRNA produced little if any sigma G, and transferring the spoIIIG gene without the spoIIG promoter into the amyE locus resulted in a Spo+ phenotype. Significant translation of spoIIIG began at t2.5-3 with use of an mRNA whose 5' end is just upstream of the spoIIIG coding sequence. Synthesis of this spoIIIG-specific mRNA was not abolished by a deletion in spoIIIG itself. Similar results were obtained when a spoIIIG-lacZ translational fusion lacking the spoIIG promoter was integrated at the amyE locus. These data suggest that synthesis of sigma G is dependent neither on transcription from the spoIIG promoter nor on sigma G itself but can be due to another transcription factor. This transcription factor may be sigma F, the product of the spoIIAC locus, since a spoIIAC mutation blocked spoIIIG expression, and sequences upstream of the 5' end of the spoIIIG-specific mRNA agree well with the recognition sequence for sigma F. RNA polymerase containing sigma F (E sigma F) initiated transcription in vitro on a spoIIIG template at the 5' end found in vivo, as did E sigma G. However, E sigma F showed a greater than 20-fold preference for spoIIIG over a known sigma G-dependent gene compared with the activity of E sigma G.

Dynamic expression of transcription factor Brn3b during mouse cranial nerve development

PubMed Central

Sajgo, Szilard; Ali, Seid; Popescu, Octavian; Badea, Tudor Constantin

2015-01-01

During development transcription factor combinatorial codes define a large variety of morphologically and physiologically distinct neurons. Such a combinatorial code has been proposed for the differentiation of projection neurons of the somatic and visceral components of cranial nerves. It is possible that individual neuronal cell types are not specified by unique transcription factors, but rather emerge through the intersection of their expression domains. Brn3a, Brn3b and Brn3c, in combination with each other and/or transcription factors of other families, can define subgroups of Retinal Ganglion Cells (RGC), Spiral and Vestibular Ganglia, inner ear and vestibular hair cell neurons in the vestibuloacoustic system, and groups of somatosensory neurons in the Dorsal Root Ganglia (DRG). In the present study we investigated the expression and potential role of the Brn3b transcription factor in cranial nerves and associated nuclei of the brainstem. We report the dynamic expression of Brn3b in the somatosensory component of cranial nerves II, V, VII and VIII and visceromotor nuclei of nerves VII, IX, X, as well as other brainstem nuclei during different stages of development into adult stage. We find that genetically identified Brn3bKO RGC axons show correct but delayed pathfinding during the early stages of embryonic development. However loss of Brn3b does not affect the anatomy of the other cranial nerves normally expressing this transcription factor. PMID:26356988

Differential Regulation of Brain-Derived Neurotrophic Factor Transcripts during the Consolidation of Fear Learning

ERIC Educational Resources Information Center

Ressler, Kerry J.; Rattiner, Lisa M.; Davis, Michael

2004-01-01

Brain-derived neurotrophic factor (BDNF) has been implicated as a molecular mediator of learning and memory. The BDNF gene contains four differentially regulated promoters that generate four distinct mRNA transcripts, each containing a unique noncoding 5[prime]-exon and a common 3[prime]-coding exon. This study describes novel evidence for the…

Identification of novel non-coding RNA-based negative feedback regulating the expression of the oncogenic transcription factor GLI1.

PubMed

Villegas, Victoria E; Rahman, Mohammed Ferdous-Ur; Fernandez-Barrena, Maite G; Diao, Yumei; Liapi, Eleni; Sonkoly, Enikö; Ståhle, Mona; Pivarcsi, Andor; Annaratone, Laura; Sapino, Anna; Ramírez Clavijo, Sandra; Bürglin, Thomas R; Shimokawa, Takashi; Ramachandran, Saraswathi; Kapranov, Philipp; Fernandez-Zapico, Martin E; Zaphiropoulos, Peter G

2014-07-01

Non-coding RNAs are a complex class of nucleic acids, with growing evidence supporting regulatory roles in gene expression. Here we identify a non-coding RNA located head-to-head with the gene encoding the Glioma-associated oncogene 1 (GLI1), a transcriptional effector of multiple cancer-associated signaling pathways. The expression of this three-exon GLI1 antisense (GLI1AS) RNA in cancer cells was concordant with GLI1 levels. siRNAs knockdown of GLI1AS up-regulated GLI1 and increased cellular proliferation and tumor growth in a xenograft model system. Conversely, GLI1AS overexpression decreased the levels of GLI1, its target genes PTCH1 and PTCH2, and cellular proliferation. Additionally, we demonstrate that GLI1 knockdown reduced GLI1AS, while GLI1 overexpression increased GLI1AS, supporting the role of GLI1AS as a target gene of the GLI1 transcription factor. Activation of TGFβ and Hedgehog signaling, two known regulators of GLI1 expression, conferred a concordant up-regulation of GLI1 and GLI1AS in cancer cells. Finally, analysis of the mechanism underlying the interplay between GLI1 and GLI1AS indicates that the non-coding RNA elicits a local alteration of chromatin structure by increasing the silencing mark H3K27me3 and decreasing the recruitment of RNA polymerase II to this locus. Taken together, the data demonstrate the existence of a novel non-coding RNA-based negative feedback loop controlling GLI1 levels, thus expanding the repertoire of mechanisms regulating the expression of this oncogenic transcription factor. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Predicting phonetic transcription agreement: Insights from research in infant vocalizations

PubMed Central

RAMSDELL, HEATHER L.; OLLER, D. KIMBROUGH; ETHINGTON, CORINNA A.

2010-01-01

The purpose of this study is to provide new perspectives on correlates of phonetic transcription agreement. Our research focuses on phonetic transcription and coding of infant vocalizations. The findings are presumed to be broadly applicable to other difficult cases of transcription, such as found in severe disorders of speech, which similarly result in low reliability for a variety of reasons. We evaluated the predictiveness of two factors not previously documented in the literature as influencing transcription agreement: canonicity and coder confidence. Transcribers coded samples of infant vocalizations, judging both canonicity and confidence. Correlation results showed that canonicity and confidence were strongly related to agreement levels, and regression results showed that canonicity and confidence both contributed significantly to explanation of variance. Specifically, the results suggest that canonicity plays a major role in transcription agreement when utterances involve supraglottal articulation, with coder confidence offering additional power in predicting transcription agreement. PMID:17882695

Plant Mediator complex and its critical functions in transcription regulation.

PubMed

Yang, Yan; Li, Ling; Qu, Li-Jia

2016-02-01

The Mediator complex is an important component of the eukaryotic transcriptional machinery. As an essential link between transcription factors and RNA polymerase II, the Mediator complex transduces diverse signals to genes involved in different pathways. The plant Mediator complex was recently purified and comprises conserved and specific subunits. It functions in concert with transcription factors to modulate various responses. In this review, we summarize the recent advances in understanding the plant Mediator complex and its diverse roles in plant growth, development, defense, non-coding RNA production, response to abiotic stresses, flowering, genomic stability and metabolic homeostasis. In addition, the transcription factors interacting with the Mediator complex are also highlighted. © 2015 Institute of Botany, Chinese Academy of Sciences.

[Regulation of heat shock gene expression in response to stress].

PubMed

Garbuz, D G

2017-01-01

Heat shock (HS) genes, or stress genes, code for a number of proteins that collectively form the most ancient and universal stress defense system. The system determines the cell capability of adaptation to various adverse factors and performs a variety of auxiliary functions in normal physiological conditions. Common stress factors, such as higher temperatures, hypoxia, heavy metals, and others, suppress transcription and translation for the majority of genes, while HS genes are upregulated. Transcription of HS genes is controlled by transcription factors of the HS factor (HSF) family. Certain HSFs are activated on exposure to higher temperatures or other adverse factors to ensure stress-induced HS gene expression, while other HSFs are specifically activated at particular developmental stages. The regulation of the main mammalian stress-inducible factor HSF1 and Drosophila melanogaster HSF includes many components, such as a variety of early warning signals indicative of abnormal cell activity (e.g., increases in intracellular ceramide, cytosolic calcium ions, or partly denatured proteins); protein kinases, which phosphorylate HSFs at various Ser residues; acetyltransferases; and regulatory proteins, such as SUMO and HSBP1. Transcription factors other than HSFs are also involved in activating HS gene transcription; the set includes D. melanogaster GAF, mammalian Sp1 and NF-Y, and other factors. Transcription of several stress genes coding for molecular chaperones of the glucose-regulated protein (GRP) family is predominantly regulated by another stress-detecting system, which is known as the unfolded protein response (UPR) system and is activated in response to massive protein misfolding in the endoplasmic reticulum and mitochondrial matrix. A translational fine tuning of HS protein expression occurs via changing the phosphorylation status of several proteins involved in translation initiation. In addition, specific signal sequences in the 5'-UTRs of some HS protein mRNAs ensure their preferential translation in stress.

Expression of metastasis suppressor gene AES driven by a Yin Yang (YY) element in a CpG island promoter and transcription factor YY2.

PubMed

Kakizaki, Fumihiko; Sonoshita, Masahiro; Miyoshi, Hiroyuki; Itatani, Yoshiro; Ito, Shinji; Kawada, Kenji; Sakai, Yoshiharu; Taketo, M Mark

2016-11-01

We recently found that the product of the AES gene functions as a metastasis suppressor of colorectal cancer (CRC) in both humans and mice. Expression of amino-terminal enhancer of split (AES) protein is significantly decreased in liver metastatic lesions compared with primary colon tumors. To investigate its downregulation mechanism in metastases, we searched for transcriptional regulators of AES in human CRC and found that its expression is reduced mainly by transcriptional dysregulation and, in some cases, by additional haploidization of its coding gene. The AES promoter-enhancer is in a typical CpG island, and contains a Yin-Yang transcription factor recognition sequence (YY element). In human epithelial cells of normal colon and primary tumors, transcription factor YY2, a member of the YY family, binds directly to the YY element, and stimulates expression of AES. In a transplantation mouse model of liver metastases, however, expression of Yy2 (and therefore of Aes) is downregulated. In human CRC metastases to the liver, the levels of AES protein are correlated with those of YY2. In addition, we noticed copy-number reduction for the AES coding gene in chromosome 19p13.3 in 12% (5/42) of human CRC cell lines. We excluded other mechanisms such as point or indel mutations in the coding or regulatory regions of the AES gene, CpG methylation in the AES promoter enhancer, expression of microRNAs, and chromatin histone modifications. These results indicate that Aes may belong to a novel family of metastasis suppressors with a CpG-island promoter enhancer, and it is regulated transcriptionally. © 2016 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

Developmental roles of 21 Drosophila transcription factors are determined by quantitative differences in binding to an overlapping set of thousands of genomic regions

DOE Office of Scientific and Technical Information (OSTI.GOV)

MacArthur, Stewart; Li, Xiao-Yong; Li, Jingyi

2009-05-15

BACKGROUND: We previously established that six sequence-specific transcription factors that initiate anterior/posterior patterning in Drosophila bind to overlapping sets of thousands of genomic regions in blastoderm embryos. While regions bound at high levels include known and probable functional targets, more poorly bound regions are preferentially associated with housekeeping genes and/or genes not transcribed in the blastoderm, and are frequently found in protein coding sequences or in less conserved non-coding DNA, suggesting that many are likely non-functional. RESULTS: Here we show that an additional 15 transcription factors that regulate other aspects of embryo patterning show a similar quantitative continuum of functionmore » and binding to thousands of genomic regions in vivo. Collectively, the 21 regulators show a surprisingly high overlap in the regions they bind given that they belong to 11 DNA binding domain families, specify distinct developmental fates, and can act via different cis-regulatory modules. We demonstrate, however, that quantitative differences in relative levels of binding to shared targets correlate with the known biological and transcriptional regulatory specificities of these factors. CONCLUSIONS: It is likely that the overlap in binding of biochemically and functionally unrelated transcription factors arises from the high concentrations of these proteins in nuclei, which, coupled with their broad DNA binding specificities, directs them to regions of open chromatin. We suggest that most animal transcription factors will be found to show a similar broad overlapping pattern of binding in vivo, with specificity achieved by modulating the amount, rather than the identity, of bound factor.« less

Transcriptional role of androgen receptor in the expression of long non-coding RNA Sox2OT in neurogenesis

PubMed Central

Tosetti, Valentina; Sassone, Jenny; Ferri, Anna L. M.; Taiana, Michela; Bedini, Gloria; Nava, Sara; Brenna, Greta; Di Resta, Chiara; Pareyson, Davide; Di Giulio, Anna Maria; Carelli, Stephana

2017-01-01

The complex architecture of adult brain derives from tightly regulated migration and differentiation of precursor cells generated during embryonic neurogenesis. Changes at transcriptional level of genes that regulate migration and differentiation may lead to neurodevelopmental disorders. Androgen receptor (AR) is a transcription factor that is already expressed during early embryonic days. However, AR role in the regulation of gene expression at early embryonic stage is yet to be determinate. Long non-coding RNA (lncRNA) Sox2 overlapping transcript (Sox2OT) plays a crucial role in gene expression control during development but its transcriptional regulation is still to be clearly defined. Here, using Bicalutamide in order to pharmacologically inactivated AR, we investigated whether AR participates in the regulation of the transcription of the lncRNASox2OTat early embryonic stage. We identified a new DNA binding region upstream of Sox2 locus containing three androgen response elements (ARE), and found that AR binds such a sequence in embryonic neural stem cells and in mouse embryonic brain. Our data suggest that through this binding, AR can promote the RNA polymerase II dependent transcription of Sox2OT. Our findings also suggest that AR participates in embryonic neurogenesis through transcriptional control of the long non-coding RNA Sox2OT. PMID:28704421

Transcriptional role of androgen receptor in the expression of long non-coding RNA Sox2OT in neurogenesis.

PubMed

Tosetti, Valentina; Sassone, Jenny; Ferri, Anna L M; Taiana, Michela; Bedini, Gloria; Nava, Sara; Brenna, Greta; Di Resta, Chiara; Pareyson, Davide; Di Giulio, Anna Maria; Carelli, Stephana; Parati, Eugenio A; Gorio, Alfredo

2017-01-01

The complex architecture of adult brain derives from tightly regulated migration and differentiation of precursor cells generated during embryonic neurogenesis. Changes at transcriptional level of genes that regulate migration and differentiation may lead to neurodevelopmental disorders. Androgen receptor (AR) is a transcription factor that is already expressed during early embryonic days. However, AR role in the regulation of gene expression at early embryonic stage is yet to be determinate. Long non-coding RNA (lncRNA) Sox2 overlapping transcript (Sox2OT) plays a crucial role in gene expression control during development but its transcriptional regulation is still to be clearly defined. Here, using Bicalutamide in order to pharmacologically inactivated AR, we investigated whether AR participates in the regulation of the transcription of the lncRNASox2OTat early embryonic stage. We identified a new DNA binding region upstream of Sox2 locus containing three androgen response elements (ARE), and found that AR binds such a sequence in embryonic neural stem cells and in mouse embryonic brain. Our data suggest that through this binding, AR can promote the RNA polymerase II dependent transcription of Sox2OT. Our findings also suggest that AR participates in embryonic neurogenesis through transcriptional control of the long non-coding RNA Sox2OT.

Coding and non-coding gene regulatory networks underlie the immune response in liver cirrhosis.

PubMed

Gao, Bo; Zhang, Xueming; Huang, Yongming; Yang, Zhengpeng; Zhang, Yuguo; Zhang, Weihui; Gao, Zu-Hua; Xue, Dongbo

2017-01-01

Liver cirrhosis is recognized as being the consequence of immune-mediated hepatocyte damage and repair processes. However, the regulation of these immune responses underlying liver cirrhosis has not been elucidated. In this study, we used GEO datasets and bioinformatics methods to established coding and non-coding gene regulatory networks including transcription factor-/lncRNA-microRNA-mRNA, and competing endogenous RNA interaction networks. Our results identified 2224 mRNAs, 70 lncRNAs and 46 microRNAs were differentially expressed in liver cirrhosis. The transcription factor -/lncRNA- microRNA-mRNA network we uncovered that results in immune-mediated liver cirrhosis is comprised of 5 core microRNAs (e.g., miR-203; miR-219-5p), 3 transcription factors (i.e., FOXP3, ETS1 and FOS) and 7 lncRNAs (e.g., ENTS00000671336, ENST00000575137). The competing endogenous RNA interaction network we identified includes a complex immune response regulatory subnetwork that controls the entire liver cirrhosis network. Additionally, we found 10 overlapping GO terms shared by both liver cirrhosis and hepatocellular carcinoma including "immune response" as well. Interestingly, the overlapping differentially expressed genes in liver cirrhosis and hepatocellular carcinoma were enriched in immune response-related functional terms. In summary, a complex gene regulatory network underlying immune response processes may play an important role in the development and progression of liver cirrhosis, and its development into hepatocellular carcinoma.

Quantifying the Effect of DNA Packaging on Gene Expression Level

NASA Astrophysics Data System (ADS)

Kim, Harold

2010-10-01

Gene expression, the process by which the genetic code comes alive in the form of proteins, is one of the most important biological processes in living cells, and begins when transcription factors bind to specific DNA sequences in the promoter region upstream of a gene. The relationship between gene expression output and transcription factor input which is termed the gene regulation function is specific to each promoter, and predicting this gene regulation function from the locations of transcription factor binding sites is one of the challenges in biology. In eukaryotic organisms (for example, animals, plants, fungi etc), DNA is highly compacted into nucleosomes, 147-bp segments of DNA tightly wrapped around histone protein core, and therefore, the accessibility of transcription factor binding sites depends on their locations with respect to nucleosomes - sites inside nucleosomes are less accessible than those outside nucleosomes. To understand how transcription factor binding sites contribute to gene expression in a quantitative manner, we obtain gene regulation functions of promoters with various configurations of transcription factor binding sites by using fluorescent protein reporters to measure transcription factor input and gene expression output in single yeast cells. In this talk, I will show that the affinity of a transcription factor binding site inside and outside the nucleosome controls different aspects of the gene regulation function, and explain this finding based on a mass-action kinetic model that includes competition between nucleosomes and transcription factors.

Chromatin-Specific Regulation of Mammalian rDNA Transcription by Clustered TTF-I Binding Sites

PubMed Central

Diermeier, Sarah D.; Németh, Attila; Rehli, Michael; Grummt, Ingrid; Längst, Gernot

2013-01-01

Enhancers and promoters often contain multiple binding sites for the same transcription factor, suggesting that homotypic clustering of binding sites may serve a role in transcription regulation. Here we show that clustering of binding sites for the transcription termination factor TTF-I downstream of the pre-rRNA coding region specifies transcription termination, increases the efficiency of transcription initiation and affects the three-dimensional structure of rRNA genes. On chromatin templates, but not on free rDNA, clustered binding sites promote cooperative binding of TTF-I, loading TTF-I to the downstream terminators before it binds to the rDNA promoter. Interaction of TTF-I with target sites upstream and downstream of the rDNA transcription unit connects these distal DNA elements by forming a chromatin loop between the rDNA promoter and the terminators. The results imply that clustered binding sites increase the binding affinity of transcription factors in chromatin, thus influencing the timing and strength of DNA-dependent processes. PMID:24068958

A novel TBP-TAF complex on RNA polymerase II-transcribed snRNA genes.

PubMed

Zaborowska, Justyna; Taylor, Alice; Roeder, Robert G; Murphy, Shona

2012-01-01

Initiation of transcription of most human genes transcribed by RNA polymerase II (RNAP II) requires the formation of a preinitiation complex comprising TFIIA, B, D, E, F, H and RNAP II. The general transcription factor TFIID is composed of the TATA-binding protein and up to 13 TBP-associated factors. During transcription of snRNA genes, RNAP II does not appear to make the transition to long-range productive elongation, as happens during transcription of protein-coding genes. In addition, recognition of the snRNA gene-type specific 3' box RNA processing element requires initiation from an snRNA gene promoter. These characteristics may, at least in part, be driven by factors recruited to the promoter. For example, differences in the complement of TAFs might result in differential recruitment of elongation and RNA processing factors. As precedent, it already has been shown that the promoters of some protein-coding genes do not recruit all the TAFs found in TFIID. Although TAF5 has been shown to be associated with RNAP II-transcribed snRNA genes, the full complement of TAFs associated with these genes has remained unclear. Here we show, using a ChIP and siRNA-mediated approach, that the TBP/TAF complex on snRNA genes differs from that found on protein-coding genes. Interestingly, the largest TAF, TAF1, and the core TAFs, TAF10 and TAF4, are not detected on snRNA genes. We propose that this snRNA gene-specific TAF subset plays a key role in gene type-specific control of expression.

Activity-Dependent Human Brain Coding/Noncoding Gene Regulatory Networks

PubMed Central

Lipovich, Leonard; Dachet, Fabien; Cai, Juan; Bagla, Shruti; Balan, Karina; Jia, Hui; Loeb, Jeffrey A.

2012-01-01

While most gene transcription yields RNA transcripts that code for proteins, a sizable proportion of the genome generates RNA transcripts that do not code for proteins, but may have important regulatory functions. The brain-derived neurotrophic factor (BDNF) gene, a key regulator of neuronal activity, is overlapped by a primate-specific, antisense long noncoding RNA (lncRNA) called BDNFOS. We demonstrate reciprocal patterns of BDNF and BDNFOS transcription in highly active regions of human neocortex removed as a treatment for intractable seizures. A genome-wide analysis of activity-dependent coding and noncoding human transcription using a custom lncRNA microarray identified 1288 differentially expressed lncRNAs, of which 26 had expression profiles that matched activity-dependent coding genes and an additional 8 were adjacent to or overlapping with differentially expressed protein-coding genes. The functions of most of these protein-coding partner genes, such as ARC, include long-term potentiation, synaptic activity, and memory. The nuclear lncRNAs NEAT1, MALAT1, and RPPH1, composing an RNAse P-dependent lncRNA-maturation pathway, were also upregulated. As a means to replicate human neuronal activity, repeated depolarization of SY5Y cells resulted in sustained CREB activation and produced an inverse pattern of BDNF-BDNFOS co-expression that was not achieved with a single depolarization. RNAi-mediated knockdown of BDNFOS in human SY5Y cells increased BDNF expression, suggesting that BDNFOS directly downregulates BDNF. Temporal expression patterns of other lncRNA-messenger RNA pairs validated the effect of chronic neuronal activity on the transcriptome and implied various lncRNA regulatory mechanisms. lncRNAs, some of which are unique to primates, thus appear to have potentially important regulatory roles in activity-dependent human brain plasticity. PMID:22960213

Coding and non-coding gene regulatory networks underlie the immune response in liver cirrhosis

PubMed Central

Zhang, Xueming; Huang, Yongming; Yang, Zhengpeng; Zhang, Yuguo; Zhang, Weihui; Gao, Zu-hua; Xue, Dongbo

2017-01-01

Liver cirrhosis is recognized as being the consequence of immune-mediated hepatocyte damage and repair processes. However, the regulation of these immune responses underlying liver cirrhosis has not been elucidated. In this study, we used GEO datasets and bioinformatics methods to established coding and non-coding gene regulatory networks including transcription factor-/lncRNA-microRNA-mRNA, and competing endogenous RNA interaction networks. Our results identified 2224 mRNAs, 70 lncRNAs and 46 microRNAs were differentially expressed in liver cirrhosis. The transcription factor -/lncRNA- microRNA-mRNA network we uncovered that results in immune-mediated liver cirrhosis is comprised of 5 core microRNAs (e.g., miR-203; miR-219-5p), 3 transcription factors (i.e., FOXP3, ETS1 and FOS) and 7 lncRNAs (e.g., ENTS00000671336, ENST00000575137). The competing endogenous RNA interaction network we identified includes a complex immune response regulatory subnetwork that controls the entire liver cirrhosis network. Additionally, we found 10 overlapping GO terms shared by both liver cirrhosis and hepatocellular carcinoma including “immune response” as well. Interestingly, the overlapping differentially expressed genes in liver cirrhosis and hepatocellular carcinoma were enriched in immune response-related functional terms. In summary, a complex gene regulatory network underlying immune response processes may play an important role in the development and progression of liver cirrhosis, and its development into hepatocellular carcinoma. PMID:28355233

P‐TEFb goes viral

PubMed Central

Zaborowska, Justyna; Isa, Nur F.

2015-01-01

Positive transcription elongation factor b (P‐TEFb), which comprises cyclin‐dependent kinase 9 (CDK9) kinase and cyclin T subunits, is an essential kinase complex in human cells. Phosphorylation of the negative elongation factors by P‐TEFb is required for productive elongation of transcription of protein‐coding genes by RNA polymerase II (pol II). In addition, P‐TEFb‐mediated phosphorylation of the carboxyl‐terminal domain (CTD) of the largest subunit of pol II mediates the recruitment of transcription and RNA processing factors during the transcription cycle. CDK9 also phosphorylates p53, a tumor suppressor that plays a central role in cellular responses to a range of stress factors. Many viral factors affect transcription by recruiting or modulating the activity of CDK9. In this review, we will focus on how the function of CDK9 is regulated by viral gene products. The central role of CDK9 in viral life cycles suggests that drugs targeting the interaction between viral products and P‐TEFb could be effective anti‐viral agents. PMID:27398404

The histone modifications governing TFF1 transcription mediated by estrogen receptor.

PubMed

Li, Yanyan; Sun, Luyang; Zhang, Yu; Wang, Dandan; Wang, Feng; Liang, Jing; Gui, Bin; Shang, Yongfeng

2011-04-22

Transcription regulation by histone modifications is a major contributing factor to the structural and functional diversity in biology. These modifications are encrypted as histone codes or histone languages and function to establish and maintain heritable epigenetic codes that define the identity and the fate of the cell. Despite recent advances revealing numerous histone modifications associated with transcription regulation, how such modifications dictate the process of transcription is not fully understood. Here we describe spatial and temporal analyses of the histone modifications that are introduced during estrogen receptor α (ERα)-activated transcription. We demonstrated that aborting RNA polymerase II caused a disruption of the histone modifications that are associated with transcription elongation but had a minimal effect on modifications deposited during transcription initiation. We also found that the histone H3S10 phosphorylation mark is catalyzed by mitogen- and stress-activated protein kinase 1 (MSK1) and is recognized by a 14-3-3ζ/14-3-3ε heterodimer through its interaction with H3K4 trimethyltransferase SMYD3 and the p52 subunit of TFIIH. We showed that H3S10 phosphorylation is a prerequisite for H3K4 trimethylation. In addition, we demonstrated that SET8/PR-Set7/KMT5A is required for ERα-regulated transcription and its catalyzed H4K20 monomethylation is implicated in both transcription initiation and elongation. Our experiments provide a relatively comprehensive analysis of histone modifications associated with ERα-regulated transcription and define the biological meaning of several key components of the histone code that governs ERα-regulated transcription.

ChIPBase v2.0: decoding transcriptional regulatory networks of non-coding RNAs and protein-coding genes from ChIP-seq data.

PubMed

Zhou, Ke-Ren; Liu, Shun; Sun, Wen-Ju; Zheng, Ling-Ling; Zhou, Hui; Yang, Jian-Hua; Qu, Liang-Hu

2017-01-04

The abnormal transcriptional regulation of non-coding RNAs (ncRNAs) and protein-coding genes (PCGs) is contributed to various biological processes and linked with human diseases, but the underlying mechanisms remain elusive. In this study, we developed ChIPBase v2.0 (http://rna.sysu.edu.cn/chipbase/) to explore the transcriptional regulatory networks of ncRNAs and PCGs. ChIPBase v2.0 has been expanded with ∼10 200 curated ChIP-seq datasets, which represent about 20 times expansion when comparing to the previous released version. We identified thousands of binding motif matrices and their binding sites from ChIP-seq data of DNA-binding proteins and predicted millions of transcriptional regulatory relationships between transcription factors (TFs) and genes. We constructed 'Regulator' module to predict hundreds of TFs and histone modifications that were involved in or affected transcription of ncRNAs and PCGs. Moreover, we built a web-based tool, Co-Expression, to explore the co-expression patterns between DNA-binding proteins and various types of genes by integrating the gene expression profiles of ∼10 000 tumor samples and ∼9100 normal tissues and cell lines. ChIPBase also provides a ChIP-Function tool and a genome browser to predict functions of diverse genes and visualize various ChIP-seq data. This study will greatly expand our understanding of the transcriptional regulations of ncRNAs and PCGs. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Identification of BSAP (Pax-5) target genes in early B-cell development by loss- and gain-of-function experiments.

PubMed Central

Nutt, S L; Morrison, A M; Dörfler, P; Rolink, A; Busslinger, M

1998-01-01

The Pax-5 gene codes for the transcription factor BSAP which is essential for the progression of adult B lymphopoiesis beyond an early progenitor (pre-BI) cell stage. Although several genes have been proposed to be regulated by BSAP, CD19 is to date the only target gene which has been genetically confirmed to depend on this transcription factor for its expression. We have now taken advantage of cultured pre-BI cells of wild-type and Pax-5 mutant bone marrow to screen a large panel of B lymphoid genes for additional BSAP target genes. Four differentially expressed genes were shown to be under the direct control of BSAP, as their expression was rapidly regulated in Pax-5-deficient pre-BI cells by a hormone-inducible BSAP-estrogen receptor fusion protein. The genes coding for the B-cell receptor component Ig-alpha (mb-1) and the transcription factors N-myc and LEF-1 are positively regulated by BSAP, while the gene coding for the cell surface protein PD-1 is efficiently repressed. Distinct regulatory mechanisms of BSAP were revealed by reconstituting Pax-5-deficient pre-BI cells with full-length BSAP or a truncated form containing only the paired domain. IL-7 signalling was able to efficiently induce the N-myc gene only in the presence of full-length BSAP, while complete restoration of CD19 synthesis was critically dependent on the BSAP protein concentration. In contrast, the expression of the mb-1 and LEF-1 genes was already reconstituted by the paired domain polypeptide lacking any transactivation function, suggesting that the DNA-binding domain of BSAP is sufficient to recruit other transcription factors to the regulatory regions of these two genes. In conclusion, these loss- and gain-of-function experiments demonstrate that BSAP regulates four newly identified target genes as a transcriptional activator, repressor or docking protein depending on the specific regulatory sequence context. PMID:9545244

Genome-wide identification of conserved intronic non-coding sequences using a Bayesian segmentation approach.

PubMed

Algama, Manjula; Tasker, Edward; Williams, Caitlin; Parslow, Adam C; Bryson-Richardson, Robert J; Keith, Jonathan M

2017-03-27

Computational identification of non-coding RNAs (ncRNAs) is a challenging problem. We describe a genome-wide analysis using Bayesian segmentation to identify intronic elements highly conserved between three evolutionarily distant vertebrate species: human, mouse and zebrafish. We investigate the extent to which these elements include ncRNAs (or conserved domains of ncRNAs) and regulatory sequences. We identified 655 deeply conserved intronic sequences in a genome-wide analysis. We also performed a pathway-focussed analysis on genes involved in muscle development, detecting 27 intronic elements, of which 22 were not detected in the genome-wide analysis. At least 87% of the genome-wide and 70% of the pathway-focussed elements have existing annotations indicative of conserved RNA secondary structure. The expression of 26 of the pathway-focused elements was examined using RT-PCR, providing confirmation that they include expressed ncRNAs. Consistent with previous studies, these elements are significantly over-represented in the introns of transcription factors. This study demonstrates a novel, highly effective, Bayesian approach to identifying conserved non-coding sequences. Our results complement previous findings that these sequences are enriched in transcription factors. However, in contrast to previous studies which suggest the majority of conserved sequences are regulatory factor binding sites, the majority of conserved sequences identified using our approach contain evidence of conserved RNA secondary structures, and our laboratory results suggest most are expressed. Functional roles at DNA and RNA levels are not mutually exclusive, and many of our elements possess evidence of both. Moreover, ncRNAs play roles in transcriptional and post-transcriptional regulation, and this may contribute to the over-representation of these elements in introns of transcription factors. We attribute the higher sensitivity of the pathway-focussed analysis compared to the genome-wide analysis to improved alignment quality, suggesting that enhanced genomic alignments may reveal many more conserved intronic sequences.

A Positive Regulatory Loop between a Wnt-Regulated Non-coding RNA and ASCL2 Controls Intestinal Stem Cell Fate.

PubMed

Giakountis, Antonis; Moulos, Panagiotis; Zarkou, Vasiliki; Oikonomou, Christina; Harokopos, Vaggelis; Hatzigeorgiou, Artemis G; Reczko, Martin; Hatzis, Pantelis

2016-06-21

The canonical Wnt pathway plays a central role in stem cell maintenance, differentiation, and proliferation in the intestinal epithelium. Constitutive, aberrant activity of the TCF4/β-catenin transcriptional complex is the primary transforming factor in colorectal cancer. We identify a nuclear long non-coding RNA, termed WiNTRLINC1, as a direct target of TCF4/β-catenin in colorectal cancer cells. WiNTRLINC1 positively regulates the expression of its genomic neighbor ASCL2, a transcription factor that controls intestinal stem cell fate. WiNTRLINC1 interacts with TCF4/β-catenin to mediate the juxtaposition of its promoter with the regulatory regions of ASCL2. ASCL2, in turn, regulates WiNTRLINC1 transcriptionally, closing a feedforward regulatory loop that controls stem cell-related gene expression. This regulatory circuitry is highly amplified in colorectal cancer and correlates with increased metastatic potential and decreased patient survival. Our results uncover the interplay between non-coding RNA-mediated regulation and Wnt signaling and point to the diagnostic and therapeutic potential of WiNTRLINC1. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

The Mediator complex and transcription regulation

PubMed Central

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

Differential expression of lncRNAs during the HIV replication cycle: an underestimated layer in the HIV-host interplay.

PubMed

Trypsteen, Wim; Mohammadi, Pejman; Van Hecke, Clarissa; Mestdagh, Pieter; Lefever, Steve; Saeys, Yvan; De Bleser, Pieter; Vandesompele, Jo; Ciuffi, Angela; Vandekerckhove, Linos; De Spiegelaere, Ward

2016-10-26

Studying the effects of HIV infection on the host transcriptome has typically focused on protein-coding genes. However, recent advances in the field of RNA sequencing revealed that long non-coding RNAs (lncRNAs) add an extensive additional layer to the cell's molecular network. Here, we performed transcriptome profiling throughout a primary HIV infection in vitro to investigate lncRNA expression at the different HIV replication cycle processes (reverse transcription, integration and particle production). Subsequently, guilt-by-association, transcription factor and co-expression analysis were performed to infer biological roles for the lncRNAs identified in the HIV-host interplay. Many lncRNAs were suggested to play a role in mechanisms relying on proteasomal and ubiquitination pathways, apoptosis, DNA damage responses and cell cycle regulation. Through transcription factor binding analysis, we found that lncRNAs display a distinct transcriptional regulation profile as compared to protein coding mRNAs, suggesting that mRNAs and lncRNAs are independently modulated. In addition, we identified five differentially expressed lncRNA-mRNA pairs with mRNA involvement in HIV pathogenesis with possible cis regulatory lncRNAs that control nearby mRNA expression and function. Altogether, the present study demonstrates that lncRNAs add a new dimension to the HIV-host interplay and should be further investigated as they may represent targets for controlling HIV replication.

G =  MAT: linking transcription factor expression and DNA binding data.

PubMed

Tretyakov, Konstantin; Laur, Sven; Vilo, Jaak

2011-01-31

Transcription factors are proteins that bind to motifs on the DNA and thus affect gene expression regulation. The qualitative description of the corresponding processes is therefore important for a better understanding of essential biological mechanisms. However, wet lab experiments targeted at the discovery of the regulatory interplay between transcription factors and binding sites are expensive. We propose a new, purely computational method for finding putative associations between transcription factors and motifs. This method is based on a linear model that combines sequence information with expression data. We present various methods for model parameter estimation and show, via experiments on simulated data, that these methods are reliable. Finally, we examine the performance of this model on biological data and conclude that it can indeed be used to discover meaningful associations. The developed software is available as a web tool and Scilab source code at http://biit.cs.ut.ee/gmat/.

G = MAT: Linking Transcription Factor Expression and DNA Binding Data

PubMed Central

Tretyakov, Konstantin; Laur, Sven; Vilo, Jaak

2011-01-01

Transcription factors are proteins that bind to motifs on the DNA and thus affect gene expression regulation. The qualitative description of the corresponding processes is therefore important for a better understanding of essential biological mechanisms. However, wet lab experiments targeted at the discovery of the regulatory interplay between transcription factors and binding sites are expensive. We propose a new, purely computational method for finding putative associations between transcription factors and motifs. This method is based on a linear model that combines sequence information with expression data. We present various methods for model parameter estimation and show, via experiments on simulated data, that these methods are reliable. Finally, we examine the performance of this model on biological data and conclude that it can indeed be used to discover meaningful associations. The developed software is available as a web tool and Scilab source code at http://biit.cs.ut.ee/gmat/. PMID:21297945

Pan-cancer transcriptomic analysis associates long non-coding RNAs with key mutational driver events

PubMed Central

Ashouri, Arghavan; Sayin, Volkan I.; Van den Eynden, Jimmy; Singh, Simranjit X.; Papagiannakopoulos, Thales; Larsson, Erik

2016-01-01

Thousands of long non-coding RNAs (lncRNAs) lie interspersed with coding genes across the genome, and a small subset has been implicated as downstream effectors in oncogenic pathways. Here we make use of transcriptome and exome sequencing data from thousands of tumours across 19 cancer types, to identify lncRNAs that are induced or repressed in relation to somatic mutations in key oncogenic driver genes. Our screen confirms known coding and non-coding effectors and also associates many new lncRNAs to relevant pathways. The associations are often highly reproducible across cancer types, and while many lncRNAs are co-expressed with their protein-coding hosts or neighbours, some are intergenic and independent. We highlight lncRNAs with possible functions downstream of the tumour suppressor TP53 and the master antioxidant transcription factor NFE2L2. Our study provides a comprehensive overview of lncRNA transcriptional alterations in relation to key driver mutational events in human cancers. PMID:28959951

Transcription Factor Binding Profiles Reveal Cyclic Expression of Human Protein-coding Genes and Non-coding RNAs

PubMed Central

Cheng, Chao; Ung, Matthew; Grant, Gavin D.; Whitfield, Michael L.

2013-01-01

Cell cycle is a complex and highly supervised process that must proceed with regulatory precision to achieve successful cellular division. Despite the wide application, microarray time course experiments have several limitations in identifying cell cycle genes. We thus propose a computational model to predict human cell cycle genes based on transcription factor (TF) binding and regulatory motif information in their promoters. We utilize ENCODE ChIP-seq data and motif information as predictors to discriminate cell cycle against non-cell cycle genes. Our results show that both the trans- TF features and the cis- motif features are predictive of cell cycle genes, and a combination of the two types of features can further improve prediction accuracy. We apply our model to a complete list of GENCODE promoters to predict novel cell cycle driving promoters for both protein-coding genes and non-coding RNAs such as lincRNAs. We find that a similar percentage of lincRNAs are cell cycle regulated as protein-coding genes, suggesting the importance of non-coding RNAs in cell cycle division. The model we propose here provides not only a practical tool for identifying novel cell cycle genes with high accuracy, but also new insights on cell cycle regulation by TFs and cis-regulatory elements. PMID:23874175

Repression of YdaS Toxin Is Mediated by Transcriptional Repressor RacR in the Cryptic rac Prophage of Escherichia coli K-12.

PubMed

Krishnamurthi, Revathy; Ghosh, Swagatha; Khedkar, Supriya; Seshasayee, Aswin Sai Narain

2017-01-01

Horizontal gene transfer is a major driving force behind the genomic diversity seen in prokaryotes. The cryptic rac prophage in Escherichia coli K-12 carries the gene for a putative transcription factor RacR, whose deletion is lethal. We have shown that the essentiality of racR in E. coli K-12 is attributed to its role in transcriptionally repressing toxin gene(s) called ydaS and ydaT , which are adjacent to and coded divergently to racR . IMPORTANCE Transcription factors in the bacterium E. coli are rarely essential, and when they are essential, they are largely toxin-antitoxin systems. While studying transcription factors encoded in horizontally acquired regions in E. coli , we realized that the protein RacR, a putative transcription factor encoded by a gene on the rac prophage, is an essential protein. Here, using genetics, biochemistry, and bioinformatics, we show that its essentiality derives from its role as a transcriptional repressor of the ydaS and ydaT genes, whose products are toxic to the cell. Unlike type II toxin-antitoxin systems in which transcriptional regulation involves complexes of the toxin and antitoxin, repression by RacR is sufficient to keep ydaS transcriptionally silent.

A novel statistical approach for identification of the master regulator transcription factor.

PubMed

Sikdar, Sinjini; Datta, Susmita

2017-02-02

Transcription factors are known to play key roles in carcinogenesis and therefore, are gaining popularity as potential therapeutic targets in drug development. A 'master regulator' transcription factor often appears to control most of the regulatory activities of the other transcription factors and the associated genes. This 'master regulator' transcription factor is at the top of the hierarchy of the transcriptomic regulation. Therefore, it is important to identify and target the master regulator transcription factor for proper understanding of the associated disease process and identifying the best therapeutic option. We present a novel two-step computational approach for identification of master regulator transcription factor in a genome. At the first step of our method we test whether there exists any master regulator transcription factor in the system. We evaluate the concordance of two ranked lists of transcription factors using a statistical measure. In case the concordance measure is statistically significant, we conclude that there is a master regulator. At the second step, our method identifies the master regulator transcription factor, if there exists one. In the simulation scenario, our method performs reasonably well in validating the existence of a master regulator when the number of subjects in each treatment group is reasonably large. In application to two real datasets, our method ensures the existence of master regulators and identifies biologically meaningful master regulators. An R code for implementing our method in a sample test data can be found in http://www.somnathdatta.org/software . We have developed a screening method of identifying the 'master regulator' transcription factor just using only the gene expression data. Understanding the regulatory structure and finding the master regulator help narrowing the search space for identifying biomarkers for complex diseases such as cancer. In addition to identifying the master regulator our method provides an overview of the regulatory structure of the transcription factors which control the global gene expression profiles and consequently the cell functioning.

TFIIS-Dependent Non-coding Transcription Regulates Developmental Genome Rearrangements

PubMed Central

Maliszewska-Olejniczak, Kamila; Gruchota, Julita; Gromadka, Robert; Denby Wilkes, Cyril; Arnaiz, Olivier; Mathy, Nathalie; Duharcourt, Sandra; Bétermier, Mireille; Nowak, Jacek K.

2015-01-01

Because of their nuclear dimorphism, ciliates provide a unique opportunity to study the role of non-coding RNAs (ncRNAs) in the communication between germline and somatic lineages. In these unicellular eukaryotes, a new somatic nucleus develops at each sexual cycle from a copy of the zygotic (germline) nucleus, while the old somatic nucleus degenerates. In the ciliate Paramecium tetraurelia, the genome is massively rearranged during this process through the reproducible elimination of repeated sequences and the precise excision of over 45,000 short, single-copy Internal Eliminated Sequences (IESs). Different types of ncRNAs resulting from genome-wide transcription were shown to be involved in the epigenetic regulation of genome rearrangements. To understand how ncRNAs are produced from the entire genome, we have focused on a homolog of the TFIIS elongation factor, which regulates RNA polymerase II transcriptional pausing. Six TFIIS-paralogs, representing four distinct families, can be found in P. tetraurelia genome. Using RNA interference, we showed that TFIIS4, which encodes a development-specific TFIIS protein, is essential for the formation of a functional somatic genome. Molecular analyses and high-throughput DNA sequencing upon TFIIS4 RNAi demonstrated that TFIIS4 is involved in all kinds of genome rearrangements, including excision of ~48% of IESs. Localization of a GFP-TFIIS4 fusion revealed that TFIIS4 appears specifically in the new somatic nucleus at an early developmental stage, before IES excision. RT-PCR experiments showed that TFIIS4 is necessary for the synthesis of IES-containing non-coding transcripts. We propose that these IES+ transcripts originate from the developing somatic nucleus and serve as pairing substrates for germline-specific short RNAs that target elimination of their homologous sequences. Our study, therefore, connects the onset of zygotic non coding transcription to the control of genome plasticity in Paramecium, and establishes for the first time a specific role of TFIIS in non-coding transcription in eukaryotes. PMID:26177014

Transcription Factors Bind Thousands of Active and InactiveRegions in the Drosophila Blastoderm

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Xiao-Yong; MacArthur, Stewart; Bourgon, Richard

2008-01-10

Identifying the genomic regions bound by sequence-specific regulatory factors is central both to deciphering the complex DNA cis-regulatory code that controls transcription in metazoans and to determining the range of genes that shape animal morphogenesis. Here, we use whole-genome tiling arrays to map sequences bound in Drosophila melanogaster embryos by the six maternal and gap transcription factors that initiate anterior-posterior patterning. We find that these sequence-specific DNA binding proteins bind with quantitatively different specificities to highly overlapping sets of several thousand genomic regions in blastoderm embryos. Specific high- and moderate-affinity in vitro recognition sequences for each factor are enriched inmore » bound regions. This enrichment, however, is not sufficient to explain the pattern of binding in vivo and varies in a context-dependent manner, demonstrating that higher-order rules must govern targeting of transcription factors. The more highly bound regions include all of the over forty well-characterized enhancers known to respond to these factors as well as several hundred putative new cis-regulatory modules clustered near developmental regulators and other genes with patterned expression at this stage of embryogenesis. The new targets include most of the microRNAs (miRNAs) transcribed in the blastoderm, as well as all major zygotically transcribed dorsal-ventral patterning genes, whose expression we show to be quantitatively modulated by anterior-posterior factors. In addition to these highly bound regions, there are several thousand regions that are reproducibly bound at lower levels. However, these poorly bound regions are, collectively, far more distant from genes transcribed in the blastoderm than highly bound regions; are preferentially found in protein-coding sequences; and are less conserved than highly bound regions. Together these observations suggest that many of these poorly-bound regions are not involved in early-embryonic transcriptional regulation, and a significant proportion may be nonfunctional. Surprisingly, for five of the six factors, their recognition sites are not unambiguously more constrained evolutionarily than the immediate flanking DNA, even in more highly bound and presumably functional regions, indicating that comparative DNA sequence analysis is limited in its ability to identify functional transcription factor targets.« less

Circular RNAs are long-lived and display only minimal early alterations in response to a growth factor

PubMed Central

Enuka, Yehoshua; Lauriola, Mattia; Feldman, Morris E.; Sas-Chen, Aldema; Ulitsky, Igor; Yarden, Yosef

2016-01-01

Circular RNAs (circRNAs) are widespread circles of non-coding RNAs with largely unknown function. Because stimulation of mammary cells with the epidermal growth factor (EGF) leads to dynamic changes in the abundance of coding and non-coding RNA molecules, and culminates in the acquisition of a robust migratory phenotype, this cellular model might disclose functions of circRNAs. Here we show that circRNAs of EGF-stimulated mammary cells are stably expressed, while mRNAs and microRNAs change within minutes. In general, the circRNAs we detected are relatively long-lived and weakly expressed. Interestingly, they are almost ubiquitously co-expressed with the corresponding linear transcripts, and the respective, shared promoter regions are more active compared to genes producing linear isoforms with no detectable circRNAs. These findings imply that altered abundance of circRNAs, unlike changes in the levels of other RNAs, might not play critical roles in signaling cascades and downstream transcriptional networks that rapidly commit cells to specific outcomes. PMID:26657629

Relationships between Translation and Transcription Processes during fMRI Connectivity Scanning and Coded Translation and Transcription in Writing Products after Scanning in Children with and without Transcription Disabilities

PubMed Central

Wallis, Peter; Richards, Todd; Boord, Peter; Abbott, Robert; Berninger, Virginia

2018-01-01

Students with transcription disabilities (dysgraphia/impaired handwriting, n = 13 or dyslexia/impaired word spelling, n = 16) or without transcription disabilities (controls) completed transcription and translation (idea generating, planning, and creating) writing tasks during fMRI connectivity scanning and compositions after scanning, which were coded for transcription and translation variables. Compositions in both groups showed diversity in genre beyond usual narrative-expository distinction; groups differed in coded transcription but not translation variables. For the control group specific transcription or translation tasks during scanning correlated with corresponding coded transcription or translation skills in composition, but connectivity during scanning was not correlated with coded handwriting during composing in dysgraphia group and connectivity during translating was not correlated with any coded variable during composing in dyslexia group. Results are discussed in reference to the trend in neuroscience to use connectivity from relevant seed points while performing tasks and trends in education to recognize the generativity (creativity) of composing at both the genre and syntax levels. PMID:29600113

Novel insights into the response of Atlantic salmon (Salmo salar) to Piscirickettsia salmonis: Interplay of coding genes and lncRNAs during bacterial infection.

PubMed

Valenzuela-Miranda, Diego; Gallardo-Escárate, Cristian

2016-12-01

Despite the high prevalence and impact to Chilean salmon aquaculture of the intracellular bacterium Piscirickettsia salmonis, the molecular underpinnings of host-pathogen interactions remain unclear. Herein, the interplay of coding and non-coding transcripts has been proposed as a key mechanism involved in immune response. Therefore, the aim of this study was to evidence how coding and non-coding transcripts are modulated during the infection process of Atlantic salmon with P. salmonis. For this, RNA-seq was conducted in brain, spleen, and head kidney samples, revealing different transcriptional profiles according to bacterial load. Additionally, while most of the regulated genes annotated for diverse biological processes during infection, a common response associated with clathrin-mediated endocytosis and iron homeostasis was present in all tissues. Interestingly, while endocytosis-promoting factors and clathrin inductions were upregulated, endocytic receptors were mainly downregulated. Furthermore, the regulation of genes related to iron homeostasis suggested an intracellular accumulation of iron, a process in which heme biosynthesis/degradation pathways might play an important role. Regarding the non-coding response, 918 putative long non-coding RNAs were identified, where 425 were newly characterized for S. salar. Finally, co-localization and co-expression analyses revealed a strong correlation between the modulations of long non-coding RNAs and genes associated with endocytosis and iron homeostasis. These results represent the first comprehensive study of putative interplaying mechanisms of coding and non-coding RNAs during bacterial infection in salmonids. Copyright © 2016 Elsevier Ltd. All rights reserved.

BRD4 assists elongation of both coding and enhancer RNAs guided by histone acetylation

PubMed Central

Kanno, Tomohiko; Kanno, Yuka; LeRoy, Gary; Campos, Eric; Sun, Hong-Wei; Brooks, Stephen R; Vahedi, Golnaz; Heightman, Tom D; Garcia, Benjamin A; Reinberg, Danny; Siebenlist, Ulrich; O’Shea, John J; Ozato, Keiko

2016-01-01

Small-molecule BET inhibitors interfere with the epigenetic interactions between acetylated histones and the bromodomains of the BET family proteins, including BRD4, and they potently inhibit growth of malignant cells by targeting cancer-promoting genes. BRD4 interacts with the pause-release factor P-TEFb, and has been proposed to release Pol II from promoter-proximal pausing. We show that BRD4 occupied widespread genomic regions in mouse cells, and directly stimulated elongation of both protein-coding transcripts and non-coding enhancer RNAs (eRNAs), dependent on the function of bromodomains. BRD4 interacted physically with elongating Pol II complexes, and assisted Pol II progression through hyper-acetylated nucleosomes by interacting with acetylated histones via bromodomains. On active enhancers, the BET inhibitor JQ1 antagonized BRD4-associated eRNA synthesis. Thus, BRD4 is involved in multiple steps of the transcription hierarchy, primarily by assisting transcript elongation both at enhancers and on gene bodies. PMID:25383670

A combinatorial code for pattern formation in Drosophila oogenesis.

PubMed

Yakoby, Nir; Bristow, Christopher A; Gong, Danielle; Schafer, Xenia; Lembong, Jessica; Zartman, Jeremiah J; Halfon, Marc S; Schüpbach, Trudi; Shvartsman, Stanislav Y

2008-11-01

Two-dimensional patterning of the follicular epithelium in Drosophila oogenesis is required for the formation of three-dimensional eggshell structures. Our analysis of a large number of published gene expression patterns in the follicle cells suggests that they follow a simple combinatorial code based on six spatial building blocks and the operations of union, difference, intersection, and addition. The building blocks are related to the distribution of inductive signals, provided by the highly conserved epidermal growth factor receptor and bone morphogenetic protein signaling pathways. We demonstrate the validity of the code by testing it against a set of patterns obtained in a large-scale transcriptional profiling experiment. Using the proposed code, we distinguish 36 distinct patterns for 81 genes expressed in the follicular epithelium and characterize their joint dynamics over four stages of oogenesis. The proposed combinatorial framework allows systematic analysis of the diversity and dynamics of two-dimensional transcriptional patterns and guides future studies of gene regulation.

The Nrf2-antioxidant response element pathway: a target for regulating energy metabolism

USDA-ARS?s Scientific Manuscript database

The nuclear factor E2-related factor 2 (Nrf2) is a transcription factor that responds to oxidative stress by binding to the antioxidant response element (ARE) in the promoter of genes coding for antioxidant enzymes like NAD(P)H:quinone oxidoreductase 1 (NQO1) and proteins for glutathione synthesis. ...

TEAD and YAP regulate the enhancer network of human embryonic pancreatic progenitors.

PubMed

Cebola, Inês; Rodríguez-Seguí, Santiago A; Cho, Candy H-H; Bessa, José; Rovira, Meritxell; Luengo, Mario; Chhatriwala, Mariya; Berry, Andrew; Ponsa-Cobas, Joan; Maestro, Miguel Angel; Jennings, Rachel E; Pasquali, Lorenzo; Morán, Ignasi; Castro, Natalia; Hanley, Neil A; Gomez-Skarmeta, Jose Luis; Vallier, Ludovic; Ferrer, Jorge

2015-05-01

The genomic regulatory programmes that underlie human organogenesis are poorly understood. Pancreas development, in particular, has pivotal implications for pancreatic regeneration, cancer and diabetes. We have now characterized the regulatory landscape of embryonic multipotent progenitor cells that give rise to all pancreatic epithelial lineages. Using human embryonic pancreas and embryonic-stem-cell-derived progenitors we identify stage-specific transcripts and associated enhancers, many of which are co-occupied by transcription factors that are essential for pancreas development. We further show that TEAD1, a Hippo signalling effector, is an integral component of the transcription factor combinatorial code of pancreatic progenitor enhancers. TEAD and its coactivator YAP activate key pancreatic signalling mediators and transcription factors, and regulate the expansion of pancreatic progenitors. This work therefore uncovers a central role for TEAD and YAP as signal-responsive regulators of multipotent pancreatic progenitors, and provides a resource for the study of embryonic development of the human pancreas.

Divergent transcription is associated with promoters of transcriptional regulators

PubMed Central

2013-01-01

Background Divergent transcription is a wide-spread phenomenon in mammals. For instance, short bidirectional transcripts are a hallmark of active promoters, while longer transcripts can be detected antisense from active genes in conditions where the RNA degradation machinery is inhibited. Moreover, many described long non-coding RNAs (lncRNAs) are transcribed antisense from coding gene promoters. However, the general significance of divergent lncRNA/mRNA gene pair transcription is still poorly understood. Here, we used strand-specific RNA-seq with high sequencing depth to thoroughly identify antisense transcripts from coding gene promoters in primary mouse tissues. Results We found that a substantial fraction of coding-gene promoters sustain divergent transcription of long non-coding RNA (lncRNA)/mRNA gene pairs. Strikingly, upstream antisense transcription is significantly associated with genes related to transcriptional regulation and development. Their promoters share several characteristics with those of transcriptional developmental genes, including very large CpG islands, high degree of conservation and epigenetic regulation in ES cells. In-depth analysis revealed a unique GC skew profile at these promoter regions, while the associated coding genes were found to have large first exons, two genomic features that might enforce bidirectional transcription. Finally, genes associated with antisense transcription harbor specific H3K79me2 epigenetic marking and RNA polymerase II enrichment profiles linked to an intensified rate of early transcriptional elongation. Conclusions We concluded that promoters of a class of transcription regulators are characterized by a specialized transcriptional control mechanism, which is directly coupled to relaxed bidirectional transcription. PMID:24365181

Cell cycle, oncogenic and tumor suppressor pathways regulate numerous long and macro non-protein-coding RNAs

PubMed Central

2014-01-01

Background The genome is pervasively transcribed but most transcripts do not code for proteins, constituting non-protein-coding RNAs. Despite increasing numbers of functional reports of individual long non-coding RNAs (lncRNAs), assessing the extent of functionality among the non-coding transcriptional output of mammalian cells remains intricate. In the protein-coding world, transcripts differentially expressed in the context of processes essential for the survival of multicellular organisms have been instrumental in the discovery of functionally relevant proteins and their deregulation is frequently associated with diseases. We therefore systematically identified lncRNAs expressed differentially in response to oncologically relevant processes and cell-cycle, p53 and STAT3 pathways, using tiling arrays. Results We found that up to 80% of the pathway-triggered transcriptional responses are non-coding. Among these we identified very large macroRNAs with pathway-specific expression patterns and demonstrated that these are likely continuous transcripts. MacroRNAs contain elements conserved in mammals and sauropsids, which in part exhibit conserved RNA secondary structure. Comparing evolutionary rates of a macroRNA to adjacent protein-coding genes suggests a local action of the transcript. Finally, in different grades of astrocytoma, a tumor disease unrelated to the initially used cell lines, macroRNAs are differentially expressed. Conclusions It has been shown previously that the majority of expressed non-ribosomal transcripts are non-coding. We now conclude that differential expression triggered by signaling pathways gives rise to a similar abundance of non-coding content. It is thus unlikely that the prevalence of non-coding transcripts in the cell is a trivial consequence of leaky or random transcription events. PMID:24594072

Function of the growth-regulated transcription initiation factor TIF-IA in initiation complex formation at the murine ribosomal gene promoter.

PubMed

Schnapp, A; Schnapp, G; Erny, B; Grummt, I

1993-11-01

Alterations in the rate of cell proliferation are accompanied by changes in the transcription of rRNA genes. In mammals, this growth-dependent regulation of transcription of genes coding for rRNA (rDNA) is due to reduction of the amount or activity of an essential transcription factor, called TIF-IA. Extracts prepared from quiescent cells lack this factor activity and, therefore, are transcriptionally inactive. We have purified TIF-IA from exponentially growing cells and have shown that it is a polypeptide with a molecular mass of 75 kDa which exists as a monomer in solution. Using a reconstituted transcription system consisting of purified transcription factors, we demonstrate that TIF-IA is a bona fide transcription initiation factor which interacts with RNA polymerase I. Preinitiation complexes can be assembled in the absence of TIF-IA, but formation of the first phosphodiester bonds of nascent rRNA is precluded. After initiation, TIF-IA is liberated from the initiation complex and facilitates transcription from templates bearing preinitiation complexes which lack TIF-IA. Despite the pronounced species specificity of class I gene transcription, this growth-dependent factor has been identified not only in mouse but also in human cells. Murine TIF-IA complements extracts from both growth-inhibited mouse and human cells. The analogous human activity appears to be similar or identical to that of TIF-IA. Therefore, despite the fact that the RNA polymerase transcription system has evolved sufficiently rapidly that an rDNA promoter from one species will not function in another species, the basic mechanisms that adapt ribosome synthesis to cell proliferation have been conserved.

Neuropsychological correlates of transcription factor AP-2Beta, and its interaction with COMT and MAOA in healthy females.

PubMed

Schabram, Ina; Eggermann, Thomas; Siegel, Steven J; Gründer, Gerhard; Zerres, Klaus; Vernaleken, Ingo

2013-01-01

The transcription factor AP-2β has been shown to impact clinical and neuropsychological properties. Apparently, it regulates the transcription of genes that code for molecules which are part of the catecholaminergic transmission system. This investigation focuses on possible effects of the transcription factor AP-2β intron 2 polymorphism on cognitive performance parameters. This hypothesis-driven investigation examined the effects and interactions of the transcription factor AP-2β intron 2 polymorphism, the Val158Met catechol-O-methyltransferase (COMT) polymorphism, and the variable number of tandem repeat polymorphism of monoamine oxidase A (MAOA) on cognitive performance parameters within a group of 200 healthy women (age: mean ± SD, 23.93 ± 3.33 years). The AP-2β polymorphism significantly influenced cognitive performance (in particular, the Trail Making Test part B), whereas the MAOA and COMT polymorphisms did not. However, there was an interaction effect of the AP-2β × MAOA × COMT genotypes on the decision bias β of the degraded-stimulus version of the continuous performance task. Only the Val158Met COMT polymorphism showed an influence on personality questionnaires (openness and self-transcendence; NEO Five-Factor Inventory, Temperament and Character Inventory). The transcription factor AP-2β intron 2 polymorphism had more influence on cognition than the MAOA and COMT polymorphisms. Possibly, the AP-2β genotype might influence cognition through pathways other than those that regulate MAOA and COMT transcription. Interactions of transcription factor AP-2β, COMT, and MAOA polymorphisms suggest higher leverage effects of transcription factor AP-2β in subjects with high dopamine availability. Copyright © 2013 S. Karger AG, Basel.

Natural variation in non-coding regions underlying phenotypic diversity in budding yeast

PubMed Central

Salinas, Francisco; de Boer, Carl G.; Abarca, Valentina; García, Verónica; Cuevas, Mara; Araos, Sebastian; Larrondo, Luis F.; Martínez, Claudio; Cubillos, Francisco A.

2016-01-01

Linkage mapping studies in model organisms have typically focused their efforts in polymorphisms within coding regions, ignoring those within regulatory regions that may contribute to gene expression variation. In this context, differences in transcript abundance are frequently proposed as a source of phenotypic diversity between individuals, however, until now, little molecular evidence has been provided. Here, we examined Allele Specific Expression (ASE) in six F1 hybrids from Saccharomyces cerevisiae derived from crosses between representative strains of the four main lineages described in yeast. ASE varied between crosses with levels ranging between 28% and 60%. Part of the variation in expression levels could be explained by differences in transcription factors binding to polymorphic cis-regulations and to differences in trans-activation depending on the allelic form of the TF. Analysis on highly expressed alleles on each background suggested ASN1 as a candidate transcript underlying nitrogen consumption differences between two strains. Further promoter allele swap analysis under fermentation conditions confirmed that coding and non-coding regions explained aspartic and glutamic acid consumption differences, likely due to a polymorphism affecting Uga3 binding. Together, we provide a new catalogue of variants to bridge the gap between genotype and phenotype. PMID:26898953

Antisense Transcription Is Pervasive but Rarely Conserved in Enteric Bacteria

PubMed Central

Raghavan, Rahul; Sloan, Daniel B.; Ochman, Howard

2012-01-01

ABSTRACT Noncoding RNAs, including antisense RNAs (asRNAs) that originate from the complementary strand of protein-coding genes, are involved in the regulation of gene expression in all domains of life. Recent application of deep-sequencing technologies has revealed that the transcription of asRNAs occurs genome-wide in bacteria. Although the role of the vast majority of asRNAs remains unknown, it is often assumed that their presence implies important regulatory functions, similar to those of other noncoding RNAs. Alternatively, many antisense transcripts may be produced by chance transcription events from promoter-like sequences that result from the degenerate nature of bacterial transcription factor binding sites. To investigate the biological relevance of antisense transcripts, we compared genome-wide patterns of asRNA expression in closely related enteric bacteria, Escherichia coli and Salmonella enterica serovar Typhimurium, by performing strand-specific transcriptome sequencing. Although antisense transcripts are abundant in both species, less than 3% of asRNAs are expressed at high levels in both species, and only about 14% appear to be conserved among species. And unlike the promoters of protein-coding genes, asRNA promoters show no evidence of sequence conservation between, or even within, species. Our findings suggest that many or even most bacterial asRNAs are nonadaptive by-products of the cell’s transcription machinery. PMID:22872780

A common class of transcripts with 5'-intron depletion, distinct early coding sequence features, and N1-methyladenosine modification.

PubMed

Cenik, Can; Chua, Hon Nian; Singh, Guramrit; Akef, Abdalla; Snyder, Michael P; Palazzo, Alexander F; Moore, Melissa J; Roth, Frederick P

2017-03-01

Introns are found in 5' untranslated regions (5'UTRs) for 35% of all human transcripts. These 5'UTR introns are not randomly distributed: Genes that encode secreted, membrane-bound and mitochondrial proteins are less likely to have them. Curiously, transcripts lacking 5'UTR introns tend to harbor specific RNA sequence elements in their early coding regions. To model and understand the connection between coding-region sequence and 5'UTR intron status, we developed a classifier that can predict 5'UTR intron status with >80% accuracy using only sequence features in the early coding region. Thus, the classifier identifies transcripts with 5 ' proximal- i ntron- m inus-like-coding regions ("5IM" transcripts). Unexpectedly, we found that the early coding sequence features defining 5IM transcripts are widespread, appearing in 21% of all human RefSeq transcripts. The 5IM class of transcripts is enriched for non-AUG start codons, more extensive secondary structure both preceding the start codon and near the 5' cap, greater dependence on eIF4E for translation, and association with ER-proximal ribosomes. 5IM transcripts are bound by the exon junction complex (EJC) at noncanonical 5' proximal positions. Finally, N 1 -methyladenosines are specifically enriched in the early coding regions of 5IM transcripts. Taken together, our analyses point to the existence of a distinct 5IM class comprising ∼20% of human transcripts. This class is defined by depletion of 5' proximal introns, presence of specific RNA sequence features associated with low translation efficiency, N 1 -methyladenosines in the early coding region, and enrichment for noncanonical binding by the EJC. © 2017 Cenik et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

Long Non-Coding RNAs Differentially Expressed between Normal versus Primary Breast Tumor Tissues Disclose Converse Changes to Breast Cancer-Related Protein-Coding Genes

PubMed Central

Reiche, Kristin; Kasack, Katharina; Schreiber, Stephan; Lüders, Torben; Due, Eldri U.; Naume, Bjørn; Riis, Margit; Kristensen, Vessela N.; Horn, Friedemann; Børresen-Dale, Anne-Lise; Hackermüller, Jörg; Baumbusch, Lars O.

2014-01-01

Breast cancer, the second leading cause of cancer death in women, is a highly heterogeneous disease, characterized by distinct genomic and transcriptomic profiles. Transcriptome analyses prevalently assessed protein-coding genes; however, the majority of the mammalian genome is expressed in numerous non-coding transcripts. Emerging evidence supports that many of these non-coding RNAs are specifically expressed during development, tumorigenesis, and metastasis. The focus of this study was to investigate the expression features and molecular characteristics of long non-coding RNAs (lncRNAs) in breast cancer. We investigated 26 breast tumor and 5 normal tissue samples utilizing a custom expression microarray enclosing probes for mRNAs as well as novel and previously identified lncRNAs. We identified more than 19,000 unique regions significantly differentially expressed between normal versus breast tumor tissue, half of these regions were non-coding without any evidence for functional open reading frames or sequence similarity to known proteins. The identified non-coding regions were primarily located in introns (53%) or in the intergenic space (33%), frequently orientated in antisense-direction of protein-coding genes (14%), and commonly distributed at promoter-, transcription factor binding-, or enhancer-sites. Analyzing the most diverse mRNA breast cancer subtypes Basal-like versus Luminal A and B resulted in 3,025 significantly differentially expressed unique loci, including 682 (23%) for non-coding transcripts. A notable number of differentially expressed protein-coding genes displayed non-synonymous expression changes compared to their nearest differentially expressed lncRNA, including an antisense lncRNA strongly anticorrelated to the mRNA coding for histone deacetylase 3 (HDAC3), which was investigated in more detail. Previously identified chromatin-associated lncRNAs (CARs) were predominantly downregulated in breast tumor samples, including CARs located in the protein-coding genes for CALD1, FTX, and HNRNPH1. In conclusion, a number of differentially expressed lncRNAs have been identified with relation to cancer-related protein-coding genes. PMID:25264628

Long non-coding RNAs differentially expressed between normal versus primary breast tumor tissues disclose converse changes to breast cancer-related protein-coding genes.

PubMed

Reiche, Kristin; Kasack, Katharina; Schreiber, Stephan; Lüders, Torben; Due, Eldri U; Naume, Bjørn; Riis, Margit; Kristensen, Vessela N; Horn, Friedemann; Børresen-Dale, Anne-Lise; Hackermüller, Jörg; Baumbusch, Lars O

2014-01-01

Breast cancer, the second leading cause of cancer death in women, is a highly heterogeneous disease, characterized by distinct genomic and transcriptomic profiles. Transcriptome analyses prevalently assessed protein-coding genes; however, the majority of the mammalian genome is expressed in numerous non-coding transcripts. Emerging evidence supports that many of these non-coding RNAs are specifically expressed during development, tumorigenesis, and metastasis. The focus of this study was to investigate the expression features and molecular characteristics of long non-coding RNAs (lncRNAs) in breast cancer. We investigated 26 breast tumor and 5 normal tissue samples utilizing a custom expression microarray enclosing probes for mRNAs as well as novel and previously identified lncRNAs. We identified more than 19,000 unique regions significantly differentially expressed between normal versus breast tumor tissue, half of these regions were non-coding without any evidence for functional open reading frames or sequence similarity to known proteins. The identified non-coding regions were primarily located in introns (53%) or in the intergenic space (33%), frequently orientated in antisense-direction of protein-coding genes (14%), and commonly distributed at promoter-, transcription factor binding-, or enhancer-sites. Analyzing the most diverse mRNA breast cancer subtypes Basal-like versus Luminal A and B resulted in 3,025 significantly differentially expressed unique loci, including 682 (23%) for non-coding transcripts. A notable number of differentially expressed protein-coding genes displayed non-synonymous expression changes compared to their nearest differentially expressed lncRNA, including an antisense lncRNA strongly anticorrelated to the mRNA coding for histone deacetylase 3 (HDAC3), which was investigated in more detail. Previously identified chromatin-associated lncRNAs (CARs) were predominantly downregulated in breast tumor samples, including CARs located in the protein-coding genes for CALD1, FTX, and HNRNPH1. In conclusion, a number of differentially expressed lncRNAs have been identified with relation to cancer-related protein-coding genes.

Control of light-dependent keto carotenoid biosynthesis in Nostoc 7120 by the transcription factor NtcA.

PubMed

Sandmann, Gerhard; Mautz, Jürgen; Breitenbach, Jürgen

2016-09-01

In Nostoc PCC 7120, two different ketolases, CrtW and CrtO are involved in the formation of keto carotenoids from β-carotene. In contrast to other cyanobacteria, CrtW catalyzes the formation of monoketo echinenone whereas CrtO is the only enzyme for the synthesis of diketo canthaxanthin. This is the major photo protective carotenoid in this cyanobacterium. Under high-light conditions, basic canthaxanthin formation was transcriptionally up-regulated. Upon transfer to high light, the transcript levels of all investigated carotenogenic genes including those coding for phytoene synthase, phytoene desaturase and both ketolases were increased. These transcription changes proceeded via binding of the transcription factor NtcA to the promoter regions of the carotenogenic genes. The binding was absolutely dependent on the presence of reductants and oxo-glutarate. Light-stimulated transcript formation was inhibited by DCMU. Therefore, photosynthetic electron transport is proposed as the sensor for high-light and a changing redox state as a signal for NtcA binding.

Tumor hypoxia induces nuclear paraspeckle formation through HIF-2α dependent transcriptional activation of NEAT1 leading to cancer cell survival

PubMed Central

Choudhry, H; Albukhari, A; Morotti, M; Haider, S; Moralli, D; Smythies, J; Schödel, J; Green, C M; Camps, C; Buffa, F; Ratcliffe, P; Ragoussis, J; Harris, A L; Mole, D R

2015-01-01

Activation of cellular transcriptional responses, mediated by hypoxia-inducible factor (HIF), is common in many types of cancer, and generally confers a poor prognosis. Known to induce many hundreds of protein-coding genes, HIF has also recently been shown to be a key regulator of the non-coding transcriptional response. Here, we show that NEAT1 long non-coding RNA (lncRNA) is a direct transcriptional target of HIF in many breast cancer cell lines and in solid tumors. Unlike previously described lncRNAs, NEAT1 is regulated principally by HIF-2 rather than by HIF-1. NEAT1 is a nuclear lncRNA that is an essential structural component of paraspeckles and the hypoxic induction of NEAT1 induces paraspeckle formation in a manner that is dependent upon both NEAT1 and on HIF-2. Paraspeckles are multifunction nuclear structures that sequester transcriptionally active proteins as well as RNA transcripts that have been subjected to adenosine-to-inosine (A-to-I) editing. We show that the nuclear retention of one such transcript, F11R (also known as junctional adhesion molecule 1, JAM1), in hypoxia is dependent upon the hypoxic increase in NEAT1, thereby conferring a novel mechanism of HIF-dependent gene regulation. Induction of NEAT1 in hypoxia also leads to accelerated cellular proliferation, improved clonogenic survival and reduced apoptosis, all of which are hallmarks of increased tumorigenesis. Furthermore, in patients with breast cancer, high tumor NEAT1 expression correlates with poor survival. Taken together, these results indicate a new role for HIF transcriptional pathways in the regulation of nuclear structure and that this contributes to the pro-tumorigenic hypoxia-phenotype in breast cancer. PMID:25417700

High-resolution transcriptional analysis of the regulatory influence of cell-to-cell signalling reveals novel genes that contribute to Xanthomonas phytopathogenesis

PubMed Central

An, Shi-Qi; Febrer, Melanie; McCarthy, Yvonne; Tang, Dong-Jie; Clissold, Leah; Kaithakottil, Gemy; Swarbreck, David; Tang, Ji-Liang; Rogers, Jane; Dow, J Maxwell; Ryan, Robert P

2013-01-01

The bacterium Xanthomonas campestris is an economically important pathogen of many crop species and a model for the study of bacterial phytopathogenesis. In X. campestris, a regulatory system mediated by the signal molecule DSF controls virulence to plants. The synthesis and recognition of the DSF signal depends upon different Rpf proteins. DSF signal generation requires RpfF whereas signal perception and transduction depends upon a system comprising the sensor RpfC and regulator RpfG. Here we have addressed the action and role of Rpf/DSF signalling in phytopathogenesis by high-resolution transcriptional analysis coupled to functional genomics. We detected transcripts for many genes that were unidentified by previous computational analysis of the genome sequence. Novel transcribed regions included intergenic transcripts predicted as coding or non-coding as well as those that were antisense to coding sequences. In total, mutation of rpfF, rpfG and rpfC led to alteration in transcript levels (more than fourfold) of approximately 480 genes. The regulatory influence of RpfF and RpfC demonstrated considerable overlap. Contrary to expectation, the regulatory influence of RpfC and RpfG had limited overlap, indicating complexities of the Rpf signalling system. Importantly, functional analysis revealed over 160 new virulence factors within the group of Rpf-regulated genes. PMID:23617851

Identification of a functionally distinct truncated BDNF mRNA splice variant and protein in Trachemys scripta elegans.

PubMed

Ambigapathy, Ganesh; Zheng, Zhaoqing; Li, Wei; Keifer, Joyce

2013-01-01

Brain-derived neurotrophic factor (BDNF) has a diverse functional role and complex pattern of gene expression. Alternative splicing of mRNA transcripts leads to further diversity of mRNAs and protein isoforms. Here, we describe the regulation of BDNF mRNA transcripts in an in vitro model of eyeblink classical conditioning and a unique transcript that forms a functionally distinct truncated BDNF protein isoform. Nine different mRNA transcripts from the BDNF gene of the pond turtle Trachemys scripta elegans (tBDNF) are selectively regulated during classical conditioning: exon I mRNA transcripts show no change, exon II transcripts are downregulated, while exon III transcripts are upregulated. One unique transcript that codes from exon II, tBDNF2a, contains a 40 base pair deletion in the protein coding exon that generates a truncated tBDNF protein. The truncated transcript and protein are expressed in the naïve untrained state and are fully repressed during conditioning when full-length mature tBDNF is expressed, thereby having an alternate pattern of expression in conditioning. Truncated BDNF is not restricted to turtles as a truncated mRNA splice variant has been described for the human BDNF gene. Further studies are required to determine the ubiquity of truncated BDNF alternative splice variants across species and the mechanisms of regulation and function of this newly recognized BDNF protein.

Identification of a Functionally Distinct Truncated BDNF mRNA Splice Variant and Protein in Trachemys scripta elegans

PubMed Central

Ambigapathy, Ganesh; Zheng, Zhaoqing; Li, Wei; Keifer, Joyce

2013-01-01

Brain-derived neurotrophic factor (BDNF) has a diverse functional role and complex pattern of gene expression. Alternative splicing of mRNA transcripts leads to further diversity of mRNAs and protein isoforms. Here, we describe the regulation of BDNF mRNA transcripts in an in vitro model of eyeblink classical conditioning and a unique transcript that forms a functionally distinct truncated BDNF protein isoform. Nine different mRNA transcripts from the BDNF gene of the pond turtle Trachemys scripta elegans (tBDNF) are selectively regulated during classical conditioning: exon I mRNA transcripts show no change, exon II transcripts are downregulated, while exon III transcripts are upregulated. One unique transcript that codes from exon II, tBDNF2a, contains a 40 base pair deletion in the protein coding exon that generates a truncated tBDNF protein. The truncated transcript and protein are expressed in the naïve untrained state and are fully repressed during conditioning when full-length mature tBDNF is expressed, thereby having an alternate pattern of expression in conditioning. Truncated BDNF is not restricted to turtles as a truncated mRNA splice variant has been described for the human BDNF gene. Further studies are required to determine the ubiquity of truncated BDNF alternative splice variants across species and the mechanisms of regulation and function of this newly recognized BDNF protein. PMID:23825634

Searching for transcription factor binding sites in vector spaces

PubMed Central

2012-01-01

Background Computational approaches to transcription factor binding site identification have been actively researched in the past decade. Learning from known binding sites, new binding sites of a transcription factor in unannotated sequences can be identified. A number of search methods have been introduced over the years. However, one can rarely find one single method that performs the best on all the transcription factors. Instead, to identify the best method for a particular transcription factor, one usually has to compare a handful of methods. Hence, it is highly desirable for a method to perform automatic optimization for individual transcription factors. Results We proposed to search for transcription factor binding sites in vector spaces. This framework allows us to identify the best method for each individual transcription factor. We further introduced two novel methods, the negative-to-positive vector (NPV) and optimal discriminating vector (ODV) methods, to construct query vectors to search for binding sites in vector spaces. Extensive cross-validation experiments showed that the proposed methods significantly outperformed the ungapped likelihood under positional background method, a state-of-the-art method, and the widely-used position-specific scoring matrix method. We further demonstrated that motif subtypes of a TF can be readily identified in this framework and two variants called the k NPV and k ODV methods benefited significantly from motif subtype identification. Finally, independent validation on ChIP-seq data showed that the ODV and NPV methods significantly outperformed the other compared methods. Conclusions We conclude that the proposed framework is highly flexible. It enables the two novel methods to automatically identify a TF-specific subspace to search for binding sites. Implementations are available as source code at: http://biogrid.engr.uconn.edu/tfbs_search/. PMID:23244338

Genetic Code Expansion as a Tool to Study Regulatory Processes of Transcription

NASA Astrophysics Data System (ADS)

Schmidt, Moritz; Summerer, Daniel

2014-02-01

The expansion of the genetic code with noncanonical amino acids (ncAA) enables the chemical and biophysical properties of proteins to be tailored, inside cells, with a previously unattainable level of precision. A wide range of ncAA with functions not found in canonical amino acids have been genetically encoded in recent years and have delivered insights into biological processes that would be difficult to access with traditional approaches of molecular biology. A major field for the development and application of novel ncAA-functions has been transcription and its regulation. This is particularly attractive, since advanced DNA sequencing- and proteomics-techniques continue to deliver vast information on these processes on a global level, but complementing methodologies to study them on a detailed, molecular level and in living cells have been comparably scarce. In a growing number of studies, genetic code expansion has now been applied to precisely control the chemical properties of transcription factors, RNA polymerases and histones, and this has enabled new insights into their interactions, conformational changes, cellular localizations and the functional roles of posttranslational modifications.

Differential protein-coding gene and long noncoding RNA expression in smoking-related lung squamous cell carcinoma.

PubMed

Li, Shicheng; Sun, Xiao; Miao, Shuncheng; Liu, Jia; Jiao, Wenjie

2017-11-01

Cigarette smoking is one of the greatest preventable risk factors for developing cancer, and most cases of lung squamous cell carcinoma (lung SCC) are associated with smoking. The pathogenesis mechanism of tumor progress is unclear. This study aimed to identify biomarkers in smoking-related lung cancer, including protein-coding gene, long noncoding RNA, and transcription factors. We selected and obtained messenger RNA microarray datasets and clinical data from the Gene Expression Omnibus database to identify gene expression altered by cigarette smoking. Integrated bioinformatic analysis was used to clarify biological functions of the identified genes, including Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, the construction of a protein-protein interaction network, transcription factor, and statistical analyses. Subsequent quantitative real-time PCR was utilized to verify these bioinformatic analyses. Five hundred and ninety-eight differentially expressed genes and 21 long noncoding RNA were identified in smoking-related lung SCC. GO and KEGG pathway analysis showed that identified genes were enriched in the cancer-related functions and pathways. The protein-protein interaction network revealed seven hub genes identified in lung SCC. Several transcription factors and their binding sites were predicted. The results of real-time quantitative PCR revealed that AURKA and BIRC5 were significantly upregulated and LINC00094 was downregulated in the tumor tissues of smoking patients. Further statistical analysis indicated that dysregulation of AURKA, BIRC5, and LINC00094 indicated poor prognosis in lung SCC. Protein-coding genes AURKA, BIRC5, and LINC00094 could be biomarkers or therapeutic targets for smoking-related lung SCC. © 2017 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.

Network perturbation by recurrent regulatory variants in cancer

PubMed Central

Cho, Ara; Lee, Insuk; Choi, Jung Kyoon

2017-01-01

Cancer driving genes have been identified as recurrently affected by variants that alter protein-coding sequences. However, a majority of cancer variants arise in noncoding regions, and some of them are thought to play a critical role through transcriptional perturbation. Here we identified putative transcriptional driver genes based on combinatorial variant recurrence in cis-regulatory regions. The identified genes showed high connectivity in the cancer type-specific transcription regulatory network, with high outdegree and many downstream genes, highlighting their causative role during tumorigenesis. In the protein interactome, the identified transcriptional drivers were not as highly connected as coding driver genes but appeared to form a network module centered on the coding drivers. The coding and regulatory variants associated via these interactions between the coding and transcriptional drivers showed exclusive and complementary occurrence patterns across tumor samples. Transcriptional cancer drivers may act through an extensive perturbation of the regulatory network and by altering protein network modules through interactions with coding driver genes. PMID:28333928

ICAM-1-related long non-coding RNA: promoter analysis and expression in human retinal endothelial cells.

PubMed

Lumsden, Amanda L; Ma, Yuefang; Ashander, Liam M; Stempel, Andrew J; Keating, Damien J; Smith, Justine R; Appukuttan, Binoy

2018-05-09

Regulation of intercellular adhesion molecule (ICAM)-1 in retinal endothelial cells is a promising druggable target for retinal vascular diseases. The ICAM-1-related (ICR) long non-coding RNA stabilizes ICAM-1 transcript, increasing protein expression. However, studies of ICR involvement in disease have been limited as the promoter is uncharacterized. To address this issue, we undertook a comprehensive in silico analysis of the human ICR gene promoter region. We used genomic evolutionary rate profiling to identify a 115 base pair (bp) sequence within 500 bp upstream of the transcription start site of the annotated human ICR gene that was conserved across 25 eutherian genomes. A second constrained sequence upstream of the orthologous mouse gene (68 bp; conserved across 27 Eutherian genomes including human) was also discovered. Searching these elements identified 33 matrices predictive of binding sites for transcription factors known to be responsive to a broad range of pathological stimuli, including hypoxia, and metabolic and inflammatory proteins. Five phenotype-associated single nucleotide polymorphisms (SNPs) in the immediate vicinity of these elements included four SNPs (i.e. rs2569693, rs281439, rs281440 and rs11575074) predicted to impact binding motifs of transcription factors, and thus the expression of ICR and ICAM-1 genes, with potential to influence disease susceptibility. We verified that human retinal endothelial cells expressed ICR, and observed induction of expression by tumor necrosis factor-α.

Auto-Regulatory RNA Editing Fine-Tunes mRNA Re-Coding and Complex Behaviour in Drosophila

PubMed Central

Savva, Yiannis A.; Jepson, James E.C; Sahin, Asli; Sugden, Arthur U.; Dorsky, Jacquelyn S.; Alpert, Lauren; Lawrence, Charles; Reenan, Robert A.

2014-01-01

Auto-regulatory feedback loops are a common molecular strategy used to optimize protein function. In Drosophila many mRNAs involved in neuro-transmission are re-coded at the RNA level by the RNA editing enzyme dADAR, leading to the incorporation of amino acids that are not directly encoded by the genome. dADAR also re-codes its own transcript, but the consequences of this auto-regulation in vivo are unclear. Here we show that hard-wiring or abolishing endogenous dADAR auto-regulation dramatically remodels the landscape of re-coding events in a site-specific manner. These molecular phenotypes correlate with altered localization of dADAR within the nuclear compartment. Furthermore, auto-editing exhibits sexually dimorphic patterns of spatial regulation and can be modified by abiotic environmental factors. Finally, we demonstrate that modifying dAdar auto-editing affects adaptive complex behaviors. Our results reveal the in vivo relevance of auto-regulatory control over post-transcriptional mRNA re-coding events in fine-tuning brain function and organismal behavior. PMID:22531175

Constitutive expression of a salinity-induced wheat WRKY transcription factor enhances salinity and ionic stress tolerance in transgenic Arabidopsis thaliana.

PubMed

Qin, Yuxiang; Tian, Yanchen; Han, Lu; Yang, Xinchao

2013-10-25

The isolation and characterization of TaWRKY79, a wheat class II WRKY transcription factor, is described. Its 1297 bp coding region includes a 987 bp long open reading frame. TaWRKY79 was induced by stressing seedlings with either NaCl or abscisic acid (ABA). When a fusion between an 843 bp segment upstream of the TaWRKY79 coding sequence and GUS was introduced into Arabidopsis thaliana, GUS staining indicated that this upstream segment captured the sequence(s) required to respond to ABA or NaCl treatment. When TaWRKY79 was constitutively expressed as a transgene in A. thaliana, the transgenic plants showed an improved capacity to extend their primary root in the presence of either 100 mM NaCl, 10 mM LiCl or 2 μM ABA. The inference was that TaWRKY79 enhanced the level of tolerance to both salinity and ionic stress, while reducing the level of sensitivity to ABA. The ABA-related genes ABA1, ABA2 ABI1 and ABI5 were all up-regulated in the TaWRKY79 transgenic plants, suggesting that the transcription factor operates in an ABA-dependent pathway. Copyright © 2013. Published by Elsevier Inc.

Modular Evolution of DNA-Binding Preference of a Tbrain Transcription Factor Provides a Mechanism for Modifying Gene Regulatory Networks

PubMed Central

Cheatle Jarvela, Alys M.; Brubaker, Lisa; Vedenko, Anastasia; Gupta, Anisha; Armitage, Bruce A.; Bulyk, Martha L.; Hinman, Veronica F.

2014-01-01

Gene regulatory networks (GRNs) describe the progression of transcriptional states that take a single-celled zygote to a multicellular organism. It is well documented that GRNs can evolve extensively through mutations to cis-regulatory modules (CRMs). Transcription factor proteins that bind these CRMs may also evolve to produce novelty. Coding changes are considered to be rarer, however, because transcription factors are multifunctional and hence are more constrained to evolve in ways that will not produce widespread detrimental effects. Recent technological advances have unearthed a surprising variation in DNA-binding abilities, such that individual transcription factors may recognize both a preferred primary motif and an additional secondary motif. This provides a source of modularity in function. Here, we demonstrate that orthologous transcription factors can also evolve a changed preference for a secondary binding motif, thereby offering an unexplored mechanism for GRN evolution. Using protein-binding microarray, surface plasmon resonance, and in vivo reporter assays, we demonstrate an important difference in DNA-binding preference between Tbrain protein orthologs in two species of echinoderms, the sea star, Patiria miniata, and the sea urchin, Strongylocentrotus purpuratus. Although both orthologs recognize the same primary motif, only the sea star Tbr also has a secondary binding motif. Our in vivo assays demonstrate that this difference may allow for greater evolutionary change in timing of regulatory control. This uncovers a layer of transcription factor binding divergence that could exist for many pairs of orthologs. We hypothesize that this divergence provides modularity that allows orthologous transcription factors to evolve novel roles in GRNs through modification of binding to secondary sites. PMID:25016582

Gene Expression Profiling of Transcription Factors of Helicobacter pylori under Different Environmental Conditions.

PubMed

De la Cruz, Miguel A; Ares, Miguel A; von Bargen, Kristine; Panunzi, Leonardo G; Martínez-Cruz, Jessica; Valdez-Salazar, Hilda A; Jiménez-Galicia, César; Torres, Javier

2017-01-01

Helicobacter pylori is a Gram-negative bacterium that colonizes the human gastric mucosa and causes peptic ulcers and gastric carcinoma. H. pylori strain 26695 has a small genome (1.67 Mb), which codes for few known transcriptional regulators that control bacterial metabolism and virulence. We analyzed by qRT-PCR the expression of 16 transcriptional regulators in H. pylori 26695, including the three sigma factors under different environmental conditions. When bacteria were exposed to acidic pH, urea, nickel, or iron, the sigma factors were differentially expressed with a particularly strong induction of fliA . The regulatory genes hrcA, hup , and crdR were highly induced in the presence of urea, nickel, and iron. In terms of biofilm formation fliA, flgR, hp1021, fur, nikR , and crdR were induced in sessile bacteria. Transcriptional expression levels of rpoD, flgR, hspR, hp1043 , and cheY were increased in contact with AGS epithelial cells. Kanamycin, chloramphenicol, and tetracycline increased or decreased expression of regulatory genes, showing that these antibiotics affect the transcription of H. pylori . Our data indicate that environmental cues which may be present in the human stomach modulate H. pylori transcription.

Different domains of the murine RNA polymerase I-specific termination factor mTTF-I serve distinct functions in transcription termination.

PubMed

Evers, R; Smid, A; Rudloff, U; Lottspeich, F; Grummt, I

1995-03-15

Termination of mouse ribosomal gene transcription by RNA polymerase I (Pol I) requires the specific interaction of a DNA binding protein, mTTF-I, with an 18 bp sequence element located downstream of the rRNA coding region. Here we describe the molecular cloning and functional characterization of the cDNA encoding this transcription termination factor. Recombinant mTTF-I binds specifically to the murine terminator elements and terminates Pol I transcription in a reconstituted in vitro system. Deletion analysis has defined a modular structure of mTTF-I comprising a dispensable N-terminal half, a large C-terminal DNA binding region and an internal domain which is required for transcription termination. Significantly, the C-terminal region of mTTF-I reveals striking homology to the DNA binding domains of the proto-oncogene c-Myb and the yeast transcription factor Reb1p. Site-directed mutagenesis of one of the tryptophan residues that is conserved in the homology region of c-Myb, Reb1p and mTTF-I abolishes specific DNA binding, a finding which underscores the functional relevance of these residues in DNA-protein interactions.

Different domains of the murine RNA polymerase I-specific termination factor mTTF-I serve distinct functions in transcription termination.

PubMed Central

Evers, R; Smid, A; Rudloff, U; Lottspeich, F; Grummt, I

1995-01-01

Termination of mouse ribosomal gene transcription by RNA polymerase I (Pol I) requires the specific interaction of a DNA binding protein, mTTF-I, with an 18 bp sequence element located downstream of the rRNA coding region. Here we describe the molecular cloning and functional characterization of the cDNA encoding this transcription termination factor. Recombinant mTTF-I binds specifically to the murine terminator elements and terminates Pol I transcription in a reconstituted in vitro system. Deletion analysis has defined a modular structure of mTTF-I comprising a dispensable N-terminal half, a large C-terminal DNA binding region and an internal domain which is required for transcription termination. Significantly, the C-terminal region of mTTF-I reveals striking homology to the DNA binding domains of the proto-oncogene c-Myb and the yeast transcription factor Reb1p. Site-directed mutagenesis of one of the tryptophan residues that is conserved in the homology region of c-Myb, Reb1p and mTTF-I abolishes specific DNA binding, a finding which underscores the functional relevance of these residues in DNA-protein interactions. Images PMID:7720715

A genome-wide survey of maternal and embryonic transcripts during Xenopus tropicalis development.

PubMed

Paranjpe, Sarita S; Jacobi, Ulrike G; van Heeringen, Simon J; Veenstra, Gert Jan C

2013-11-06

Dynamics of polyadenylation vs. deadenylation determine the fate of several developmentally regulated genes. Decay of a subset of maternal mRNAs and new transcription define the maternal-to-zygotic transition, but the full complement of polyadenylated and deadenylated coding and non-coding transcripts has not yet been assessed in Xenopus embryos. To analyze the dynamics and diversity of coding and non-coding transcripts during development, both polyadenylated mRNA and ribosomal RNA-depleted total RNA were harvested across six developmental stages and subjected to high throughput sequencing. The maternally loaded transcriptome is highly diverse and consists of both polyadenylated and deadenylated transcripts. Many maternal genes show peak expression in the oocyte and include genes which are known to be the key regulators of events like oocyte maturation and fertilization. Of all the transcripts that increase in abundance between early blastula and larval stages, about 30% of the embryonic genes are induced by fourfold or more by the late blastula stage and another 35% by late gastrulation. Using a gene model validation and discovery pipeline, we identified novel transcripts and putative long non-coding RNAs (lncRNA). These lncRNA transcripts were stringently selected as spliced transcripts generated from independent promoters, with limited coding potential and a codon bias characteristic of noncoding sequences. Many lncRNAs are conserved and expressed in a developmental stage-specific fashion. These data reveal dynamics of transcriptome polyadenylation and abundance and provides a high-confidence catalogue of novel and long non-coding RNAs.

Global insights into high temperature and drought stress regulated genes by RNA-Seq in economically important oilseed crop Brassica juncea.

PubMed

Bhardwaj, Ankur R; Joshi, Gopal; Kukreja, Bharti; Malik, Vidhi; Arora, Priyanka; Pandey, Ritu; Shukla, Rohit N; Bankar, Kiran G; Katiyar-Agarwal, Surekha; Goel, Shailendra; Jagannath, Arun; Kumar, Amar; Agarwal, Manu

2015-01-21

Brassica juncea var. Varuna is an economically important oilseed crop of family Brassicaceae which is vulnerable to abiotic stresses at specific stages in its life cycle. Till date no attempts have been made to elucidate genome-wide changes in its transcriptome against high temperature or drought stress. To gain global insights into genes, transcription factors and kinases regulated by these stresses and to explore information on coding transcripts that are associated with traits of agronomic importance, we utilized a combinatorial approach of next generation sequencing and de-novo assembly to discover B. juncea transcriptome associated with high temperature and drought stresses. We constructed and sequenced three transcriptome libraries namely Brassica control (BC), Brassica high temperature stress (BHS) and Brassica drought stress (BDS). More than 180 million purity filtered reads were generated which were processed through quality parameters and high quality reads were assembled de-novo using SOAPdenovo assembler. A total of 77750 unique transcripts were identified out of which 69,245 (89%) were annotated with high confidence. We established a subset of 19110 transcripts, which were differentially regulated by either high temperature and/or drought stress. Furthermore, 886 and 2834 transcripts that code for transcription factors and kinases, respectively, were also identified. Many of these were responsive to high temperature, drought or both stresses. Maximum number of up-regulated transcription factors in high temperature and drought stress belonged to heat shock factors (HSFs) and dehydration responsive element-binding (DREB) families, respectively. We also identified 239 metabolic pathways, which were perturbed during high temperature and drought treatments. Analysis of gene ontologies associated with differentially regulated genes forecasted their involvement in diverse biological processes. Our study provides first comprehensive discovery of B. juncea transcriptome under high temperature and drought stress conditions. Transcriptome resource generated in this study will enhance our understanding on the molecular mechanisms involved in defining the response of B. juncea against two important abiotic stresses. Furthermore this information would benefit designing of efficient crop improvement strategies for tolerance against conditions of high temperature regimes and water scarcity.

Trypanosome RNA polymerases and transcription factors: sensible trypanocidal drug targets?

PubMed

Vanhamme, Luc

2008-11-01

Trypanosomes and Leishmaniae are the agents of several important parasitic diseases threatening hundreds of million human beings worldwide. As they diverged early in evolution, they display original molecular characteristics. These peculiarities are each defining putative specific targets for anti-parasitic drugs. Transcription displays its lot of unique characteristics in trypanosomes and will be taken as an example to uncover these targets. Unique features of transcription in trypanosomes include constitutive and poly-cistronic transcription by RNA polymerase II as well as transcription of protein-coding genes by RNA polymerase I. It is becoming clear that these unique mechanisms are performed by dedicated molecular players. The first of them have been recently characterized. They are reviewed and their suitability as drug targets is commented.

Lipopolysaccharide-induced innate immune factors in the bottlenose dolphin (Tursiops truncatus) detected in expression sequence tag analysis.

PubMed

Ohishi, Kazue; Shishido, Reiko; Iwata, Yasunao; Saitoh, Masafumi; Takenaka, Ryota; Ohtsu, Dai; Okutsu, Kenji; Maruyama, Tadashi

2011-11-01

EST analysis based on the megaclone-megasorting method was performed using leukocytes from the bottlenose dolphin (Tursiops truncatus) with or without LPS stimulation. A total of 849 upregulated and 384 downregulated EST clones were sequenced, annotated, and functionally classified. Ferritin heavy peptide I was the most abundant upregulated transcript, suggesting that LPS stimulation induced high production of reactive oxygen species, which were sequestered in ferritin. Among the immune factors, the transcripts coding for an IL-1Ra, homologs to bovine serum amyloid A3, and canine intercellular adhesion molecule-1 were highly expressed. Markedly downregulated transcripts of immune factors were those for homologs of calcium-binding proteins belonging to the S100 family, S100A12, S100A8, and S100A6. Time-course experiments on the expression of some immune factors including IL-1Ra suggested that these factors interact and control cetacean innate immunity. © 2011 The Societies and Blackwell Publishing Asia Pty Ltd.

Problem-Based Test: The Effect of Fibroblast Growth Factor on Gene Expression

ERIC Educational Resources Information Center

Szeberenyi, Jozsef

2011-01-01

This paper shows the results of an experiment in which the effects of fibroblast growth factor (FGF), actinomycin D (Act D; an inhibitor of transcription), and cycloheximide (CHX; an inhibitor of translation) were studied on the expression of two genes: a gene called "Fnk" and the gene coding for glyceraldehyde-3-phosphate dehydrogenase (GAPDH).…

Comparative genomics and transcriptional profiles of Saccharopolyspora erythraea NRRL 2338 and a classically improved erythromycin over-producing strain

PubMed Central

2012-01-01

Background The molecular mechanisms altered by the traditional mutation and screening approach during the improvement of antibiotic-producing microorganisms are still poorly understood although this information is essential to design rational strategies for industrial strain improvement. In this study, we applied comparative genomics to identify all genetic changes occurring during the development of an erythromycin overproducer obtained using the traditional mutate-and- screen method. Results Compared with the parental Saccharopolyspora erythraea NRRL 2338, the genome of the overproducing strain presents 117 deletion, 78 insertion and 12 transposition sites, with 71 insertion/deletion sites mapping within coding sequences (CDSs) and generating frame-shift mutations. Single nucleotide variations are present in 144 CDSs. Overall, the genomic variations affect 227 proteins of the overproducing strain and a considerable number of mutations alter genes of key enzymes in the central carbon and nitrogen metabolism and in the biosynthesis of secondary metabolites, resulting in the redirection of common precursors toward erythromycin biosynthesis. Interestingly, several mutations inactivate genes coding for proteins that play fundamental roles in basic transcription and translation machineries including the transcription anti-termination factor NusB and the transcription elongation factor Efp. These mutations, along with those affecting genes coding for pleiotropic or pathway-specific regulators, affect global expression profile as demonstrated by a comparative analysis of the parental and overproducer expression profiles. Genomic data, finally, suggest that the mutate-and-screen process might have been accelerated by mutations in DNA repair genes. Conclusions This study helps to clarify the mechanisms underlying antibiotic overproduction providing valuable information about new possible molecular targets for rationale strain improvement. PMID:22401291

Multiple Transcript Properties Related to Translation Affect mRNA Degradation Rates in Saccharomyces cerevisiae

PubMed Central

Neymotin, Benjamin; Ettorre, Victoria; Gresham, David

2016-01-01

Degradation of mRNA contributes to variation in transcript abundance. Studies of individual mRNAs have shown that both cis and trans factors affect mRNA degradation rates. However, the factors underlying transcriptome-wide variation in mRNA degradation rates are poorly understood. We investigated the contribution of different transcript properties to transcriptome-wide degradation rate variation in the budding yeast, Saccharomyces cerevisiae, using multiple regression analysis. We find that multiple transcript properties are significantly associated with variation in mRNA degradation rates, and that a model incorporating these properties explains ∼50% of the genome-wide variance. Predictors of mRNA degradation rates include transcript length, ribosome density, biased codon usage, and GC content of the third position in codons. To experimentally validate these factors, we studied individual transcripts expressed from identical promoters. We find that decreasing ribosome density by mutating the first translational start site of a transcript increases its degradation rate. Using coding sequence variants of green fluorescent protein (GFP) that differ only at synonymous sites, we show that increased GC content of the third position of codons results in decreased rates of mRNA degradation. Thus, in steady-state conditions, a large fraction of genome-wide variation in mRNA degradation rates is determined by inherent properties of transcripts, many of which are related to translation, rather than specific regulatory mechanisms. PMID:27633789

Bijective transformation circular codes and nucleotide exchanging RNA transcription.

PubMed

Michel, Christian J; Seligmann, Hervé

2014-04-01

The C(3) self-complementary circular code X identified in genes of prokaryotes and eukaryotes is a set of 20 trinucleotides enabling reading frame retrieval and maintenance, i.e. a framing code (Arquès and Michel, 1996; Michel, 2012, 2013). Some mitochondrial RNAs correspond to DNA sequences when RNA transcription systematically exchanges between nucleotides (Seligmann, 2013a,b). We study here the 23 bijective transformation codes ΠX of X which may code nucleotide exchanging RNA transcription as suggested by this mitochondrial observation. The 23 bijective transformation codes ΠX are C(3) trinucleotide circular codes, seven of them are also self-complementary. Furthermore, several correlations are observed between the Reading Frame Retrieval (RFR) probability of bijective transformation codes ΠX and the different biological properties of ΠX related to their numbers of RNAs in GenBank's EST database, their polymerization rate, their number of amino acids and the chirality of amino acids they code. Results suggest that the circular code X with the functions of reading frame retrieval and maintenance in regular RNA transcription, may also have, through its bijective transformation codes ΠX, the same functions in nucleotide exchanging RNA transcription. Associations with properties such as amino acid chirality suggest that the RFR of X and its bijective transformations molded the origins of the genetic code's machinery. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Natural Antisense Transcripts: Molecular Mechanisms and Implications in Breast Cancers

PubMed Central

Latgé, Guillaume; Poulet, Christophe; Bours, Vincent; Jerusalem, Guy

2018-01-01

Natural antisense transcripts are RNA sequences that can be transcribed from both DNA strands at the same locus but in the opposite direction from the gene transcript. Because strand-specific high-throughput sequencing of the antisense transcriptome has only been available for less than a decade, many natural antisense transcripts were first described as long non-coding RNAs. Although the precise biological roles of natural antisense transcripts are not known yet, an increasing number of studies report their implication in gene expression regulation. Their expression levels are altered in many physiological and pathological conditions, including breast cancers. Among the potential clinical utilities of the natural antisense transcripts, the non-coding|coding transcript pairs are of high interest for treatment. Indeed, these pairs can be targeted by antisense oligonucleotides to specifically tune the expression of the coding-gene. Here, we describe the current knowledge about natural antisense transcripts, their varying molecular mechanisms as gene expression regulators, and their potential as prognostic or predictive biomarkers in breast cancers. PMID:29301303

Natural Antisense Transcripts: Molecular Mechanisms and Implications in Breast Cancers.

PubMed

Latgé, Guillaume; Poulet, Christophe; Bours, Vincent; Josse, Claire; Jerusalem, Guy

2018-01-02

Natural antisense transcripts are RNA sequences that can be transcribed from both DNA strands at the same locus but in the opposite direction from the gene transcript. Because strand-specific high-throughput sequencing of the antisense transcriptome has only been available for less than a decade, many natural antisense transcripts were first described as long non-coding RNAs. Although the precise biological roles of natural antisense transcripts are not known yet, an increasing number of studies report their implication in gene expression regulation. Their expression levels are altered in many physiological and pathological conditions, including breast cancers. Among the potential clinical utilities of the natural antisense transcripts, the non-coding|coding transcript pairs are of high interest for treatment. Indeed, these pairs can be targeted by antisense oligonucleotides to specifically tune the expression of the coding-gene. Here, we describe the current knowledge about natural antisense transcripts, their varying molecular mechanisms as gene expression regulators, and their potential as prognostic or predictive biomarkers in breast cancers.

Growth hormone and Pit-1 expression in bovine fetal lymphoid cells.

PubMed

Chen, H T; Schuler, L A; Schultz, R D

1997-11-01

Bovine fetal lymphoid cells were examined for growth hormone (GH) and the transcription factor Pit-1/GHF-1 mRNA. GH and Pit-1/GHF-1 transcripts were detected in thymocytes and splenocytes from fetuses at 60, 90, 120, and 270 d of gestation using reverse transcription-polymerase chain reaction (RT-PCR). Northern analysis indicated that the lymphoid GH mRNA was approximately 350 nucleotides larger than in the pituitary. RT-PCR analysis demonstrated that the coding regions as well as 3' untranslated region of the lymphocyte GH and pituitary transcripts were the same. Analysis of the 5'-untranslated region of the lymphocyte GH mRNA showed that transcription began upstream from the start site in the pituitary gland, suggesting differences in regulation in these tissues. Fetal thymocytes and splenocytes expressed Pit-1/GHF-1 mRNA; however, they contained only the 2.5-kb transcript. The GH and Pit-1/GHF-1 mRNA in fetal lymphoid cells supports the hypothesis that lymphocyte-derived GH may function as an autocrine and/or paracrine factor during the development and maturation of the bovine fetal immune system.

Constitutive expression of a salinity-induced wheat WRKY transcription factor enhances salinity and ionic stress tolerance in transgenic Arabidopsis thaliana

DOE Office of Scientific and Technical Information (OSTI.GOV)

Qin, Yuxiang, E-mail: yuxiangqin@126.com; Tian, Yanchen; Han, Lu

Highlights: •A class II WRKY transcription factor, TaWRKY79 was isolated and characterized. •TaWRKY79 was induced by NaCl or abscisic acid. •843 bp regulatory segment was sufficient to respond to ABA or NaCl treatment. •TaWRKY79 enhanced salinity and ionic tolerance while reduced sensitivity to ABA. •TaWRKY79 increased salinity and ionic tolerance in an ABA-dependent pathway. -- Abstract: The isolation and characterization of TaWRKY79, a wheat class II WRKY transcription factor, is described. Its 1297 bp coding region includes a 987 bp long open reading frame. TaWRKY79 was induced by stressing seedlings with either NaCl or abscisic acid (ABA). When a fusionmore » between an 843 bp segment upstream of the TaWRKY79 coding sequence and GUS was introduced into Arabidopsis thaliana, GUS staining indicated that this upstream segment captured the sequence(s) required to respond to ABA or NaCl treatment. When TaWRKY79 was constitutively expressed as a transgene in A. thaliana, the transgenic plants showed an improved capacity to extend their primary root in the presence of either 100 mM NaCl, 10 mM LiCl or 2 μM ABA. The inference was that TaWRKY79 enhanced the level of tolerance to both salinity and ionic stress, while reducing the level of sensitivity to ABA. The ABA-related genes ABA1, ABA2 ABI1 and ABI5 were all up-regulated in the TaWRKY79 transgenic plants, suggesting that the transcription factor operates in an ABA-dependent pathway.« less

Microarray analysis of long non-coding RNA expression profiles in monocytic myeloid-derived suppressor cells in Echinococcus granulosus-infected mice.

PubMed

Yu, Aiping; Wang, Ying; Yin, Jianhai; Zhang, Jing; Cao, Shengkui; Cao, Jianping; Shen, Yujuan

2018-05-30

Cystic echinococcosis is a worldwide chronic zoonotic disease caused by infection with the larval stage of Echinococcus granulosus. Previously, we found significant accumulation of myeloid-derived suppressor cells (MDSCs) in E. granulosus infection mouse models and that they play a key role in immunosuppressing T lymphocytes. Here, we compared the long non-coding RNA (lncRNA) and mRNA expression patterns between the splenic monocytic MDSCs (M-MDSCs) of E. granulosus protoscoleces-infected mice and normal mice using microarray analysis. LncRNA functions were predicted using Gene Ontology enrichment and the Kyoto Encyclopedia of Genes and Genomes pathway analysis. Cis- and trans-regulation analyses revealed potential relationships between the lncRNAs and their target genes or related transcription factors. We found that 649 lncRNAs were differentially expressed (fold change ≥ 2, P < 0.05): 582 lncRNAs were upregulated and 67 lncRNAs were downregulated; respectively, 28 upregulated mRNAs and 1043 downregulated mRNAs were differentially expressed. The microarray data was validated by quantitative reverse transcription-PCR. The results indicated that mRNAs co-expressed with the lncRNAs are mainly involved in regulating the actin cytoskeleton, Salmonella infection, leishmaniasis, and the vascular endothelial growth factor (VEGF) signaling pathway. The lncRNA NONMMUT021591 was predicted to cis-regulate the retinoblastoma gene (Rb1), whose expression is associated with abnormal M-MDSCs differentiation. We found that 372 lncRNAs were predicted to interact with 60 transcription factors; among these, C/EBPβ (CCAAT/enhancer binding protein beta) was previously demonstrated to be a transcription factor of MDSCs. Our study identified dysregulated lncRNAs in the M-MDSCs of E. granulosus infection mouse models; they might be involved in M-MDSC-derived immunosuppression in related diseases.

Antisense transcription is pervasive but rarely conserved in enteric bacteria.

PubMed

Raghavan, Rahul; Sloan, Daniel B; Ochman, Howard

2012-01-01

Noncoding RNAs, including antisense RNAs (asRNAs) that originate from the complementary strand of protein-coding genes, are involved in the regulation of gene expression in all domains of life. Recent application of deep-sequencing technologies has revealed that the transcription of asRNAs occurs genome-wide in bacteria. Although the role of the vast majority of asRNAs remains unknown, it is often assumed that their presence implies important regulatory functions, similar to those of other noncoding RNAs. Alternatively, many antisense transcripts may be produced by chance transcription events from promoter-like sequences that result from the degenerate nature of bacterial transcription factor binding sites. To investigate the biological relevance of antisense transcripts, we compared genome-wide patterns of asRNA expression in closely related enteric bacteria, Escherichia coli and Salmonella enterica serovar Typhimurium, by performing strand-specific transcriptome sequencing. Although antisense transcripts are abundant in both species, less than 3% of asRNAs are expressed at high levels in both species, and only about 14% appear to be conserved among species. And unlike the promoters of protein-coding genes, asRNA promoters show no evidence of sequence conservation between, or even within, species. Our findings suggest that many or even most bacterial asRNAs are nonadaptive by-products of the cell's transcription machinery. IMPORTANCE Application of high-throughput methods has revealed the expression throughout bacterial genomes of transcripts encoded on the strand complementary to protein-coding genes. Because transcription is costly, it is usually assumed that these transcripts, termed antisense RNAs (asRNAs), serve some function; however, the role of most asRNAs is unclear, raising questions about their relevance in cellular processes. Because natural selection conserves functional elements, comparisons between related species provide a method for assessing functionality genome-wide. Applying such an approach, we assayed all transcripts in two closely related bacteria, Escherichia coli and Salmonella enterica serovar Typhimurium, and demonstrate that, although the levels of genome-wide antisense transcription are similarly high in both bacteria, only a small fraction of asRNAs are shared across species. Moreover, the promoters associated with asRNAs show no evidence of sequence conservation between, or even within, species. These findings indicate that despite the genome-wide transcription of asRNAs, many of these transcripts are likely nonfunctional.

Whole transcriptome analysis reveals dysregulated oncogenic lncRNAs in natural killer/T-cell lymphoma and establishes MIR155HG as a target of PRDM1.

PubMed

Baytak, Esra; Gong, Qiang; Akman, Burcu; Yuan, Hongling; Chan, Wing C; Küçük, Can

2017-05-01

Natural killer/T-cell lymphoma is a rare but aggressive neoplasm with poor prognosis. Despite previous reports that showed potential tumor suppressors, such as PRDM1 or oncogenes associated with the etiology of this malignancy, the role of long non-coding RNAs in natural killer/T-cell lymphoma pathobiology has not been addressed to date. Here, we aim to identify cancer-associated dysregulated long non-coding RNAs and signaling pathways or biological processes associated with these long non-coding RNAs in natural killer/T-cell lymphoma cases and to identify the long non-coding RNAs transcriptionally regulated by PRDM1. RNA-Seq analysis revealed 166 and 66 long non-coding RNAs to be significantly overexpressed or underexpressed, respectively, in natural killer/T-cell lymphoma cases compared with resting or activated normal natural killer cells. Novel long non-coding RNAs as well as the cancer-associated ones such as SNHG5, ZFAS1, or MIR155HG were dysregulated. Interestingly, antisense transcripts of many growth-regulating genes appeared to be transcriptionally deregulated. Expression of ZFAS1, which is upregulated in natural killer/T-cell lymphoma cases, showed association with growth-regulating pathways such as stabilization of P53, regulation of apoptosis, cell cycle, or nuclear factor-kappa B signaling in normal and neoplastic natural killer cell samples. Consistent with the tumor suppressive role of PRDM1, we identified MIR155HG and TERC to be transcriptionally downregulated by PRDM1 in two PRDM1-null NK-cell lines when it is ectopically expressed. In conclusion, this is the first study that identified long non-coding RNAs whose expression is dysregulated in natural killer/T-cell lymphoma cases. These findings suggest that ZFAS1 and other dysregulated long non-coding RNAs may be involved in natural killer/T-cell lymphoma pathobiology through regulation of cancer-related genes, and loss-of-PRDM1 expression in natural killer/T-cell lymphomas may contribute to overexpression of MIR155HG; thereby promoting tumorigenesis.

Transcription Factors Encoded on Core and Accessory Chromosomes of Fusarium oxysporum Induce Expression of Effector Genes

PubMed Central

van der Does, H. Charlotte; Schmidt, Sarah M.; Langereis, Léon; Hughes, Timothy R.

2016-01-01

Proteins secreted by pathogens during host colonization largely determine the outcome of pathogen-host interactions and are commonly called ‘effectors’. In fungal plant pathogens, coordinated transcriptional up-regulation of effector genes is a key feature of pathogenesis and effectors are often encoded in genomic regions with distinct repeat content, histone code and rate of evolution. In the tomato pathogen Fusarium oxysporum f. sp. lycopersici (Fol), effector genes reside on one of four accessory chromosomes, known as the ‘pathogenicity’ chromosome, which can be exchanged between strains through horizontal transfer. The three other accessory chromosomes in the Fol reference strain may also be important for virulence towards tomato. Expression of effector genes in Fol is highly up-regulated upon infection and requires Sge1, a transcription factor encoded on the core genome. Interestingly, the pathogenicity chromosome itself contains 13 predicted transcription factor genes and for all except one, there is a homolog on the core genome. We determined DNA binding specificity for nine transcription factors using oligonucleotide arrays. The binding sites for homologous transcription factors were highly similar, suggesting that extensive neofunctionalization of DNA binding specificity has not occurred. Several DNA binding sites are enriched on accessory chromosomes, and expression of FTF1, its core homolog FTF2 and SGE1 from a constitutive promoter can induce expression of effector genes. The DNA binding sites of only these three transcription factors are enriched among genes up-regulated during infection. We further show that Ftf1, Ftf2 and Sge1 can activate transcription from their binding sites in yeast. RNAseq analysis revealed that in strains with constitutive expression of FTF1, FTF2 or SGE1, expression of a similar set of plant-responsive genes on the pathogenicity chromosome is induced, including most effector genes. We conclude that the Fol pathogenicity chromosome may be partially transcriptionally autonomous, but there are also extensive transcriptional connections between core and accessory chromosomes. PMID:27855160

Differentiated evolutionary conservatism and lack of polymorphism of crucial sex determination genes (SRY and SOX9) in four species of the family Canidae.

PubMed

Nowacka-Woszuk, Joanna; Switonski, Marek

2009-01-01

The sex determination process is under the control of several genes of which two (SRY and SOX9), encoding transcription factors, play a crucial role. It is well-known that mutations at these genes may cause the development of an intersexual phenotype. The aim of this study was to conduct a comparative analysis of the coding sequence and 5'-flanking regions of both genes in four species of the family Canidae (the dog, red fox, arctic fox and Chinese raccoon dog). Similarity of the coding sequence of the SOX9 gene among the studied species was higher (99.7-99.9%) than in the case of the SRY gene (96.7-97.3%). Only single nucleotide changes were found in the compared coding sequences, whereas in the 5'-flanking region of both genes nucleotide substitutions, as well as insertions and deletions were observed. None of the changes detected in the 5'-flanking region occurred within the potential consensus sequences for transcription factors. No polymorphism was found for either of these genes in any of the analyzed species.

Diversity of Antisense and Other Non-Coding RNAs in Archaea Revealed by Comparative Small RNA Sequencing in Four Pyrobaculum Species

PubMed Central

Bernick, David L.; Dennis, Patrick P.; Lui, Lauren M.; Lowe, Todd M.

2012-01-01

A great diversity of small, non-coding RNA (ncRNA) molecules with roles in gene regulation and RNA processing have been intensely studied in eukaryotic and bacterial model organisms, yet our knowledge of possible parallel roles for small RNAs (sRNA) in archaea is limited. We employed RNA-seq to identify novel sRNA across multiple species of the hyperthermophilic genus Pyrobaculum, known for unusual RNA gene characteristics. By comparing transcriptional data collected in parallel among four species, we were able to identify conserved RNA genes fitting into known and novel families. Among our findings, we highlight three novel cis-antisense sRNAs encoded opposite to key regulatory (ferric uptake regulator), metabolic (triose-phosphate isomerase), and core transcriptional apparatus genes (transcription factor B). We also found a large increase in the number of conserved C/D box sRNA genes over what had been previously recognized; many of these genes are encoded antisense to protein coding genes. The conserved opposition to orthologous genes across the Pyrobaculum genus suggests similarities to other cis-antisense regulatory systems. Furthermore, the genus-specific nature of these sRNAs indicates they are relatively recent, stable adaptations. PMID:22783241

RNAi mediates post-transcriptional repression of gene expression in fission yeast Schizosaccharomyces pombe

DOE Office of Scientific and Technical Information (OSTI.GOV)

Smialowska, Agata, E-mail: smialowskaa@gmail.com; School of Life Sciences, Södertörn Högskola, Huddinge 141-89; Djupedal, Ingela

Highlights: • Protein coding genes accumulate anti-sense sRNAs in fission yeast S. pombe. • RNAi represses protein-coding genes in S. pombe. • RNAi-mediated gene repression is post-transcriptional. - Abstract: RNA interference (RNAi) is a gene silencing mechanism conserved from fungi to mammals. Small interfering RNAs are products and mediators of the RNAi pathway and act as specificity factors in recruiting effector complexes. The Schizosaccharomyces pombe genome encodes one of each of the core RNAi proteins, Dicer, Argonaute and RNA-dependent RNA polymerase (dcr1, ago1, rdp1). Even though the function of RNAi in heterochromatin assembly in S. pombe is established, its rolemore » in controlling gene expression is elusive. Here, we report the identification of small RNAs mapped anti-sense to protein coding genes in fission yeast. We demonstrate that these genes are up-regulated at the protein level in RNAi mutants, while their mRNA levels are not significantly changed. We show that the repression by RNAi is not a result of heterochromatin formation. Thus, we conclude that RNAi is involved in post-transcriptional gene silencing in S. pombe.« less

Gene-specific cell labeling using MiMIC transposons

PubMed Central

Gnerer, Joshua P.; Venken, Koen J. T.; Dierick, Herman A.

2015-01-01

Binary expression systems such as GAL4/UAS, LexA/LexAop and QF/QUAS have greatly enhanced the power of Drosophila as a model organism by allowing spatio-temporal manipulation of gene function as well as cell and neural circuit function. Tissue-specific expression of these heterologous transcription factors relies on random transposon integration near enhancers or promoters that drive the binary transcription factor embedded in the transposon. Alternatively, gene-specific promoter elements are directly fused to the binary factor within the transposon followed by random or site-specific integration. However, such insertions do not consistently recapitulate endogenous expression. We used Minos-Mediated Integration Cassette (MiMIC) transposons to convert host loci into reliable gene-specific binary effectors. MiMIC transposons allow recombinase-mediated cassette exchange to modify the transposon content. We developed novel exchange cassettes to convert coding intronic MiMIC insertions into gene-specific binary factor protein-traps. In addition, we expanded the set of binary factor exchange cassettes available for non-coding intronic MiMIC insertions. We show that binary factor conversions of different insertions in the same locus have indistinguishable expression patterns, suggesting that they reliably reflect endogenous gene expression. We show the efficacy and broad applicability of these new tools by dissecting the cellular expression patterns of the Drosophila serotonin receptor gene family. PMID:25712101

Polymerization of non-complementary RNA: systematic symmetric nucleotide exchanges mainly involving uracil produce mitochondrial RNA transcripts coding for cryptic overlapping genes.

PubMed

Seligmann, Hervé

2013-03-01

Usual DNA→RNA transcription exchanges T→U. Assuming different systematic symmetric nucleotide exchanges during translation, some GenBank RNAs match exactly human mitochondrial sequences (exchange rules listed in decreasing transcript frequencies): C↔U, A↔U, A↔U+C↔G (two nucleotide pairs exchanged), G↔U, A↔G, C↔G, none for A↔C, A↔G+C↔U, and A↔C+G↔U. Most unusual transcripts involve exchanging uracil. Independent measures of rates of rare replicational enzymatic DNA nucleotide misinsertions predict frequencies of RNA transcripts systematically exchanging the corresponding misinserted nucleotides. Exchange transcripts self-hybridize less than other gene regions, self-hybridization increases with length, suggesting endoribonuclease-limited elongation. Blast detects stop codon depleted putative protein coding overlapping genes within exchange-transcribed mitochondrial genes. These align with existing GenBank proteins (mainly metazoan origins, prokaryotic and viral origins underrepresented). These GenBank proteins frequently interact with RNA/DNA, are membrane transporters, or are typical of mitochondrial metabolism. Nucleotide exchange transcript frequencies increase with overlapping gene densities and stop densities, indicating finely tuned counterbalancing regulation of expression of systematic symmetric nucleotide exchange-encrypted proteins. Such expression necessitates combined activities of suppressor tRNAs matching stops, and nucleotide exchange transcription. Two independent properties confirm predicted exchanged overlap coding genes: discrepancy of third codon nucleotide contents from replicational deamination gradients, and codon usage according to circular code predictions. Predictions from both properties converge, especially for frequent nucleotide exchange types. Nucleotide exchanging transcription apparently increases coding densities of protein coding genes without lengthening genomes, revealing unsuspected functional DNA coding potential. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

A purified transcription factor (TIF-IB) binds to essential sequences of the mouse rDNA promoter.

PubMed Central

Clos, J; Buttgereit, D; Grummt, I

1986-01-01

A transcription factor that is specific for mouse rDNA has been partially purified from Ehrlich ascites cells. This factor [designated transcription initiation factor (TIF)-IB] is required for accurate in vitro synthesis of mouse rRNA in addition to RNA polymerase I and another regulatory factor, TIF-IA. TIF-IB activity is present in extracts both from growing and nongrowing cells in comparable amounts. Prebinding competition experiments with wild-type and mutant templates suggest that TIF-IB interacts with the core control element of the rDNA promoter, which is located immediately upstream of the initiation site. The specific binding of TIF-IB to the RNA polymerase I promoter is demonstrated by exonuclease III protection experiments. The 3' border of the sequences protected by TIF-IB is shown to be on the coding strand at position -21 and on the noncoding strand at position -7. The results suggest that direct binding of TIF-IB to sequences in the core promoter element is the mechanism by which this factor imparts promoter selectivity to RNA polymerase I. Images PMID:3456157

The artificial zinc finger coding gene 'Jazz' binds the utrophin promoter and activates transcription.

PubMed

Corbi, N; Libri, V; Fanciulli, M; Tinsley, J M; Davies, K E; Passananti, C

2000-06-01

Up-regulation of utrophin gene expression is recognized as a plausible therapeutic approach in the treatment of Duchenne muscular dystrophy (DMD). We have designed and engineered new zinc finger-based transcription factors capable of binding and activating transcription from the promoter of the dystrophin-related gene, utrophin. Using the recognition 'code' that proposes specific rules between zinc finger primary structure and potential DNA binding sites, we engineered a new gene named 'Jazz' that encodes for a three-zinc finger peptide. Jazz belongs to the Cys2-His2 zinc finger type and was engineered to target the nine base pair DNA sequence: 5'-GCT-GCT-GCG-3', present in the promoter region of both the human and mouse utrophin gene. The entire zinc finger alpha-helix region, containing the amino acid positions that are crucial for DNA binding, was specifically chosen on the basis of the contacts more frequently represented in the available list of the 'code'. Here we demonstrate that Jazz protein binds specifically to the double-stranded DNA target, with a dissociation constant of about 32 nM. Band shift and super-shift experiments confirmed the high affinity and specificity of Jazz protein for its DNA target. Moreover, we show that chimeric proteins, named Gal4-Jazz and Sp1-Jazz, are able to drive the transcription of a test gene from the human utrophin promoter.

Ectopic expression of different cytokinin-regulated transcription factor genes of Arabidopsis thaliana alters plant growth and development.

PubMed

Köllmer, Ireen; Werner, Tomáš; Schmülling, Thomas

2011-08-15

The plant hormone cytokinin rapidly alters the steady state transcript levels of a number of transcription factor genes suggesting that these might have a function in mediating cytokinin effects. Here we report the analysis of Arabidopsis thaliana plants with an altered expression level of four different cytokinin-regulated transcription factor genes. These include GATA22 (also known as CGA1/GNL), two genes coding for members of the homeodomain zip (HD zip) class II transcription factor family (HAT4, HAT22), and bHLH64. Ectopic expression of the GATA22 gene induced the development of chloroplasts in root tissue where it is normally suppressed and led to the formation of shorter and less branched roots. Overexpression of HAT22 lowered the seedlings chlorophyll content and caused an earlier onset of leaf senescence. Enhanced expression of the HAT4 gene led to severe defects in inflorescence stem development and to a decrease in root growth and branching, while hat4 insertional mutants developed a larger root system. 35S:bHLH64 transgenic plants showed a pleiotropic phenotype, consisting of larger rosettes, reduced chlorophyll content and an elongated and thickened hypocotyl. Flower development was strongly disturbed leading to sterile plants. The results are consistent with specific functions of these transcription factor genes in regulating part of the cytokinin activities and suggest their action as convergence point with other signalling pathways, particularly those of gibberellin and light. Copyright © 2011 Elsevier GmbH. All rights reserved.

Molecular Phylogenetic and Expression Analysis of the Complete WRKY Transcription Factor Family in Maize

PubMed Central

Wei, Kai-Fa; Chen, Juan; Chen, Yan-Feng; Wu, Ling-Juan; Xie, Dao-Xin

2012-01-01

The WRKY transcription factors function in plant growth and development, and response to the biotic and abiotic stresses. Although many studies have focused on the functional identification of the WRKY transcription factors, much less is known about molecular phylogenetic and global expression analysis of the complete WRKY family in maize. In this study, we identified 136 WRKY proteins coded by 119 genes in the B73 inbred line from the complete genome and named them in an orderly manner. Then, a comprehensive phylogenetic analysis of five species was performed to explore the origin and evolutionary patterns of these WRKY genes, and the result showed that gene duplication is the major driving force for the origin of new groups and subgroups and functional divergence during evolution. Chromosomal location analysis of maize WRKY genes indicated that 20 gene clusters are distributed unevenly in the genome. Microarray-based expression analysis has revealed that 131 WRKY transcripts encoded by 116 genes may participate in the regulation of maize growth and development. Among them, 102 transcripts are stably expressed with a coefficient of variation (CV) value of <15%. The remaining 29 transcripts produced by 25 WRKY genes with the CV value of >15% are further analysed to discover new organ- or tissue-specific genes. In addition, microarray analyses of transcriptional responses to drought stress and fungal infection showed that maize WRKY proteins are involved in stress responses. All these results contribute to a deep probing into the roles of WRKY transcription factors in maize growth and development and stress tolerance. PMID:22279089

Molecular phylogenetic and expression analysis of the complete WRKY transcription factor family in maize.

PubMed

Wei, Kai-Fa; Chen, Juan; Chen, Yan-Feng; Wu, Ling-Juan; Xie, Dao-Xin

2012-04-01

The WRKY transcription factors function in plant growth and development, and response to the biotic and abiotic stresses. Although many studies have focused on the functional identification of the WRKY transcription factors, much less is known about molecular phylogenetic and global expression analysis of the complete WRKY family in maize. In this study, we identified 136 WRKY proteins coded by 119 genes in the B73 inbred line from the complete genome and named them in an orderly manner. Then, a comprehensive phylogenetic analysis of five species was performed to explore the origin and evolutionary patterns of these WRKY genes, and the result showed that gene duplication is the major driving force for the origin of new groups and subgroups and functional divergence during evolution. Chromosomal location analysis of maize WRKY genes indicated that 20 gene clusters are distributed unevenly in the genome. Microarray-based expression analysis has revealed that 131 WRKY transcripts encoded by 116 genes may participate in the regulation of maize growth and development. Among them, 102 transcripts are stably expressed with a coefficient of variation (CV) value of <15%. The remaining 29 transcripts produced by 25 WRKY genes with the CV value of >15% are further analysed to discover new organ- or tissue-specific genes. In addition, microarray analyses of transcriptional responses to drought stress and fungal infection showed that maize WRKY proteins are involved in stress responses. All these results contribute to a deep probing into the roles of WRKY transcription factors in maize growth and development and stress tolerance.

Perspectives on the mechanism of transcriptional regulation by long non-coding RNAs.

PubMed

Roberts, Thomas C; Morris, Kevin V; Weinberg, Marc S

2014-01-01

Long non-coding RNAs (lncRNAs) are increasingly being recognized as epigenetic regulators of gene transcription. The diversity and complexity of lncRNA genes means that they exert their regulatory effects by a variety of mechanisms. Although there is still much to be learned about the mechanism of lncRNA function, general principles are starting to emerge. In particular, the application of high throughput (deep) sequencing methodologies has greatly advanced our understanding of lncRNA gene function. lncRNAs function as adaptors that link specific chromatin loci with chromatin-remodeling complexes and transcription factors. lncRNAs can act in cis or trans to guide epigenetic-modifier complexes to distinct genomic sites, or act as scaffolds which recruit multiple proteins simultaneously, thereby coordinating their activities. In this review we discuss the genomic organization of lncRNAs, the importance of RNA secondary structure to lncRNA functionality, the multitude of ways in which they interact with the genome, and what evolutionary conservation tells us about their function.

Long Noncoding RNAs: New Players in the Osteogenic Differentiation of Bone Marrow- and Adipose-Derived Mesenchymal Stem Cells.

PubMed

Yang, Qiaolin; Jia, Lingfei; Li, Xiaobei; Guo, Runzhi; Huang, Yiping; Zheng, Yunfei; Li, Weiran

2018-06-01

Mesenchymal stem cells (MSCs) are an important population of multipotent stem cells that differentiate into multiple lineages and display great potential in bone regeneration and repair. Although the role of protein-coding genes in the osteogenic differentiation of MSCs has been extensively studied, the functions of noncoding RNAs in the osteogenic differentiation of MSCs are unclear. The recent application of next-generation sequencing to MSC transcriptomes has revealed that long noncoding RNAs (lncRNAs) are associated with the osteogenic differentiation of MSCs. LncRNAs are a class of non-coding transcripts of more than 200 nucleotides in length. Noncoding RNAs are thought to play a key role in osteoblast differentiation through various regulatory mechanisms including chromatin modification, transcription factor binding, competent endogenous mechanism, and other post-transcriptional mechanisms. Here, we review the roles of lncRNAs in the osteogenic differentiation of bone marrow- and adipose-derived stem cells and provide a theoretical foundation for future research.

Improving the genome annotation of the acarbose producer Actinoplanes sp. SE50/110 by sequencing enriched 5'-ends of primary transcripts.

PubMed

Schwientek, Patrick; Neshat, Armin; Kalinowski, Jörn; Klein, Andreas; Rückert, Christian; Schneiker-Bekel, Susanne; Wendler, Sergej; Stoye, Jens; Pühler, Alfred

2014-11-20

Actinoplanes sp. SE50/110 is the producer of the alpha-glucosidase inhibitor acarbose, which is an economically relevant and potent drug in the treatment of type-2 diabetes mellitus. In this study, we present the detection of transcription start sites on this genome by sequencing enriched 5'-ends of primary transcripts. Altogether, 1427 putative transcription start sites were initially identified. With help of the annotated genome sequence, 661 transcription start sites were found to belong to the leader region of protein-coding genes with the surprising result that roughly 20% of these genes rank among the class of leaderless transcripts. Next, conserved promoter motifs were identified for protein-coding genes with and without leader sequences. The mapped transcription start sites were finally used to improve the annotation of the Actinoplanes sp. SE50/110 genome sequence. Concerning protein-coding genes, 41 translation start sites were corrected and 9 novel protein-coding genes could be identified. In addition to this, 122 previously undetermined non-coding RNA (ncRNA) genes of Actinoplanes sp. SE50/110 were defined. Focusing on antisense transcription start sites located within coding genes or their leader sequences, it was discovered that 96 of those ncRNA genes belong to the class of antisense RNA (asRNA) genes. The remaining 26 ncRNA genes were found outside of known protein-coding genes. Four chosen examples of prominent ncRNA genes, namely the transfer messenger RNA gene ssrA, the ribonuclease P class A RNA gene rnpB, the cobalamin riboswitch RNA gene cobRS, and the selenocysteine-specific tRNA gene selC, are presented in more detail. This study demonstrates that sequencing of enriched 5'-ends of primary transcripts and the identification of transcription start sites are valuable tools for advanced genome annotation of Actinoplanes sp. SE50/110 and most probably also for other bacteria. Copyright © 2014 Elsevier B.V. All rights reserved.

Current knowledge of microRNA-mediated regulation of drug metabolism in humans.

PubMed

Nakano, Masataka; Nakajima, Miki

2018-05-01

Understanding the factors causing inter- and intra-individual differences in drug metabolism potencies is required for the practice of personalized or precision medicine, as well as for the promotion of efficient drug development. The expression of drug-metabolizing enzymes is controlled by transcriptional regulation by nuclear receptors and transcriptional factors, epigenetic regulation, such as DNA methylation and histone acetylation, and post-translational modification. In addition to such regulation mechanisms, recent studies revealed that microRNAs (miRNAs), endogenous ~22-nucleotide non-coding RNAs that regulate gene expression through the translational repression and degradation of mRNAs, significantly contribute to post-transcriptional regulation of drug-metabolizing enzymes. Areas covered: This review summarizes the current knowledge regarding miRNAs-dependent regulation of drug-metabolizing enzymes and transcriptional factors and its physiological and clinical significance. We also describe recent advances in miRNA-dependent regulation research, showing that the presence of pseudogenes, single-nucleotide polymorphisms, and RNA editing affects miRNA targeting. Expert opinion: It is unwavering fact that miRNAs are critical factors causing inter- and intra-individual differences in the expression of drug-metabolizing enzymes. Consideration of miRNA-dependent regulation would be a helpful tool for optimizing personalized and precision medicine.

Analysis of the TCP genes expressed in the inflorescence of the orchid Orchis italica

PubMed Central

De Paolo, Sofia; Gaudio, Luciano; Aceto, Serena

2015-01-01

TCP proteins are plant-specific transcription factors involved in many different processes. Because of their involvement in a large number of developmental pathways, their roles have been investigated in various plant species. However, there are almost no studies of this transcription factor family in orchids. Based on the available transcriptome of the inflorescence of the orchid Orchis italica, in the present study we identified 12 transcripts encoding TCP proteins. The phylogenetic analysis showed that they belong to different TCP classes (I and II) and groups (PCF, CIN and CYC/TB1), and that they display a number of conserved motifs when compared with the TCPs of Arabidopsis and Oryza. The presence of a specific cleavage site for the microRNA miRNA319, an important post-transcriptional regulator of several TCP genes in other species, was demonstrated for one transcript of O. italica, and the analysis of the expression pattern of the TCP transcripts in different inflorescence organs and in leaf tissue suggests that some TCP transcripts of O. italica exert their role only in specific tissues, while others may play multiple roles in different tissues. In addition, the evolutionary analysis showed a general purifying selection acting on the coding region of these transcripts. PMID:26531864

Analysis of the TCP genes expressed in the inflorescence of the orchid Orchis italica.

PubMed

De Paolo, Sofia; Gaudio, Luciano; Aceto, Serena

2015-11-04

TCP proteins are plant-specific transcription factors involved in many different processes. Because of their involvement in a large number of developmental pathways, their roles have been investigated in various plant species. However, there are almost no studies of this transcription factor family in orchids. Based on the available transcriptome of the inflorescence of the orchid Orchis italica, in the present study we identified 12 transcripts encoding TCP proteins. The phylogenetic analysis showed that they belong to different TCP classes (I and II) and groups (PCF, CIN and CYC/TB1), and that they display a number of conserved motifs when compared with the TCPs of Arabidopsis and Oryza. The presence of a specific cleavage site for the microRNA miRNA319, an important post-transcriptional regulator of several TCP genes in other species, was demonstrated for one transcript of O. italica, and the analysis of the expression pattern of the TCP transcripts in different inflorescence organs and in leaf tissue suggests that some TCP transcripts of O. italica exert their role only in specific tissues, while others may play multiple roles in different tissues. In addition, the evolutionary analysis showed a general purifying selection acting on the coding region of these transcripts.

A Noncoding, Regulatory Mutation Implicates HCFC1 in Nonsyndromic Intellectual Disability

PubMed Central

Huang, Lingli; Jolly, Lachlan A.; Willis-Owen, Saffron; Gardner, Alison; Kumar, Raman; Douglas, Evelyn; Shoubridge, Cheryl; Wieczorek, Dagmar; Tzschach, Andreas; Cohen, Monika; Hackett, Anna; Field, Michael; Froyen, Guy; Hu, Hao; Haas, Stefan A.; Ropers, Hans-Hilger; Kalscheuer, Vera M.; Corbett, Mark A.; Gecz, Jozef

2012-01-01

The discovery of mutations causing human disease has so far been biased toward protein-coding regions. Having excluded all annotated coding regions, we performed targeted massively parallel resequencing of the nonrepetitive genomic linkage interval at Xq28 of family MRX3. We identified in the binding site of transcription factor YY1 a regulatory mutation that leads to overexpression of the chromatin-associated transcriptional regulator HCFC1. When tested on embryonic murine neural stem cells and embryonic hippocampal neurons, HCFC1 overexpression led to a significant increase of the production of astrocytes and a considerable reduction in neurite growth. Two other nonsynonymous, potentially deleterious changes have been identified by X-exome sequencing in individuals with intellectual disability, implicating HCFC1 in normal brain function. PMID:23000143

LncRNApred: Classification of Long Non-Coding RNAs and Protein-Coding Transcripts by the Ensemble Algorithm with a New Hybrid Feature.

PubMed

Pian, Cong; Zhang, Guangle; Chen, Zhi; Chen, Yuanyuan; Zhang, Jin; Yang, Tao; Zhang, Liangyun

2016-01-01

As a novel class of noncoding RNAs, long noncoding RNAs (lncRNAs) have been verified to be associated with various diseases. As large scale transcripts are generated every year, it is significant to accurately and quickly identify lncRNAs from thousands of assembled transcripts. To accurately discover new lncRNAs, we develop a classification tool of random forest (RF) named LncRNApred based on a new hybrid feature. This hybrid feature set includes three new proposed features, which are MaxORF, RMaxORF and SNR. LncRNApred is effective for classifying lncRNAs and protein coding transcripts accurately and quickly. Moreover,our RF model only requests the training using data on human coding and non-coding transcripts. Other species can also be predicted by using LncRNApred. The result shows that our method is more effective compared with the Coding Potential Calculate (CPC). The web server of LncRNApred is available for free at http://mm20132014.wicp.net:57203/LncRNApred/home.jsp.

A compendium of transcription factor and Transcriptionally active protein coding gene families in cowpea (Vigna unguiculata L.).

PubMed

Misra, Vikram A; Wang, Yu; Timko, Michael P

2017-11-22

Cowpea (Vigna unguiculata (L.) Walp.) is the most important food and forage legume in the semi-arid tropics of sub-Saharan Africa where approximately 80% of worldwide production takes place primarily on low-input, subsistence farm sites. Among the major goals of cowpea breeding and improvement programs are the rapid manipulation of agronomic traits for seed size and quality and improved resistance to abiotic and biotic stresses to enhance productivity. Knowing the suite of transcription factors (TFs) and transcriptionally active proteins (TAPs) that control various critical plant cellular processes would contribute tremendously to these improvement aims. We used a computational approach that employed three different predictive pipelines to data mine the cowpea genome and identified over 4400 genes representing 136 different TF and TAP families. We compare the information content of cowpea to two evolutionarily close species common bean (Phaseolus vulgaris), and soybean (Glycine max) to gauge the relative informational content. Our data indicate that correcting for genome size cowpea has fewer TF and TAP genes than common bean (4408 / 5291) and soybean (4408/ 11,065). Members of the GROWTH-REGULATING FACTOR (GRF) and Auxin/indole-3-acetic acid (Aux/IAA) gene families appear to be over-represented in the genome relative to common bean and soybean, whereas members of the MADS (Minichromosome maintenance deficient 1 (MCM1), AGAMOUS, DEFICIENS, and serum response factor (SRF)) and C2C2-YABBY appear to be under-represented. Analysis of the AP2-EREBP APETALA2-Ethylene Responsive Element Binding Protein (AP2-EREBP), NAC (NAM (no apical meristem), ATAF1, 2 (Arabidopsis transcription activation factor), CUC (cup-shaped cotyledon)), and WRKY families, known to be important in defense signaling, revealed changes and phylogenetic rearrangements relative to common bean and soybean that suggest these groups may have evolved different functions. The availability of detailed information on the coding capacity of the cowpea genome and in particular the various TF and TAP gene families will facilitate future comparative analysis and development of strategies for controlling growth, differentiation, and abiotic and biotic stress resistances of cowpea.

Systematic Profiling of Poly(A)+ Transcripts Modulated by Core 3’ End Processing and Splicing Factors Reveals Regulatory Rules of Alternative Cleavage and Polyadenylation

PubMed Central

Li, Wencheng; You, Bei; Hoque, Mainul; Zheng, Dinghai; Luo, Wenting; Ji, Zhe; Park, Ji Yeon; Gunderson, Samuel I.; Kalsotra, Auinash; Manley, James L.; Tian, Bin

2015-01-01

Alternative cleavage and polyadenylation (APA) results in mRNA isoforms containing different 3’ untranslated regions (3’UTRs) and/or coding sequences. How core cleavage/polyadenylation (C/P) factors regulate APA is not well understood. Using siRNA knockdown coupled with deep sequencing, we found that several C/P factors can play significant roles in 3’UTR-APA. Whereas Pcf11 and Fip1 enhance usage of proximal poly(A) sites (pAs), CFI-25/68, PABPN1 and PABPC1 promote usage of distal pAs. Strong cis element biases were found for pAs regulated by CFI-25/68 or Fip1, and the distance between pAs plays an important role in APA regulation. In addition, intronic pAs are substantially regulated by splicing factors, with U1 mostly inhibiting C/P events in introns near the 5’ end of gene and U2 suppressing those in introns with features for efficient splicing. Furthermore, PABPN1 inhibits expression of transcripts with pAs near the transcription start site (TSS), a property possibly related to its role in RNA degradation. Finally, we found that groups of APA events regulated by C/P factors are also modulated in cell differentiation and development with distinct trends. Together, our results support an APA code where an APA event in a given cellular context is regulated by a number of parameters, including relative location to the TSS, splicing context, distance between competing pAs, surrounding cis elements and concentrations of core C/P factors. PMID:25906188

Identification and Classification of New Transcripts in Dorper and Small-Tailed Han Sheep Skeletal Muscle Transcriptomes.

PubMed

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

2016-01-01

High-throughput mRNA sequencing enables the discovery of new transcripts and additional parts of incompletely annotated transcripts. Compared with the human and cow genomes, the reference annotation level of the sheep genome is still low. An investigation of new transcripts in sheep skeletal muscle will improve our understanding of muscle development. Therefore, applying high-throughput sequencing, two cDNA libraries from the biceps brachii of small-tailed Han sheep and Dorper sheep were constructed, and whole-transcriptome analysis was performed to determine the unknown transcript catalogue of this tissue. In this study, 40,129 transcripts were finally mapped to the sheep genome. Among them, 3,467 transcripts were determined to be unannotated in the current reference sheep genome and were defined as new transcripts. Based on protein-coding capacity prediction and comparative analysis of sequence similarity, 246 transcripts were classified as portions of unannotated genes or incompletely annotated genes. Another 1,520 transcripts were predicted with high confidence to be long non-coding RNAs. Our analysis also revealed 334 new transcripts that displayed specific expression in ruminants and uncovered a number of new transcripts without intergenus homology but with specific expression in sheep skeletal muscle. The results confirmed a complex transcript pattern of coding and non-coding RNA in sheep skeletal muscle. This study provided important information concerning the sheep genome and transcriptome annotation, which could provide a basis for further study.

Modification of tRNALys UUU by Elongator Is Essential for Efficient Translation of Stress mRNAs

PubMed Central

Sansó, Miriam; Buhne, Karin; Carmona, Mercè; Paulo, Esther; Hermand, Damien; Rodríguez-Gabriel, Miguel; Ayté, José; Leidel, Sebastian; Hidalgo, Elena

2013-01-01

The Elongator complex, including the histone acetyl transferase Sin3/Elp3, was isolated as an RNA polymerase II-interacting complex, and cells deficient in Elongator subunits display transcriptional defects. However, it has also been shown that Elongator mediates the modification of some tRNAs, modulating translation efficiency. We show here that the fission yeast Sin3/Elp3 is important for oxidative stress survival. The stress transcriptional program, governed by the Sty1-Atf1-Pcr1 pathway, is affected in mutant cells, but not severely. On the contrary, cells lacking Sin3/Elp3 cannot modify the uridine wobble nucleoside of certain tRNAs, and other tRNA modifying activities such as Ctu1-Ctu2 are also essential for normal tolerance to H2O2. In particular, a plasmid over-expressing the tRNALys UUU complements the stress-related phenotypes of Sin3/Elp3 mutant cells. We have determined that the main H2O2-dependent genes, including those coding for the transcription factors Atf1 and Pcr1, are highly expressed mRNAs containing a biased number of lysine-coding codons AAA versus AAG. Thus, their mRNAs are poorly translated after stress in cells lacking Sin3/Elp3 or Ctu2, whereas a mutated atf1 transcript with AAA-to-AAG lysine codons is efficiently translated in all strain backgrounds. Our study demonstrates that the lack of a functional Elongator complex results in stress phenotypes due to its contribution to tRNA modification and subsequent translation inefficiency of certain stress-induced, highly expressed mRNAs. These results suggest that the transcriptional defects of these strain backgrounds may be a secondary consequence of the deficient expression of a transcription factor, Atf1-Pcr1, and other components of the transcriptional machinery. PMID:23874237

Modification of tRNA(Lys) UUU by elongator is essential for efficient translation of stress mRNAs.

PubMed

Fernández-Vázquez, Jorge; Vargas-Pérez, Itzel; Sansó, Miriam; Buhne, Karin; Carmona, Mercè; Paulo, Esther; Hermand, Damien; Rodríguez-Gabriel, Miguel; Ayté, José; Leidel, Sebastian; Hidalgo, Elena

2013-01-01

The Elongator complex, including the histone acetyl transferase Sin3/Elp3, was isolated as an RNA polymerase II-interacting complex, and cells deficient in Elongator subunits display transcriptional defects. However, it has also been shown that Elongator mediates the modification of some tRNAs, modulating translation efficiency. We show here that the fission yeast Sin3/Elp3 is important for oxidative stress survival. The stress transcriptional program, governed by the Sty1-Atf1-Pcr1 pathway, is affected in mutant cells, but not severely. On the contrary, cells lacking Sin3/Elp3 cannot modify the uridine wobble nucleoside of certain tRNAs, and other tRNA modifying activities such as Ctu1-Ctu2 are also essential for normal tolerance to H2O2. In particular, a plasmid over-expressing the tRNA(Lys) UUU complements the stress-related phenotypes of Sin3/Elp3 mutant cells. We have determined that the main H2O2-dependent genes, including those coding for the transcription factors Atf1 and Pcr1, are highly expressed mRNAs containing a biased number of lysine-coding codons AAA versus AAG. Thus, their mRNAs are poorly translated after stress in cells lacking Sin3/Elp3 or Ctu2, whereas a mutated atf1 transcript with AAA-to-AAG lysine codons is efficiently translated in all strain backgrounds. Our study demonstrates that the lack of a functional Elongator complex results in stress phenotypes due to its contribution to tRNA modification and subsequent translation inefficiency of certain stress-induced, highly expressed mRNAs. These results suggest that the transcriptional defects of these strain backgrounds may be a secondary consequence of the deficient expression of a transcription factor, Atf1-Pcr1, and other components of the transcriptional machinery.

Pharmacological profile of brain-derived neurotrophic factor (BDNF) splice variant translation using a novel drug screening assay: a "quantitative code".

PubMed

Vaghi, Valentina; Polacchini, Alessio; Baj, Gabriele; Pinheiro, Vera L M; Vicario, Annalisa; Tongiorgi, Enrico

2014-10-03

The neurotrophin brain-derived neurotrophic factor (BDNF) is a key regulator of neuronal development and plasticity. BDNF is a major pharmaceutical target in neurodevelopmental and psychiatric disorders. However, pharmacological modulation of this neurotrophin is challenging because BDNF is generated by multiple, alternatively spliced transcripts with different 5'- and 3'UTRs. Each BDNF mRNA variant is transcribed independently, but translation regulation is unknown. To evaluate the translatability of BDNF transcripts, we developed an in vitro luciferase assay in human neuroblastoma cells. In unstimulated cells, each BDNF 5'- and 3'UTR determined a different basal translation level of the luciferase reporter gene. However, constructs with either a 5'UTR or a 3'UTR alone showed poor translation modulation by BDNF, KCl, dihydroxyphenylglycine, AMPA, NMDA, dopamine, acetylcholine, norepinephrine, or serotonin. Constructs consisting of the luciferase reporter gene flanked by the 5'UTR of one of the most abundant BDNF transcripts in the brain (exons 1, 2c, 4, and 6) and the long 3'UTR responded selectively to stimulation with the different receptor agonists, and only transcripts 2c and 6 were increased by the antidepressants desipramine and mirtazapine. We propose that BDNF mRNA variants represent "a quantitative code" for regulated expression of the protein. Thus, to discriminate the efficacy of drugs in stimulating BDNF synthesis, it is appropriate to use variant-specific in vitro screening tests. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

8D.07: GENE EXPRESSION ANALYSIS AND BIOINFORMATICS REVEALED POTENTIAL TRANSCRIPTION FACTORS ASSOCIATED WITH RENIN-ANGIOTENSIN-ALDOSTERONE SYSTEM IN ATHEROMA.

PubMed

Nehme, A; Zibara, K; Cerutti, C; Bricca, G

2015-06-01

The implication of the renin-angiotensin-aldosterone system (RAAS) in atheroma development is well described. However, a complete view of the local RAAS in atheroma is still missing. In this study we aimed to reveal the organization of RAAS in atheroma at the transcriptomic level and identify the transcriptional regulators behind it. Extended RAAS (extRAAS) was defined as the set of 37 genes coding for classical and novel RAAS participants (Figure 1). Five microarray datasets containing overall 590 samples representing carotid and peripheral atheroma were downloaded from the GEO database. Correlation-based hierarchical clustering (R software) of extRAAS genes within each dataset allowed the identification of modules of co-expressed genes. Reproducible co-expression modules across datasets were then extracted. Transcription factors (TFs) having common binding sites (TFBSs) in the promoters of coordinated genes were identified using the Genomatix database tools and analyzed for their correlation with extRAAS genes in the microarray datasets. Expression data revealed the expressed extRAAS components and their relative abundance displaying the favored pathways in atheroma. Three co-expression modules with more than 80% reproducibility across datasets were extracted. Two of them (M1 and M2) contained genes coding for angiotensin metabolizing enzymes involved in different pathways: M1 included ACE, MME, RNPEP, and DPP3, in addition to 7 other genes; and M2 included CMA1, CTSG, and CPA3. The third module (M3) contained genes coding for receptors known to be implicated in atheroma (AGTR1, MR, GR, LNPEP, EGFR and GPER). M1 and M3 were negatively correlated in 3 of 5 datasets. We identified 19 TFs that have enriched TFBSs in the promoters of genes of M1, and two for M3, but none was found for M2. Among the extracted TFs, ELF1, MAX, and IRF5 showed significant positive correlations with peptidase-coding genes from M1 and negative correlations with receptors-coding genes from M3 (p < 0.05). The identified co-expression modules display the transcriptional organization of local extRAAS in human carotid atheroma. The identification of several TFs potentially associated to extRAAS genes may provide a frame for the discovery of atheroma-specific modulators of extRAAS activity.(Figure is included in full-text article.).

A resource for characterizing genome-wide binding and putative target genes of transcription factors expressed during secondary growth and wood formation in Populus.

PubMed

Liu, Lijun; Ramsay, Trevor; Zinkgraf, Matthew; Sundell, David; Street, Nathaniel Robert; Filkov, Vladimir; Groover, Andrew

2015-06-01

Identifying transcription factor target genes is essential for modeling the transcriptional networks underlying developmental processes. Here we report a chromatin immunoprecipitation sequencing (ChIP-seq) resource consisting of genome-wide binding regions and associated putative target genes for four Populus homeodomain transcription factors expressed during secondary growth and wood formation. Software code (programs and scripts) for processing the Populus ChIP-seq data are provided within a publically available iPlant image, including tools for ChIP-seq data quality control and evaluation adapted from the human Encyclopedia of DNA Elements (ENCODE) project. Basic information for each transcription factor (including members of Class I KNOX, Class III HD ZIP, BEL1-like families) binding are summarized, including the number and location of binding regions, distribution of binding regions relative to gene features, associated putative target genes, and enriched functional categories of putative target genes. These ChIP-seq data have been integrated within the Populus Genome Integrative Explorer (PopGenIE) where they can be analyzed using a variety of web-based tools. We present an example analysis that shows preferential binding of transcription factor ARBORKNOX1 to the nearest neighbor genes in a pre-calculated co-expression network module, and enrichment for meristem-related genes within this module including multiple orthologs of Arabidopsis KNOTTED-like Arabidopsis 2/6. © 2015 Society for Experimental Biology and John Wiley & Sons Ltd This article has been contributed to by US Government employees and their work is in the public domain in the USA.

Synergistic binding of bHLH transcription factors to the promoter of the maize NADP-ME gene used in C4 photosynthesis is based on an ancient code found in the ancestral C3 state.

PubMed

Borba, Ana Rita; Serra, Tânia S; Górska, Alicja; Gouveia, Paulo; Cordeiro, André M; Reyna-Llorens, Ivan; Knerová, Jana; Barros, Pedro M; Abreu, Isabel A; Oliveira, M Margarida; Hibberd, Julian M; Saibo, Nelson J M

2018-04-05

C4 photosynthesis has evolved repeatedly from the ancestral C3 state to generate a carbon concentrating mechanism that increases photosynthetic efficiency. This specialised form of photosynthesis is particularly common in the PACMAD clade of grasses, and is used by many of the world's most productive crops. The C4 cycle is accomplished through cell-type specific accumulation of enzymes but cis-elements and transcription factors controlling C4 photosynthesis remain largely unknown. Using the NADP-Malic Enzyme (NADP-ME) gene as a model we tested whether mechanisms impacting on transcription in C4 plants evolved from ancestral components found in C3 species. Two basic Helix-Loop-Helix (bHLH) transcription factors, ZmbHLH128 and ZmbHLH129, were shown to bind the C4NADP-ME promoter from maize. These proteins form heterodimers and ZmbHLH129 impairs trans-activation by ZmbHLH128. Electrophoretic mobility shift assays indicate that a pair of cis-elements separated by a seven base pair spacer synergistically bind either ZmbHLH128 or ZmbHLH129. This pair of cis-elements is found in both C3 and C4 Panicoid grass species of the PACMAD clade. Our analysis is consistent with this cis-element pair originating from a single motif present in the ancestral C3 state. We conclude that C4 photosynthesis has co-opted an ancient C3 regulatory code built on G-box recognition by bHLH to regulate the NADP-ME gene. More broadly, our findings also contribute to the understanding of gene regulatory networks controlling C4 photosynthesis.

lncRNA Structure: Message to the Heart.

PubMed

Fazal, Furqan M; Chang, Howard Y

2016-10-06

In this issue, Xue et al. (2016) describe the secondary structure of the heart-specific long non-coding RNA Braveheart, leading to the discovery of a short, asymmetric G-rich loop that controls cardiac lineage commitment by interacting with the transcription factor CNBP. Copyright © 2016 Elsevier Inc. All rights reserved.

Growth Cone Localization of the mRNA Encoding the Chromatin Regulator HMGN5 Modulates Neurite Outgrowth

PubMed Central

Moretti, Francesca; Rolando, Chiara; Winker, Moritz; Ivanek, Robert; Rodriguez, Javier; Von Kriegsheim, Alex; Taylor, Verdon; Bustin, Michael

2015-01-01

Neurons exploit local mRNA translation and retrograde transport of transcription factors to regulate gene expression in response to signaling events at distal neuronal ends. Whether epigenetic factors could also be involved in such regulation is not known. We report that the mRNA encoding the high-mobility group N5 (HMGN5) chromatin binding protein localizes to growth cones of both neuron-like cells and of hippocampal neurons, where it has the potential to be translated, and that HMGN5 can be retrogradely transported into the nucleus along neurites. Loss of HMGN5 function induces transcriptional changes and impairs neurite outgrowth, while HMGN5 overexpression induces neurite outgrowth and chromatin decompaction; these effects are dependent on growth cone localization of Hmgn5 mRNA. We suggest that the localization and local translation of transcripts coding for epigenetic factors couple the dynamic neuronal outgrowth process with chromatin regulation in the nucleus. PMID:25825524

Acetyl-L-carnitine supplementation to old rats partially reverts the age-related mitochondrial decay of soleus muscle by activating peroxisome proliferator-activated receptor gamma coactivator-1alpha-dependent mitochondrial biogenesis.

PubMed

Pesce, Vito; Fracasso, Flavio; Cassano, Pierluigi; Lezza, Angela Maria Serena; Cantatore, Palmiro; Gadaleta, Maria Nicola

2010-01-01

The age-related decay of mitochondrial function is a major contributor to the aging process. We tested the effects of 2-month-daily acetyl-L-carnitine (ALCAR) supplementation on mitochondrial biogenesis in the soleus muscle of aged rats. This muscle is heavily dependent on oxidative metabolism. Mitochondrial (mt) DNA content, citrate synthase activity, transcript levels of some nuclear- and mitochondrial-coded genes (cytochrome c oxidase subunit IV [COX-IV], 16S rRNA, COX-I) and of some factors involved in the mitochondrial biogenesis signaling pathway (peroxisome proliferator-activated receptor gamma [PPARgamma] coactivator-1alpha [PGC-1alpha], mitochondrial transcription factor A mitochondrial [TFAM], mitochondrial transcription factor 2B [TFB2]), as well as the protein content of PGC-1alpha were determined. The results suggest that the ALCAR treatment in old rats activates PGC-1alpha-dependent mitochondrial biogenesis, thus partially reverting the age-related mitochondrial decay.

Yeast one-hybrid system used to identify the binding proteins for rat glutathione S-transferase P enhancer I.

PubMed

Liao, Ming-Xiang; Liu, Dong-Yuan; Zuo, Jin; Fang, Fu-De

2002-03-01

To detect the trans-factors specifically binding to the strong enhancer element (GPEI) in the upstream of rat glutathione S-transferase P (GST-P) gene. Yeast one-hybrid system was used to screen rat lung MATCHMAKER cDNA library to identify potential trans-factors that can interact with core sequence of GPEI(cGPEI). Electrophoresis mobility shift assay (EMSA) was used to analyze the binding of transfactors to cGPEI. cDNA fragments coding for the C-terminal part of the transcription factor c-Jun and rat adenine nucleotide translocator (ANT) were isolated. The binding of c-Jun and ANT to GPEI core sequence were confirmed. Rat c-jun transcriptional factor and ANT may interact with cGPEI. They could play an important role in the induced expression of GST-P gene.

Gene-specific cell labeling using MiMIC transposons.

PubMed

Gnerer, Joshua P; Venken, Koen J T; Dierick, Herman A

2015-04-30

Binary expression systems such as GAL4/UAS, LexA/LexAop and QF/QUAS have greatly enhanced the power of Drosophila as a model organism by allowing spatio-temporal manipulation of gene function as well as cell and neural circuit function. Tissue-specific expression of these heterologous transcription factors relies on random transposon integration near enhancers or promoters that drive the binary transcription factor embedded in the transposon. Alternatively, gene-specific promoter elements are directly fused to the binary factor within the transposon followed by random or site-specific integration. However, such insertions do not consistently recapitulate endogenous expression. We used Minos-Mediated Integration Cassette (MiMIC) transposons to convert host loci into reliable gene-specific binary effectors. MiMIC transposons allow recombinase-mediated cassette exchange to modify the transposon content. We developed novel exchange cassettes to convert coding intronic MiMIC insertions into gene-specific binary factor protein-traps. In addition, we expanded the set of binary factor exchange cassettes available for non-coding intronic MiMIC insertions. We show that binary factor conversions of different insertions in the same locus have indistinguishable expression patterns, suggesting that they reliably reflect endogenous gene expression. We show the efficacy and broad applicability of these new tools by dissecting the cellular expression patterns of the Drosophila serotonin receptor gene family. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

The electrostatic role of the Zn-Cys2His2 complex in binding of operator DNA with transcription factors: mouse EGR-1 from the Cys2His2 family.

PubMed

Chirgadze, Y N; Boshkova, E A; Polozov, R V; Sivozhelezov, V S; Dzyabchenko, A V; Kuzminsky, M B; Stepanenko, V A; Ivanov, V V

2018-01-07

The mouse factor Zif268, known also as early growth response protein EGR-1, is a classical representative for the Cys2His2 transcription factor family. It is required for binding the RNA polymerase with operator dsDNA to initialize the transcription process. We have shown that only in this family of total six Zn-finger protein families the Zn complex plays a significant role in the protein-DNA binding. Electrostatic feature of this complex in the binding of factor Zif268 from Mus musculus with operator DNA has been considered. The factor consists of three similar Zn-finger units which bind with triplets of coding DNA. Essential contacts of the factor with the DNA phosphates are formed by three conservative His residues, one in each finger. We describe here the results of calculations of the electrostatic potentials for the Zn-Cys2His2 complex, Zn-finger unit 1, and the whole transcription factor. The potential of Zif268 has a positive area on the factor surface, and it corresponds exactly to the binding sites of each of Zn-finger units. The main part of these areas is determined by conservative His residues, which form contacts with the DNA phosphate groups. Our result shows that the electrostatic positive potential of this histidine residue is enhanced due to the Zn complex. The other contacts of the Zn-finger with DNA are related to nucleotide bases, and they are responsible for the sequence-specific binding with DNA. This result may be extended to all other members of the Cys2His2 transcription factor family.

A transcription factor collective defines the HSN serotonergic neuron regulatory landscape

PubMed Central

Artacho, Alejandro; Jimeno-Martín, Ángela; Chirivella, Laura; Weinberg, Peter

2018-01-01

Cell differentiation is controlled by individual transcription factors (TFs) that together activate a selection of enhancers in specific cell types. How these combinations of TFs identify and activate their target sequences remains poorly understood. Here, we identify the cis-regulatory transcriptional code that controls the differentiation of serotonergic HSN neurons in Caenorhabditis elegans. Activation of the HSN transcriptome is directly orchestrated by a collective of six TFs. Binding site clusters for this TF collective form a regulatory signature that is sufficient for de novo identification of HSN neuron functional enhancers. Among C. elegans neurons, the HSN transcriptome most closely resembles that of mouse serotonergic neurons. Mouse orthologs of the HSN TF collective also regulate serotonergic differentiation and can functionally substitute for their worm counterparts which suggests deep homology. Our results identify rules governing the regulatory landscape of a critically important neuronal type in two species separated by over 700 million years. PMID:29553368

Unraveling transcriptional control and cis-regulatory codes using the software suite GeneACT

PubMed Central

Cheung, Tom Hiu; Kwan, Yin Lam; Hamady, Micah; Liu, Xuedong

2006-01-01

Deciphering gene regulatory networks requires the systematic identification of functional cis-acting regulatory elements. We present a suite of web-based bioinformatics tools, called GeneACT , that can rapidly detect evolutionarily conserved transcription factor binding sites or microRNA target sites that are either unique or over-represented in differentially expressed genes from DNA microarray data. GeneACT provides graphic visualization and extraction of common regulatory sequence elements in the promoters and 3'-untranslated regions that are conserved across multiple mammalian species. PMID:17064417

Transcription factors network in root endosymbiosis establishment and development.

PubMed

Diédhiou, Issa; Diouf, Diaga

2018-02-15

Root endosymbioses are mutualistic interactions between plants and the soil microorganisms (Fungus, Frankia or Rhizobium) that lead to the formation of nitrogen-fixing root nodules and/or arbuscular mycorrhiza. These interactions enable many species to survive in different marginal lands to overcome the nitrogen-and/or phosphorus deficient environment and can potentially reduce the chemical fertilizers used in agriculture which gives them an economic, social and environmental importance. The formation and the development of these structures require the mediation of specific gene products among which the transcription factors play a key role. Three of these transcription factors, viz., CYCLOPS, NSP1 and NSP2 are well conserved between actinorhizal, legume, non-legume and mycorrhizal symbioses. They interact with DELLA proteins to induce the expression of NIN in nitrogen fixing symbiosis or RAM1 in mycorrhizal symbiosis. Recently, the small non coding RNA including micro RNAs (miRNAs) have emerged as major regulators of root endosymbioses. Among them, miRNA171 targets NSP2, a TF conserved in actinorhizal, legume, non-legume and mycorrhizal symbioses. This review will also focus on the recent advances carried out on the biological function of others transcription factors during the root pre-infection/pre-contact, infection or colonization. Their role in nodule formation and AM development will also be described.

Reflective Practice in the Clinical Setting: A Multi-Institutional Qualitative Study of Pediatric Faculty and Residents.

PubMed

Plant, Jennifer; Li, Su-Ting T; Blankenburg, Rebecca; Bogetz, Alyssa L; Long, Michele; Butani, Lavjay

2017-11-01

To explore when and in what form pediatric faculty and residents practice reflection. From February to June 2015, the authors conducted focus groups of pediatric faculty and residents at the University of California, Davis; Stanford University; and the University of California, San Francisco, until thematic saturation occurred. Transcripts were analyzed based on Mezirow's and Schon's models of reflection, using the constant comparative method associated with grounded theory. Two investigators independently coded transcripts and reconciled codes to develop themes. All investigators reviewed the codes and developed a final list of themes through consensus. Through iterative discussions, investigators developed a conceptual model of reflection in the clinical setting. Seventeen faculty and 20 residents from three institutions participated in six focus groups. Five themes emerged: triggers of reflection, intrinsic factors, extrinsic factors, timing, and outcome of reflection. Various triggers led to reflection; whether a specific trigger led to reflection depended on intrinsic and extrinsic factors. When reflection occurred, it happened in action or on action. Under optimal conditions, this reflection was goal and action directed and became critical reflection. In other instances, this process resulted in unproductive rumination or acted as an emotional release or supportive therapy. Participants reflected in clinical settings, but did not always explicitly identify it as reflection or reflect in growth-promoting ways. Strategies to enhance critical reflection include developing knowledge and skills in reflection, providing performance data to inform reflection, creating time and space for safe reflection, and providing mentorship to guide the process.

My baby, my move: examination of perceived barriers and motivating factors related to antenatal physical activity.

PubMed

Leiferman, Jenn; Swibas, Tracy; Koiness, Kacey; Marshall, Julie A; Dunn, Andrea L

2011-01-01

Based on a socioecological model, the present study examined multilevel barriers and facilitators related to physical activity engagement during pregnancy in women of low socioeconomic status. Individual and paired interviews were conducted with 25 pregnant women (aged 18-46 years, 17-40 weeks' gestation) to ask about motivational factors and to compare differences in activity level and parity. Atlas/Ti software was used to code verbatim interview transcripts by organizing codes into categories that reflect symbolic domains of meaning, relational patterns, and overarching themes. Perceived barriers and motivating factors differed between exercisers and nonexercisers at intrapersonal, interpersonal, and environmental levels. Future interventions should take into account key motivating multilevel factors and barriers to tailor more meaningful advice for pregnant women. © 2011 by the American College of Nurse-Midwives.

What Limits the Effectiveness of Antibullying Programs? A Thematic Analysis of the Perspective of Teachers

ERIC Educational Resources Information Center

Cunningham, Charles E.; Rimas, Heather; Mielko, Stephanie; Mapp, Cailin; Cunningham, Lesley; Buchanan, Don; Vaillancourt, Tracy; Chen, Yvonne; Deal, Ken; Marcus, Madalyn

2016-01-01

Prevention programs yield modest reductions in bullying in North American schools. This study explored the perspective of educators regarding factors limiting the impact of these initiatives. Transcripts from nineteen 90-min focus groups with 103 educators were coded thematically. Educators felt that off-site incidents, cyberbullying, and the…

Opposite GC skews at the 5' and 3' ends of genes in unicellular fungi

PubMed Central

2011-01-01

Background GC-skews have previously been linked to transcription in some eukaryotes. They have been associated with transcription start sites, with the coding strand G-biased in mammals and C-biased in fungi and invertebrates. Results We show a consistent and highly significant pattern of GC-skew within genes of almost all unicellular fungi. The pattern of GC-skew is asymmetrical: the coding strand of genes is typically C-biased at the 5' ends but G-biased at the 3' ends, with intermediate skews at the middle of genes. Thus, the initiation, elongation, and termination phases of transcription are associated with different skews. This pattern influences the encoded proteins by generating differential usage of amino acids at the 5' and 3' ends of genes. These biases also affect fourfold-degenerate positions and extend into promoters and 3' UTRs, indicating that skews cannot be accounted by selection for protein function or translation. Conclusions We propose two explanations, the mutational pressure hypothesis, and the adaptive hypothesis. The mutational pressure hypothesis is that different co-factors bind to RNA pol II at different phases of transcription, producing different mutational regimes. The adaptive hypothesis is that cytidine triphosphate deficiency may lead to C-avoidance at the 3' ends of transcripts to control the flow of RNA pol II molecules and reduce their frequency of collisions. PMID:22208287

Post-transcriptional regulatory network of epithelial-to-mesenchymal and mesenchymal-to-epithelial transitions

PubMed Central

2014-01-01

Epithelial-to-mesenchymal transition (EMT) and its reverse process, mesenchymal-to-epithelial transition (MET), play important roles in embryogenesis, stem cell biology, and cancer progression. EMT can be regulated by many signaling pathways and regulatory transcriptional networks. Furthermore, post-transcriptional regulatory networks regulate EMT; these networks include the long non-coding RNA (lncRNA) and microRNA (miRNA) families. Specifically, the miR-200 family, miR-101, miR-506, and several lncRNAs have been found to regulate EMT. Recent studies have illustrated that several lncRNAs are overexpressed in various cancers and that they can promote tumor metastasis by inducing EMT. MiRNA controls EMT by regulating EMT transcription factors or other EMT regulators, suggesting that lncRNAs and miRNA are novel therapeutic targets for the treatment of cancer. Further efforts have shown that non-coding-mediated EMT regulation is closely associated with epigenetic regulation through promoter methylation (e.g., miR-200 or miR-506) and protein regulation (e.g., SET8 via miR-502). The formation of gene fusions has also been found to promote EMT in prostate cancer. In this review, we discuss the post-transcriptional regulatory network that is involved in EMT and MET and how targeting EMT and MET may provide effective therapeutics for human disease. PMID:24598126

C/EBPβ contributes to transcriptional activation of long non-coding RNA NEAT1 during APL cell differentiation.

PubMed

Wang, Yewei; Fu, Lei; Sun, Ailian; Tang, Doudou; Xu, Yunxiao; Li, Zheyuan; Chen, Mingjie; Zhang, Guangsen

2018-05-05

Emerging evidences have shown that long non-coding RNAs (lncRNAs) play critical roles in cancer development and cancer therapy. LncRNA Nuclear Enriched Abundant Transcript 1 (NEAT1) is indispensable during acute promyelocytic leukemia (APL) cell differentiation induced by all-trans retinoic acid (ATRA). However, the precise mechanism of NEAT1 upregulation has not been fully understood. In this study, we performed chromatin immunoprecipitation and luciferase reporter assays to demonstrate that C/EBP family transcription factor C/EBPβ bind to and transactivate the promoter of lncRNA NEAT1 through the C/EBPβ binding sites both around -54 bp and -1453 bp upstream of the transcription start site. Moreover, the expression of C/EBPβ was increased after ATRA treatment, and the binding of C/EBPβ in the NEAT1 promoter was also dramatically increased. Finally, knockdown of C/EBPβ significantly reduced the ATRA-induced upregulation of NEAT1. In conclusion, C/EBPβ directly activates the expression of NEAT1 through binding to the promoter of NEAT1. Knockdown of C/EBPβ impairs ATRA-induced transcriptional activation of NEAT1. Our data indicate that C/EBPβ contributes to ATRA-induced activation of NEAT1 during APL cell differentiation. Our results enrich our knowledge on the regulation of lncRNAs and the regulatory role of C/EBPβ in APL cell differentiation. Copyright © 2017. Published by Elsevier Inc.

Genome-wide uniformity of human ‘open’ pre-initiation complexes

PubMed Central

Lai, William K.M.; Pugh, B. Franklin

2017-01-01

Transcription of protein-coding and noncoding DNA occurs pervasively throughout the mammalian genome. Their sites of initiation are generally inferred from transcript 5′ ends and are thought to be either locally dispersed or focused. How these two modes of initiation relate is unclear. Here, we apply permanganate treatment and chromatin immunoprecipitation (PIP-seq) of initiation factors to identify the precise location of melted DNA separately associated with the preinitiation complex (PIC) and the adjacent paused complex (PC). This approach revealed the two known modes of transcription initiation. However, in contrast to prevailing views, they co-occurred within the same promoter region: initiation originating from a focused PIC, and broad nucleosome-linked initiation. PIP-seq allowed transcriptional orientation of Pol II to be determined, which may be useful near promoters where sufficient sense/anti-sense transcript mapping information is lacking. PIP-seq detected divergently oriented Pol II at both coding and noncoding promoters, as well as at enhancers. Their occupancy levels were not necessarily coupled in the two orientations. DNA sequence and shape analysis of initiation complex sites suggest that both sequence and shape contribute to specificity, but in a context-restricted manner. That is, initiation sites have the locally “best” initiator (INR) sequence and/or shape. These findings reveal a common core to pervasive Pol II initiation throughout the human genome. PMID:27927716

Characterization and transcription of arsenic respiration and resistance genes during in situ uranium bioremediation

PubMed Central

Giloteaux, Ludovic; Holmes, Dawn E; Williams, Kenneth H; Wrighton, Kelly C; Wilkins, Michael J; Montgomery, Alison P; Smith, Jessica A; Orellana, Roberto; Thompson, Courtney A; Roper, Thomas J; Long, Philip E; Lovley, Derek R

2013-01-01

The possibility of arsenic release and the potential role of Geobacter in arsenic biogeochemistry during in situ uranium bioremediation was investigated because increased availability of organic matter has been associated with substantial releases of arsenic in other subsurface environments. In a field experiment conducted at the Rifle, CO study site, groundwater arsenic concentrations increased when acetate was added. The number of transcripts from arrA, which codes for the α-subunit of dissimilatory As(V) reductase, and acr3, which codes for the arsenic pump protein Acr3, were determined with quantitative reverse transcription-PCR. Most of the arrA (>60%) and acr3-1 (>90%) sequences that were recovered were most similar to Geobacter species, while the majority of acr3-2 (>50%) sequences were most closely related to Rhodoferax ferrireducens. Analysis of transcript abundance demonstrated that transcription of acr3-1 by the subsurface Geobacter community was correlated with arsenic concentrations in the groundwater. In contrast, Geobacter arrA transcript numbers lagged behind the major arsenic release and remained high even after arsenic concentrations declined. This suggested that factors other than As(V) availability regulated the transcription of arrA in situ, even though the presence of As(V) increased the transcription of arrA in cultures of Geobacter lovleyi, which was capable of As(V) reduction. These results demonstrate that subsurface Geobacter species can tightly regulate their physiological response to changes in groundwater arsenic concentrations. The transcriptomic approach developed here should be useful for the study of a diversity of other environments in which Geobacter species are considered to have an important influence on arsenic biogeochemistry. PMID:23038171

An RNA motif advances transcription by preventing Rho-dependent termination

PubMed Central

Sevostyanova, Anastasia; Groisman, Eduardo A.

2015-01-01

The transcription termination factor Rho associates with most nascent bacterial RNAs as they emerge from RNA polymerase. However, pharmacological inhibition of Rho derepresses only a small fraction of these transcripts. What, then, determines the specificity of Rho-dependent transcription termination? We now report the identification of a Rho-antagonizing RNA element (RARE) that hinders Rho-dependent transcription termination. We establish that RARE traps Rho in an inactive complex but does not prevent Rho binding to its recruitment sites. Although translating ribosomes normally block Rho access to an mRNA, inefficient translation of an open reading frame in the leader region of the Salmonella mgtCBR operon actually enables transcription of its associated coding region by favoring an RNA conformation that sequesters RARE. The discovery of an RNA element that inactivates Rho signifies that the specificity of nucleic-acid binding proteins is defined not only by the sequences that recruit these proteins but also by sequences that antagonize their activity. PMID:26630006

Intergenic Transcriptional Interference Is Blocked by RNA Polymerase III Transcription Factor TFIIIB in Saccharomyces cerevisiae

PubMed Central

Korde, Asawari; Rosselot, Jessica M.; Donze, David

2014-01-01

The major function of eukaryotic RNA polymerase III is to transcribe transfer RNA, 5S ribosomal RNA, and other small non-protein-coding RNA molecules. Assembly of the RNA polymerase III complex on chromosomal DNA requires the sequential binding of transcription factor complexes TFIIIC and TFIIIB. Recent evidence has suggested that in addition to producing RNA transcripts, chromatin-assembled RNA polymerase III complexes may mediate additional nuclear functions that include chromatin boundary, nucleosome phasing, and general genome organization activities. This study provides evidence of another such “extratranscriptional” activity of assembled RNA polymerase III complexes, which is the ability to block progression of intergenic RNA polymerase II transcription. We demonstrate that the RNA polymerase III complex bound to the tRNA gene upstream of the Saccharomyces cerevisiae ATG31 gene protects the ATG31 promoter against readthrough transcriptional interference from the upstream noncoding intergenic SUT467 transcription unit. This protection is predominately mediated by binding of the TFIIIB complex. When TFIIIB binding to this tRNA gene is weakened, an extended SUT467–ATG31 readthrough transcript is produced, resulting in compromised ATG31 translation. Since the ATG31 gene product is required for autophagy, strains expressing the readthrough transcript exhibit defective autophagy induction and reduced fitness under autophagy-inducing nitrogen starvation conditions. Given the recent discovery of widespread pervasive transcription in all forms of life, protection of neighboring genes from intergenic transcriptional interference may be a key extratranscriptional function of assembled RNA polymerase III complexes and possibly other DNA binding proteins. PMID:24336746

Role of Architecture in the Function and Specificity of Two Notch-Regulated Transcriptional Enhancer Modules

PubMed Central

Liu, Feng; Posakony, James W.

2012-01-01

In Drosophila melanogaster, cis-regulatory modules that are activated by the Notch cell–cell signaling pathway all contain two types of transcription factor binding sites: those for the pathway's transducing factor Suppressor of Hairless [Su(H)] and those for one or more tissue- or cell type–specific factors called “local activators.” The use of different “Su(H) plus local activator” motif combinations, or codes, is critical to ensure that only the correct subset of the broadly utilized Notch pathway's target genes are activated in each developmental context. However, much less is known about the role of enhancer “architecture”—the number, order, spacing, and orientation of its component transcription factor binding motifs—in determining the module's specificity. Here we investigate the relationship between architecture and function for two Notch-regulated enhancers with spatially distinct activities, each of which includes five high-affinity Su(H) sites. We find that the first, which is active specifically in the socket cells of external sensory organs, is largely resistant to perturbations of its architecture. By contrast, the second enhancer, active in the “non-SOP” cells of the proneural clusters from which neural precursors arise, is sensitive to even simple rearrangements of its transcription factor binding sites, responding with both loss of normal specificity and striking ectopic activity. Thus, diverse cryptic specificities can be inherent in an enhancer's particular combination of transcription factor binding motifs. We propose that for certain types of enhancer, architecture plays an essential role in determining specificity, not only by permitting factor–factor synergies necessary to generate the desired activity, but also by preventing other activator synergies that would otherwise lead to unwanted specificities. PMID:22792075

The gene coding for the B cell surface protein CD19 is localized on human chromosome 16p11.

PubMed

Stapleton, P; Kozmik, Z; Weith, A; Busslinger, M

1995-02-01

The CD19 gene codes for one of the earliest markers of the human B cell lineage and is a target for the B lymphoid-specific transcription factor BSAP (Pax-5). The transmembrane protein CD19 has been implicated in controlling proliferation of mature B lymphocytes by modulating signal transduction through the antigen receptor. In this study, we have employed Southern blot and fluorescence in situ hybridization analyses to localize the CD19 gene to human chromosome 16p11.

HSFs, Stress Sensors and Sculptors of Transcription Compartments and Epigenetic Landscapes.

PubMed

Miozzo, Federico; Sabéran-Djoneidi, Délara; Mezger, Valérie

2015-12-04

Starting as a paradigm for stress responses, the study of the transcription factor (TF) family of heat shock factors (HSFs) has quickly and widely expanded these last decades, thanks to their fascinating and significant involvement in a variety of pathophysiological processes, including development, reproduction, neurodegeneration and carcinogenesis. HSFs, originally defined as classical TFs, strikingly appeared to play a central and often pioneering role in reshaping the epigenetic landscape. In this review, we describe how HSFs are able to sense the epigenetic environment, and we review recent data that support their role as sculptors of the chromatin landscape through their complex interplay with chromatin remodelers, histone-modifying enzymes and non-coding RNAs. Copyright © 2015 Elsevier Ltd. All rights reserved.

Insights into inner ear-specific gene regulation: epigenetics and non-coding RNAs in inner ear development and regeneration

PubMed Central

Avraham, Karen B.

2016-01-01

The vertebrate inner ear houses highly specialized sensory organs, tuned to detect and encode sound, head motion and gravity. Gene expression programs under the control of transcription factors orchestrate the formation and specialization of the non-sensory inner ear labyrinth and its sensory constituents. More recently, epigenetic factors and non-coding RNAs emerged as an additional layer of gene regulation, both in inner ear development and disease. In this review, we provide an overview on how epigenetic modifications and non-coding RNAs, in particular microRNAs (miRNAs), influence gene expression and summarize recent discoveries that highlight their critical role in the proper formation of the inner ear labyrinth and its sensory organs. In contrast to non-mammalian vertebrates, adult mammals lack the ability to regenerate inner ear mechano-sensory hair cells. Finally, we discuss recent insights into how epigenetic factors and miRNAs may facilitate, or in the case of mammals, restrict sensory hair cell regeneration. PMID:27836639

Resveratrol protects primary rat hepatocytes against oxidative stress damage: activation of the Nrf2 transcription factor and augmented activities of antioxidant enzymes.

PubMed

Rubiolo, Juan Andrés; Mithieux, Gilles; Vega, Félix Victor

2008-09-04

Oxidative stress is recognized as an important factor in the development of liver pathologies. The reactive oxygen species endogenously generated or as a consequence of xenobiotic metabolism are eliminated by enzymatic and nonenzymatic cellular systems. Besides endogen defences, the antioxidant consumption in the diet has an important role in the protection against the development of diseases product of oxidative damage. Resveratrol is a naturally occurring compound which is part of the human diet. This molecule has been shown to have many biological properties, including antioxidant activity. We decided to test if resveratrol could protect primary hepatocytes in culture from oxidative stress damage and if so, to determine if this compound affects the cellular detoxifying systems and their regulation through the Nrf2 transcription factor that regulates the expression of antioxidant and phase II detoxifying enzymes. Cell death by necrosis was detected by measuring the activity of lactate dehydrogenase liberated to the medium. The activities of antioxidant and phase II enzymes were measured using previously described methods. Activation of the Nrf2 transcription factor was studied by confocal microscopy and the Nrf2 and its coding mRNA levels were determined by western blot and quantitative PCR respectively. Resveratrol pre-treatment effectively protected hepatocytes in culture exposed to oxidative stress, increasing the activities of catalase, superoxide dismutase, glutathione peroxidase, NADPH quinone oxidoreductase and glutathione-S-transferase. Resveratrol increases the level of Nrf2 and induces its translocation to the nucleus. Also, it increases the concentration of the coding mRNA for Nrf2. In this work we show that resveratrol could be a useful drug for the protection of liver cells from oxidative stress induced damage.

Cross-site comparison of ribosomal depletion kits for Illumina RNAseq library construction.

PubMed

Herbert, Zachary T; Kershner, Jamie P; Butty, Vincent L; Thimmapuram, Jyothi; Choudhari, Sulbha; Alekseyev, Yuriy O; Fan, Jun; Podnar, Jessica W; Wilcox, Edward; Gipson, Jenny; Gillaspy, Allison; Jepsen, Kristen; BonDurant, Sandra Splinter; Morris, Krystalynne; Berkeley, Maura; LeClerc, Ashley; Simpson, Stephen D; Sommerville, Gary; Grimmett, Leslie; Adams, Marie; Levine, Stuart S

2018-03-15

Ribosomal RNA (rRNA) comprises at least 90% of total RNA extracted from mammalian tissue or cell line samples. Informative transcriptional profiling using massively parallel sequencing technologies requires either enrichment of mature poly-adenylated transcripts or targeted depletion of the rRNA fraction. The latter method is of particular interest because it is compatible with degraded samples such as those extracted from FFPE and also captures transcripts that are not poly-adenylated such as some non-coding RNAs. Here we provide a cross-site study that evaluates the performance of ribosomal RNA removal kits from Illumina, Takara/Clontech, Kapa Biosystems, Lexogen, New England Biolabs and Qiagen on intact and degraded RNA samples. We find that all of the kits are capable of performing significant ribosomal depletion, though there are differences in their ease of use. All kits were able to remove ribosomal RNA to below 20% with intact RNA and identify ~ 14,000 protein coding genes from the Universal Human Reference RNA sample at >1FPKM. Analysis of differentially detected genes between kits suggests that transcript length may be a key factor in library production efficiency. These results provide a roadmap for labs on the strengths of each of these methods and how best to utilize them.

Reading of the non-template DNA by transcription elongation factors.

PubMed

Svetlov, Vladimir; Nudler, Evgeny

2018-05-14

Unlike transcription initiation and termination, which have easily discernable signals such as promoters and terminators, elongation is regulated through a dynamic network involving RNA/DNA pause signals and states- rather than sequence-specific protein interactions. A report by Nedialkov et al. (in press) provides experimental evidence for sequence-specific recruitment of elongation factor RfaH to transcribing RNA polymerase (RNAP) and outlines the mechanism of gene expression regulation by restraint ("locking") of the DNA non-template strand. According to this model, the elongation complex pauses at the so called "operon polarity sequence" (found in some long bacterial operons coding for virulence genes), when the usually flexible non-template DNA strand adopts a distinct hairpin-loop conformation on the surface of transcribing RNAP. Sequence-specific binding of RfaH to this DNA segment facilitates conversion of RfaH from its inactive closed to its active open conformation. The interaction network formed between RfaH, non-template DNA, and RNAP locks DNA in a conformation that renders the elongation complex resistant to pausing and termination. The effects of such locking on transcript elongation can be mimicked by restraint of the non-template strand due to its shortening. This work advances our understanding of regulation of transcript elongation and has important implications for the action of general transcription factors, such as NusG, which lack apparent sequence-specificity, as well as for the mechanisms of other processes linked to transcription such as transcription-coupled DNA repair. This article is protected by copyright. All rights reserved. © 2018 John Wiley & Sons Ltd.

Utrophin Up-Regulation by an Artificial Transcription Factor in Transgenic Mice

PubMed Central

Mattei, Elisabetta; Corbi, Nicoletta; Di Certo, Maria Grazia; Strimpakos, Georgios; Severini, Cinzia; Onori, Annalisa; Desantis, Agata; Libri, Valentina; Buontempo, Serena; Floridi, Aristide; Fanciulli, Maurizio; Baban, Dilair; Davies, Kay E.; Passananti, Claudio

2007-01-01

Duchenne Muscular Dystrophy (DMD) is a severe muscle degenerative disease, due to absence of dystrophin. There is currently no effective treatment for DMD. Our aim is to up-regulate the expression level of the dystrophin related gene utrophin in DMD, complementing in this way the lack of dystrophin functions. To this end we designed and engineered several synthetic zinc finger based transcription factors. In particular, we have previously shown that the artificial three zinc finger protein named Jazz, fused with the appropriate effector domain, is able to drive the transcription of a test gene from the utrophin promoter “A”. Here we report on the characterization of Vp16-Jazz-transgenic mice that specifically over-express the utrophin gene at the muscular level. A Chromatin Immunoprecipitation assay (ChIP) demonstrated the effective access/binding of the Jazz protein to active chromatin in mouse muscle and Vp16-Jazz was shown to be able to up-regulate endogenous utrophin gene expression by immunohistochemistry, western blot analyses and real-time PCR. To our knowledge, this is the first example of a transgenic mouse expressing an artificial gene coding for a zinc finger based transcription factor. The achievement of Vp16-Jazz transgenic mice validates the strategy of transcriptional targeting of endogenous genes and could represent an exclusive animal model for use in drug discovery and therapeutics. PMID:17712422

Caste- and development-associated gene expression in a lower termite

PubMed Central

Scharf, Michael E; Wu-Scharf, Dancia; Pittendrigh, Barry R; Bennett, Gary W

2003-01-01

Background Social insects such as termites express dramatic polyphenism (the occurrence of multiple forms in a species on the basis of differential gene expression) both in association with caste differentiation and between castes after differentiation. We have used cDNA macroarrays to compare gene expression between polyphenic castes and intermediary developmental stages of the termite Reticulitermes flavipes. Results We identified differentially expressed genes from nine ontogenic categories. Quantitative PCR was used to quantify precise differences in gene expression between castes and between intermediary developmental stages. We found worker and nymph-biased expression of transcripts encoding termite and endosymbiont cellulases; presoldier-biased expression of transcripts encoding the storage/hormone-binding protein vitellogenin; and soldier-biased expression of gene transcripts encoding two transcription/translation factors, two signal transduction factors and four cytoskeletal/muscle proteins. The two transcription/translation factors showed significant homology to the bicaudal and bric-a-brac developmental genes of Drosophila. Conclusions Our results show differential expression of regulatory, structural and enzyme-coding genes in association with termite castes and their developmental precursor stages. They also provide the first glimpse into how insect endosymbiont cellulase gene expression can vary in association with the caste of a host. These findings shed light on molecular processes associated with termite biology, polyphenism, caste differentiation and development and highlight potentially interesting variations in developmental themes between termites, other insects, and higher animals. PMID:14519197

Biological significance of long non-coding RNA FTX expression in human colorectal cancer.

PubMed

Guo, Xiao-Bo; Hua, Zhu; Li, Chen; Peng, Li-Pan; Wang, Jing-Shen; Wang, Bo; Zhi, Qiao-Ming

2015-01-01

The purpose of this study was to determine the expression of long non-coding RNA (lncRNA) FTX and analyze its prognostic and biological significance in colorectal cancer (CRC). A quantitative reverse transcription PCR was performed to detect the expression of long non-coding RNA FTX in 35 pairs of colorectal cancer and corresponding noncancerous tissues. The expression of long non-coding RNA FTX was detected in 187 colorectal cancer tissues and its correlations with clinicopathological factors of patients were examined. Univariate and multivariate analyses were performed to analyze the prognostic significance of Long Non-coding RNA FTX expression. The effects of long non-coding RNA FTX expression on malignant phenotypes of colorectal cancer cells and its possible biological significances were further determined. Long non-coding RNA FTX was significantly upregulated in colorectal cancer tissues, and low long non-coding RNA FTX expression was significantly correlated with differentiation grade, lymph vascular invasion, and clinical stage. Patients with high long non-coding RNA FTX showed poorer overall survival than those with low long non-coding RNA FTX. Multivariate analyses indicated that status of long non-coding RNA FTX was an independent prognostic factor for patients. Functional analyses showed that upregulation of long non-coding RNA FTX significantly promoted growth, migration, invasion, and increased colony formation in colorectal cancer cells. Therefore, long non-coding RNA FTX may be a potential biomarker for predicting the survival of colorectal cancer patients and might be a molecular target for treatment of human colorectal cancer.

A class of circadian long non-coding RNAs mark enhancers modulating long-range circadian gene regulation

PubMed Central

Fan, Zenghua; Zhao, Meng; Joshi, Parth D.; Li, Ping; Zhang, Yan; Guo, Weimin; Xu, Yichi; Wang, Haifang; Zhao, Zhihu

2017-01-01

Abstract Circadian rhythm exerts its influence on animal physiology and behavior by regulating gene expression at various levels. Here we systematically explored circadian long non-coding RNAs (lncRNAs) in mouse liver and examined their circadian regulation. We found that a significant proportion of circadian lncRNAs are expressed at enhancer regions, mostly bound by two key circadian transcription factors, BMAL1 and REV-ERBα. These circadian lncRNAs showed similar circadian phases with their nearby genes. The extent of their nuclear localization is higher than protein coding genes but less than enhancer RNAs. The association between enhancer and circadian lncRNAs is also observed in tissues other than liver. Comparative analysis between mouse and rat circadian liver transcriptomes showed that circadian transcription at lncRNA loci tends to be conserved despite of low sequence conservation of lncRNAs. One such circadian lncRNA termed lnc-Crot led us to identify a super-enhancer region interacting with a cluster of genes involved in circadian regulation of metabolism through long-range interactions. Further experiments showed that lnc-Crot locus has enhancer function independent of lnc-Crot's transcription. Our results suggest that the enhancer-associated circadian lncRNAs mark the genomic loci modulating long-range circadian gene regulation and shed new lights on the evolutionary origin of lncRNAs. PMID:28335007

Genomic positional conservation identifies topological anchor point RNAs linked to developmental loci.

PubMed

Amaral, Paulo P; Leonardi, Tommaso; Han, Namshik; Viré, Emmanuelle; Gascoigne, Dennis K; Arias-Carrasco, Raúl; Büscher, Magdalena; Pandolfini, Luca; Zhang, Anda; Pluchino, Stefano; Maracaja-Coutinho, Vinicius; Nakaya, Helder I; Hemberg, Martin; Shiekhattar, Ramin; Enright, Anton J; Kouzarides, Tony

2018-03-15

The mammalian genome is transcribed into large numbers of long noncoding RNAs (lncRNAs), but the definition of functional lncRNA groups has proven difficult, partly due to their low sequence conservation and lack of identified shared properties. Here we consider promoter conservation and positional conservation as indicators of functional commonality. We identify 665 conserved lncRNA promoters in mouse and human that are preserved in genomic position relative to orthologous coding genes. These positionally conserved lncRNA genes are primarily associated with developmental transcription factor loci with which they are coexpressed in a tissue-specific manner. Over half of positionally conserved RNAs in this set are linked to chromatin organization structures, overlapping binding sites for the CTCF chromatin organiser and located at chromatin loop anchor points and borders of topologically associating domains (TADs). We define these RNAs as topological anchor point RNAs (tapRNAs). Characterization of these noncoding RNAs and their associated coding genes shows that they are functionally connected: they regulate each other's expression and influence the metastatic phenotype of cancer cells in vitro in a similar fashion. Furthermore, we find that tapRNAs contain conserved sequence domains that are enriched in motifs for zinc finger domain-containing RNA-binding proteins and transcription factors, whose binding sites are found mutated in cancers. This work leverages positional conservation to identify lncRNAs with potential importance in genome organization, development and disease. The evidence that many developmental transcription factors are physically and functionally connected to lncRNAs represents an exciting stepping-stone to further our understanding of genome regulation.

An Ultraconserved Brain-specific Enhancer within ADGRL3 (LPHN3) Underpins ADHD Susceptibility

PubMed Central

Martinez, Ariel F.; Abe, Yu; Hong, Sungkook; Molyneux, Kevin; Yarnell, David; Löhr, Heiko; Driever, Wolfgang; Acosta, Maria T.; Arcos-Burgos, Mauricio; Muenke, Maximilian

2016-01-01

BACKGROUND Genetic factors predispose to attention deficit/hyperactivity disorder (ADHD). Previous studies have reported linkage and association to ADHD of gene variants within ADGRL3. In this study, we functionally analyzed non-coding variants in this gene as likely pathological contributors. METHODS In silico, in vitro and in vivo approaches were used to identify and characterize evolutionary conserved elements within the ADGRL3 linkage region (~207 Kb). Family-based genetic analyses on 838 individuals (372 affected and 466 unaffected) identified ADHD-associated SNPs harbored in some of these conserved elements. Luciferase assays and zebrafish GFP transgenesis tested conserved elements for transcriptional enhancer activity. Electromobility shift assays were used to verify transcription factor binding disruption by ADHD risk alleles. RESULTS An ultraconserved element was discovered (ECR47) that functions as a transcriptional enhancer. A three-variant ADHD risk haplotype in ECR47, formed by rs17226398, rs56038622 and rs2271338, reduced enhancer activity by 40% in neuroblastoma and astrocytoma cells (PBonferroni<0.0001). This enhancer also drove GFP expression in the zebrafish brain in a tissue-specific manner, sharing aspects of endogenous ADGRL3 expression. The rs2271338 risk allele disrupts binding of YY1, an important factor in the development and function of the central nervous system. Expression quantitative trait loci analysis of post-mortem human brain tissues revealed an association between rs2271338 and reduced ADGRL3 expression in the thalamus. CONCLUSIONS These results uncover the first functional evidence of common non-coding variants with potential implications for the pathology of ADHD. PMID:27692237

NRF2: Translating the Redox Code

PubMed Central

Tummala, Krishna S.; Kottakis, Filippos; Bardeesy, Nabeel

2016-01-01

Cancer requires mechanisms to mitigate reactive oxygen species (ROS) generated during rapid growth, such as induction of the antioxidant transcription factor, Nrf2. However, the targets of ROS-mediated cytotoxicity are unclear. Recent studies in pancreatic cancer show that redox control by Nrf2 prevents cysteine oxidation of the mRNA translational machinery, thereby supporting efficient protein synthesis. PMID:27555347

Extracellular Vesicle-Associated RNA as a Carrier of Epigenetic Information

PubMed Central

2017-01-01

Post-transcriptional regulation of messenger RNA (mRNA) metabolism and subcellular localization is of the utmost importance both during development and in cell differentiation. Besides carrying genetic information, mRNAs contain cis-acting signals (zip codes), usually present in their 5′- and 3′-untranslated regions (UTRs). By binding to these signals, trans-acting factors, such as RNA-binding proteins (RBPs), and/or non-coding RNAs (ncRNAs), control mRNA localization, translation and stability. RBPs can also form complexes with non-coding RNAs of different sizes. The release of extracellular vesicles (EVs) is a conserved process that allows both normal and cancer cells to horizontally transfer molecules, and hence properties, to neighboring cells. By interacting with proteins that are specifically sorted to EVs, mRNAs as well as ncRNAs can be transferred from cell to cell. In this review, we discuss the mechanisms underlying the sorting to EVs of different classes of molecules, as well as the role of extracellular RNAs and the associated proteins in altering gene expression in the recipient cells. Importantly, if, on the one hand, RBPs play a critical role in transferring RNAs through EVs, RNA itself could, on the other hand, function as a carrier to transfer proteins (i.e., chromatin modifiers, and transcription factors) that, once transferred, can alter the cell’s epigenome. PMID:28937658

Connecting the dots: chromatin and alternative splicing in EMT.

PubMed

Warns, Jessica A; Davie, James R; Dhasarathy, Archana

2016-02-01

Nature has devised sophisticated cellular machinery to process mRNA transcripts produced by RNA Polymerase II, removing intronic regions and connecting exons together, to produce mature RNAs. This process, known as splicing, is very closely linked to transcription. Alternative splicing, or the ability to produce different combinations of exons that are spliced together from the same genomic template, is a fundamental means of regulating protein complexity. Similar to transcription, both constitutive and alternative splicing can be regulated by chromatin and its associated factors in response to various signal transduction pathways activated by external stimuli. This regulation can vary between different cell types, and interference with these pathways can lead to changes in splicing, often resulting in aberrant cellular states and disease. The epithelial to mesenchymal transition (EMT), which leads to cancer metastasis, is influenced by alternative splicing events of chromatin remodelers and epigenetic factors such as DNA methylation and non-coding RNAs. In this review, we will discuss the role of epigenetic factors including chromatin, chromatin remodelers, DNA methyltransferases, and microRNAs in the context of alternative splicing, and discuss their potential involvement in alternative splicing during the EMT process.

Regulation of Global Transcription in Escherichia coli by Rsd and 6S RNA

PubMed Central

Lal, Avantika; Krishna, Sandeep; Seshasayee, Aswin Sai Narain

2018-01-01

In Escherichia coli, the sigma factor σ70 directs RNA polymerase to transcribe growth-related genes, while σ38 directs transcription of stress response genes during stationary phase. Two molecules hypothesized to regulate RNA polymerase are the protein Rsd, which binds to σ70, and the non-coding 6S RNA which binds to the RNA polymerase-σ70 holoenzyme. Despite multiple studies, the functions of Rsd and 6S RNA remain controversial. Here we use RNA-Seq in five phases of growth to elucidate their function on a genome-wide scale. We show that Rsd and 6S RNA facilitate σ38 activity throughout bacterial growth, while 6S RNA also regulates widely different genes depending upon growth phase. We discover novel interactions between 6S RNA and Rsd and show widespread expression changes in a strain lacking both regulators. Finally, we present a mathematical model of transcription which highlights the crosstalk between Rsd and 6S RNA as a crucial factor in controlling sigma factor competition and global gene expression. PMID:29686109

Regulation of Global Transcription in Escherichia coli by Rsd and 6S RNA.

PubMed

Lal, Avantika; Krishna, Sandeep; Seshasayee, Aswin Sai Narain

2018-05-31

In Escherichia coli , the sigma factor σ 70 directs RNA polymerase to transcribe growth-related genes, while σ 38 directs transcription of stress response genes during stationary phase. Two molecules hypothesized to regulate RNA polymerase are the protein Rsd, which binds to σ 70 , and the non-coding 6S RNA which binds to the RNA polymerase-σ 70 holoenzyme. Despite multiple studies, the functions of Rsd and 6S RNA remain controversial. Here we use RNA-Seq in five phases of growth to elucidate their function on a genome-wide scale. We show that Rsd and 6S RNA facilitate σ 38 activity throughout bacterial growth, while 6S RNA also regulates widely different genes depending upon growth phase. We discover novel interactions between 6S RNA and Rsd and show widespread expression changes in a strain lacking both regulators. Finally, we present a mathematical model of transcription which highlights the crosstalk between Rsd and 6S RNA as a crucial factor in controlling sigma factor competition and global gene expression. Copyright © 2018 Lal et al.

PWMScan: a fast tool for scanning entire genomes with a position-specific weight matrix.

PubMed

Ambrosini, Giovanna; Groux, Romain; Bucher, Philipp

2018-03-05

Transcription factors (TFs) regulate gene expression by binding to specific short DNA sequences of 5 to 20-bp to regulate the rate of transcription of genetic information from DNA to messenger RNA. We present PWMScan, a fast web-based tool to scan server-resident genomes for matches to a user-supplied PWM or TF binding site model from a public database. The web server and source code are available at http://ccg.vital-it.ch/pwmscan and https://sourceforge.net/projects/pwmscan, respectively. giovanna.ambrosini@epfl.ch. SUPPLEMENTARY DATA ARE AVAILABLE AT BIOINFORMATICS ONLINE.

AP-2α and AP-2β cooperatively orchestrate homeobox gene expression during branchial arch patterning.

PubMed

Van Otterloo, Eric; Li, Hong; Jones, Kenneth L; Williams, Trevor

2018-01-25

The evolution of a hinged moveable jaw with variable morphology is considered a major factor behind the successful expansion of the vertebrates. DLX homeobox transcription factors are crucial for establishing the positional code that patterns the mandible, maxilla and intervening hinge domain, but how the genes encoding these proteins are regulated remains unclear. Herein, we demonstrate that the concerted action of the AP-2α and AP-2β transcription factors within the mouse neural crest is essential for jaw patterning. In the absence of these two proteins, the hinge domain is lost and there are alterations in the size and patterning of the jaws correlating with dysregulation of homeobox gene expression, with reduced levels of Emx, Msx and Dlx paralogs accompanied by an expansion of Six1 expression. Moreover, detailed analysis of morphological features and gene expression changes indicate significant overlap with various compound Dlx gene mutants. Together, these findings reveal that the AP-2 genes have a major function in mammalian neural crest development, influencing patterning of the craniofacial skeleton via the DLX code, an effect that has implications for vertebrate facial evolution, as well as for human craniofacial disorders. © 2018. Published by The Company of Biologists Ltd.

Variations in the non-coding transcriptome as a driver of inter-strain divergence and physiological adaptation in bacteria.

PubMed

Kopf, Matthias; Klähn, Stephan; Scholz, Ingeborg; Hess, Wolfgang R; Voß, Björn

2015-04-22

In all studied organisms, a substantial portion of the transcriptome consists of non-coding RNAs that frequently execute regulatory functions. Here, we have compared the primary transcriptomes of the cyanobacteria Synechocystis sp. PCC 6714 and PCC 6803 under 10 different conditions. These strains share 2854 protein-coding genes and a 16S rRNA identity of 99.4%, indicating their close relatedness. Conserved major transcriptional start sites (TSSs) give rise to non-coding transcripts within the sigB gene, from the 5'UTRs of cmpA and isiA, and 168 loci in antisense orientation. Distinct differences include single nucleotide polymorphisms rendering promoters inactive in one of the strains, e.g., for cmpR and for the asRNA PsbA2R. Based on the genome-wide mapped location, regulation and classification of TSSs, non-coding transcripts were identified as the most dynamic component of the transcriptome. We identified a class of mRNAs that originate by read-through from an sRNA that accumulates as a discrete and abundant transcript while also serving as the 5'UTR. Such an sRNA/mRNA structure, which we name 'actuaton', represents another way for bacteria to remodel their transcriptional network. Our findings support the hypothesis that variations in the non-coding transcriptome constitute a major evolutionary element of inter-strain divergence and capability for physiological adaptation.

Modulation of gene expression via overlapping binding sites exerted by ZNF143, Notch1 and THAP11

PubMed Central

Ngondo-Mbongo, Richard Patryk; Myslinski, Evelyne; Aster, Jon C.; Carbon, Philippe

2013-01-01

ZNF143 is a zinc-finger protein involved in the transcriptional regulation of both coding and non-coding genes from polymerase II and III promoters. Our study deciphers the genome-wide regulatory role of ZNF143 in relation with the two previously unrelated transcription factors Notch1/ICN1 and thanatos-associated protein 11 (THAP11) in several human and murine cells. We show that two distinct motifs, SBS1 and SBS2, are associated to ZNF143-binding events in promoters of >3000 genes. Without co-occupation, these sites are also bound by Notch1/ICN1 in T-lymphoblastic leukaemia cells as well as by THAP11, a factor involved in self-renewal of embryonic stem cells. We present evidence that ICN1 binding overlaps with ZNF143 binding events at the SBS1 and SBS2 motifs, whereas the overlap occurs only at SBS2 for THAP11. We demonstrate that the three factors modulate expression of common target genes through the mutually exclusive occupation of overlapping binding sites. The model we propose predicts that the binding competition between the three factors controls biological processes such as rapid cell growth of both neoplastic and stem cells. Overall, our study establishes a novel relationship between ZNF143, THAP11 and ICN1 and reveals important insights into ZNF143-mediated gene regulation. PMID:23408857

Characterization and Transcription of Arsenic Respiration and Resistance Genes During In Situ Uranium Bioremediation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Giloteaux, L.; Holmes, Dawn E.; Williams, Kenneth H.

2013-02-04

The possibility of arsenic release and the potential role of Geobacter in arsenic biogeochemistry during in situ uranium bioremediation was investigated because increased availability of organic matter has been associated with substantial releases of arsenic in other subsurface environments. In a field experiment conducted at the Rifle, CO study site, groundwater arsenic concentrations increased when acetate was added. The number of transcripts from arrA, which codes for the alpha subunit of dissimilatory As(V) reductase, and acr3, which codes for the arsenic pump protein Acr3, were determined with quantitative RT-PCR. Most of the arrA (> 60%) and acr3-1 (> 90%) sequencesmore » that were recovered were most similar to Geobacter species, while the majority of acr3-2 (>50%) sequences were most closely related to Rhodoferax ferrireducens. Analysis of transcript abundance demonstrated that transcription of acr3-1 by the subsurface Geobacter community was correlated with arsenic concentrations in the groundwater. In contrast, Geobacter arrA transcript numbers lagged behind the major arsenic release and remained high even after arsenic concentrations declined. This suggested that factors other than As(V) availability regulated transcription of arrA in situ even though the presence of As(V) increased transcription of arrA in cultures of G. lovleyi, which was capable of As(V) reduction. These results demonstrate that subsurface Geobacter species can tightly regulate their physiological response to changes in groundwater arsenic concentrations. The transcriptomic approach developed here should be useful for the study of a diversity of other environments in which Geobacter species are considered to have an important influence on arsenic biogeochemistry.« less

Characterization of Transcription from TATA-Less Promoters: Identification of a New Core Promoter Element XCPE2 and Analysis of Factor Requirements

PubMed Central

Anish, Ramakrishnan; Hossain, Mohammad B.; Jacobson, Raymond H.; Takada, Shinako

2009-01-01

Background More than 80% of mammalian protein-coding genes are driven by TATA-less promoters which often show multiple transcriptional start sites (TSSs). However, little is known about the core promoter DNA sequences or mechanisms of transcriptional initiation for this class of promoters. Methodology/Principal Findings Here we identify a new core promoter element XCPE2 (X core promoter element 2) (consensus sequence: A/C/G-C-C/T-C-G/A-T-T-G/A-C-C/A+1-C/T) that can direct specific transcription from the second TSS of hepatitis B virus X gene mRNA. XCPE2 sequences can also be found in human promoter regions and typically appear to drive one of the start sites within multiple TSS-containing TATA-less promoters. To gain insight into mechanisms of transcriptional initiation from this class of promoters, we examined requirements of several general transcription factors by in vitro transcription experiments using immunodepleted nuclear extracts and purified factors. Our results show that XCPE2-driven transcription uses at least TFIIB, either TFIID or free TBP, RNA polymerase II (RNA pol II) and the MED26-containing mediator complex but not Gcn5. Therefore, XCPE2-driven transcription can be carried out by a mechanism which differs from previously described TAF-dependent mechanisms for initiator (Inr)- or downstream promoter element (DPE)-containing promoters, the TBP- and SAGA (Spt-Ada-Gcn5-acetyltransferase)-dependent mechanism for yeast TATA-containing promoters, or the TFTC (TBP-free-TAF-containing complex)-dependent mechanism for certain Inr-containing TATA-less promoters. EMSA assays using XCPE2 promoter and purified factors further suggest that XCPE2 promoter recognition requires a set of factors different from those for TATA box, Inr, or DPE promoter recognition. Conclusions/Significance We identified a new core promoter element XCPE2 that are found in multiple TSS-containing TATA-less promoters. Mechanisms of promoter recognition and transcriptional initiation for XCPE2-driven promoters appear different from previously shown mechanisms for classical promoters that show single “focused” TSSs. Our studies provide insight into novel mechanisms of RNA Pol II transcription from multiple TSS-containing TATA-less promoters. PMID:19337366

The uncharacterized transcription factor YdhM is the regulator of the nemA gene, encoding N-ethylmaleimide reductase.

PubMed

Umezawa, Yoshimasa; Shimada, Tomohiro; Kori, Ayako; Yamada, Kayoko; Ishihama, Akira

2008-09-01

N-ethylmaleimide (NEM) has been used as a specific reagent of Cys modification in proteins and thus is toxic for cell growth. On the Escherichia coli genome, the nemA gene coding for NEM reductase is located downstream of the gene encoding an as-yet-uncharacterized transcription factor, YdhM. Disruption of the ydhM gene results in reduction of nemA expression even in the induced state, indicating that the two genes form a single operon. After in vitro genomic SELEX screening, one of the target recognition sequences for YdhM was identified within the promoter region for this ydhM-nemA operon. Both YdhM binding in vitro to the ydhM promoter region and transcription repression in vivo of the ydhM-nemA operon by YdhM were markedly reduced by the addition of NEM. Taken together, we propose that YdhM is the repressor for the nemA gene, thus hereafter designated NemR. The repressor function of NemR was inactivated by the addition of not only NEM but also other Cys modification reagents, implying that Cys modification of NemR renders it inactive. This is an addition to the mode of controlling activity of transcription factors by alkylation with chemical agents.

Drosophila PAF1 Modulates PIWI/piRNA Silencing Capacity.

PubMed

Clark, Josef P; Rahman, Reazur; Yang, Nachen; Yang, Linda H; Lau, Nelson C

2017-09-11

To test the directness of factors in initiating PIWI-directed gene silencing, we employed a Piwi-interacting RNA (piRNA)-targeted reporter assay in Drosophila ovary somatic sheet (OSS) cells [1]. This assay confirmed direct silencing roles for piRNA biogenesis factors and PIWI-associated factors [2-12] but suggested that chromatin-modifying proteins may act downstream of the initial silencing event. Our data also revealed that RNA-polymerase-II-associated proteins like PAF1 and RTF1 antagonize PIWI-directed silencing. PAF1 knockdown enhances PIWI silencing of reporters when piRNAs target the transcript region proximal to the promoter. Loss of PAF1 suppresses endogenous transposable element (TE) transcript maturation, whereas a subset of gene transcripts and long-non-coding RNAs adjacent to TE insertions are affected by PAF1 knockdown in a similar fashion to piRNA-targeted reporters. Additionally, transcription activation at specific TEs and TE-adjacent loci during PIWI knockdown is suppressed when PIWI and PAF1 levels are both reduced. Our study suggests a mechanistic conservation between fission yeast PAF1 repressing AGO1/small interfering RNA (siRNA)-directed silencing [13, 14] and Drosophila PAF1 opposing PIWI/piRNA-directed silencing. Copyright © 2017 Elsevier Ltd. All rights reserved.

De novo Assembly of Leaf Transcriptome in the Medicinal Plant Andrographis paniculata

PubMed Central

Cherukupalli, Neeraja; Divate, Mayur; Mittapelli, Suresh R.; Khareedu, Venkateswara R.; Vudem, Dashavantha R.

2016-01-01

Andrographis paniculata is an important medicinal plant containing various bioactive terpenoids and flavonoids. Despite its importance in herbal medicine, no ready-to-use transcript sequence information of this plant is made available in the public data base, this study mainly deals with the sequencing of RNA from A. paniculata leaf using Illumina HiSeq™ 2000 platform followed by the de novo transcriptome assembly. A total of 189.22 million high quality paired reads were generated and 1,70,724 transcripts were predicted in the primary assembly. Secondary assembly generated a transcriptome size of ~88 Mb with 83,800 clustered transcripts. Based on the similarity searches against plant non-redundant protein database, gene ontology, and eukaryotic orthologous groups, 49,363 transcripts were annotated constituting upto 58.91% of the identified unigenes. Annotation of transcripts—using kyoto encyclopedia of genes and genomes database—revealed 5606 transcripts plausibly involved in 140 pathways including biosynthesis of terpenoids and other secondary metabolites. Transcription factor analysis showed 6767 unique transcripts belonging to 97 different transcription factor families. A total number of 124 CYP450 transcripts belonging to seven divergent clans have been identified. Transcriptome revealed 146 different transcripts coding for enzymes involved in the biosynthesis of terpenoids of which 35 contained terpene synthase motifs. This study also revealed 32,341 simple sequence repeats (SSRs) in 23,168 transcripts. Assembled sequences of transcriptome of A. paniculata generated in this study are made available, for the first time, in the TSA database, which provides useful information for functional and comparative genomic analysis besides identification of key enzymes involved in the various pathways of secondary metabolism. PMID:27582746

The perceived organizational impact of the gender gap across a Canadian department of medicine and proposed strategies to combat it: a qualitative study.

PubMed

Pattani, Reena; Marquez, Christine; Dinyarian, Camellia; Sharma, Malika; Bain, Julie; Moore, Julia E; Straus, Sharon E

2018-04-10

Despite the gender parity existing in medical schools for over three decades, women remain underrepresented in academic medical centers, particularly in senior ranks and in leadership roles. This has consequences for patient care, education, research, and workplace culture within healthcare organizations. This study was undertaken to explore the perspectives of faculty members at a single department of medicine on the impact of the existing gender gap on organizational effectiveness and workplace culture, and to identify systems-based strategies to mitigate the gap. The study took place at a large university department of medicine in Toronto, Canada, with six affiliated hospitals. In this qualitative study, semi-structured individual interviews were conducted between May and September 2016 with full-time faculty members who held clinical and university-based appointments. Transcripts of the interviews were analyzed using thematic analysis. Three authors independently reviewed the transcripts to determine a preliminary list of codes and establish a coding framework. A modified audit consensus coding approach was applied; a single analyst reviewed all the transcripts and a second analyst audited 20% of the transcripts in each round of coding. Following each round, inter-rater reliability was determined, discrepancies were resolved through discussion, and modifications were made as needed to the coding framework. The analysis revealed faculty members' perceptions of the gender gap, potential contributing factors, organizational impacts, and possible solutions to bridge the gap. Of the 43 full-time faculty members who participated in the survey (29 of whom self-identified as female), most participants were aware of the existing gender gap within academic medicine. Participants described social exclusion, reinforced stereotypes, and unprofessional behaviors as consequences of the gap on organizational effectiveness and culture. They suggested improvements in (1) the processes for recruitment, hiring, and promotion; (2) inclusiveness of the work environment; (3) structures for mentorship; and (4) ongoing monitoring of the gap. The existing gender gap in academic medicine may have negative consequences for organizational effectiveness and workplace culture but many systems-based strategies to mitigate the gap exist. Although these solutions warrant rigorous evaluation, they are feasible to institute within most healthcare organizations immediately.

Transcriptional regulation of the operon encoding stress-responsive ECF sigma factor SigH and its anti-sigma factor RshA, and control of its regulatory network in Corynebacterium glutamicum

PubMed Central

2012-01-01

Background The expression of genes in Corynebacterium glutamicum, a Gram-positive non-pathogenic bacterium used mainly for the industrial production of amino acids, is regulated by seven different sigma factors of RNA polymerase, including the stress-responsive ECF-sigma factor SigH. The sigH gene is located in a gene cluster together with the rshA gene, putatively encoding an anti-sigma factor. The aim of this study was to analyze the transcriptional regulation of the sigH and rshA gene cluster and the effects of RshA on the SigH regulon, in order to refine the model describing the role of SigH and RshA during stress response. Results Transcription analyses revealed that the sigH gene and rshA gene are cotranscribed from four sigH housekeeping promoters in C. glutamicum. In addition, a SigH-controlled rshA promoter was found to only drive the transcription of the rshA gene. To test the role of the putative anti-sigma factor gene rshA under normal growth conditions, a C. glutamicum rshA deletion strain was constructed and used for genome-wide transcription profiling with DNA microarrays. In total, 83 genes organized in 61 putative transcriptional units, including those previously detected using sigH mutant strains, exhibited increased transcript levels in the rshA deletion mutant compared to its parental strain. The genes encoding proteins related to disulphide stress response, heat stress proteins, components of the SOS-response to DNA damage and proteasome components were the most markedly upregulated gene groups. Altogether six SigH-dependent promoters upstream of the identified genes were determined by primer extension and a refined consensus promoter consisting of 45 original promoter sequences was constructed. Conclusions The rshA gene codes for an anti-sigma factor controlling the function of the stress-responsive sigma factor SigH in C. glutamicum. Transcription of rshA from a SigH-dependent promoter may serve to quickly shutdown the SigH-dependent stress response after the cells have overcome the stress condition. Here we propose a model of the regulation of oxidative and heat stress response including redox homeostasis by SigH, RshA and the thioredoxin system. PMID:22943411

Transcriptional regulation of the operon encoding stress-responsive ECF sigma factor SigH and its anti-sigma factor RshA, and control of its regulatory network in Corynebacterium glutamicum.

PubMed

Busche, Tobias; Silar, Radoslav; Pičmanová, Martina; Pátek, Miroslav; Kalinowski, Jörn

2012-09-03

The expression of genes in Corynebacterium glutamicum, a Gram-positive non-pathogenic bacterium used mainly for the industrial production of amino acids, is regulated by seven different sigma factors of RNA polymerase, including the stress-responsive ECF-sigma factor SigH. The sigH gene is located in a gene cluster together with the rshA gene, putatively encoding an anti-sigma factor. The aim of this study was to analyze the transcriptional regulation of the sigH and rshA gene cluster and the effects of RshA on the SigH regulon, in order to refine the model describing the role of SigH and RshA during stress response. Transcription analyses revealed that the sigH gene and rshA gene are cotranscribed from four sigH housekeeping promoters in C. glutamicum. In addition, a SigH-controlled rshA promoter was found to only drive the transcription of the rshA gene. To test the role of the putative anti-sigma factor gene rshA under normal growth conditions, a C. glutamicum rshA deletion strain was constructed and used for genome-wide transcription profiling with DNA microarrays. In total, 83 genes organized in 61 putative transcriptional units, including those previously detected using sigH mutant strains, exhibited increased transcript levels in the rshA deletion mutant compared to its parental strain. The genes encoding proteins related to disulphide stress response, heat stress proteins, components of the SOS-response to DNA damage and proteasome components were the most markedly upregulated gene groups. Altogether six SigH-dependent promoters upstream of the identified genes were determined by primer extension and a refined consensus promoter consisting of 45 original promoter sequences was constructed. The rshA gene codes for an anti-sigma factor controlling the function of the stress-responsive sigma factor SigH in C. glutamicum. Transcription of rshA from a SigH-dependent promoter may serve to quickly shutdown the SigH-dependent stress response after the cells have overcome the stress condition. Here we propose a model of the regulation of oxidative and heat stress response including redox homeostasis by SigH, RshA and the thioredoxin system.

Transcriptional regulation of the human mitochondrial peptide deformylase (PDF).

PubMed

Pereira-Castro, Isabel; Costa, Luís Teixeira da; Amorim, António; Azevedo, Luisa

2012-05-18

The last years of research have been particularly dynamic in establishing the importance of peptide deformylase (PDF), a protein of the N-terminal methionine excision (NME) pathway that removes formyl-methionine from mitochondrial-encoded proteins. The genomic sequence of the human PDF gene is shared with the COG8 gene, which encodes a component of the oligomeric golgi complex, a very unusual case in Eukaryotic genomes. Since PDF is crucial in maintaining mitochondrial function and given the atypical short distance between the end of COG8 coding sequence and the PDF initiation codon, we investigated whether the regulation of the human PDF is affected by the COG8 overlapping partner. Our data reveals that PDF has several transcription start sites, the most important of which only 18 bp from the initiation codon. Furthermore, luciferase-activation assays using differently-sized fragments defined a 97 bp minimal promoter region for human PDF, which is capable of very strong transcriptional activity. This fragment contains a potential Sp1 binding site highly conserved in mammalian species. We show that this binding site, whose mutation significantly reduces transcription activation, is a target for the Sp1 transcription factor, and possibly of other members of the Sp family. Importantly, the entire minimal promoter region is located after the end of COG8's coding region, strongly suggesting that the human PDF preserves an independent regulation from its overlapping partner. Copyright © 2012 Elsevier Inc. All rights reserved.

A Comparative Encyclopedia of DNA Elements in the Mouse Genome

PubMed Central

Yue, Feng; Cheng, Yong; Breschi, Alessandra; Vierstra, Jeff; Wu, Weisheng; Ryba, Tyrone; Sandstrom, Richard; Ma, Zhihai; Davis, Carrie; Pope, Benjamin D.; Shen, Yin; Pervouchine, Dmitri D.; Djebali, Sarah; Thurman, Bob; Kaul, Rajinder; Rynes, Eric; Kirilusha, Anthony; Marinov, Georgi K.; Williams, Brian A.; Trout, Diane; Amrhein, Henry; Fisher-Aylor, Katherine; Antoshechkin, Igor; DeSalvo, Gilberto; See, Lei-Hoon; Fastuca, Meagan; Drenkow, Jorg; Zaleski, Chris; Dobin, Alex; Prieto, Pablo; Lagarde, Julien; Bussotti, Giovanni; Tanzer, Andrea; Denas, Olgert; Li, Kanwei; Bender, M. A.; Zhang, Miaohua; Byron, Rachel; Groudine, Mark T.; McCleary, David; Pham, Long; Ye, Zhen; Kuan, Samantha; Edsall, Lee; Wu, Yi-Chieh; Rasmussen, Matthew D.; Bansal, Mukul S.; Keller, Cheryl A.; Morrissey, Christapher S.; Mishra, Tejaswini; Jain, Deepti; Dogan, Nergiz; Harris, Robert S.; Cayting, Philip; Kawli, Trupti; Boyle, Alan P.; Euskirchen, Ghia; Kundaje, Anshul; Lin, Shin; Lin, Yiing; Jansen, Camden; Malladi, Venkat S.; Cline, Melissa S.; Erickson, Drew T.; Kirkup, Vanessa M; Learned, Katrina; Sloan, Cricket A.; Rosenbloom, Kate R.; de Sousa, Beatriz Lacerda; Beal, Kathryn; Pignatelli, Miguel; Flicek, Paul; Lian, Jin; Kahveci, Tamer; Lee, Dongwon; Kent, W. James; Santos, Miguel Ramalho; Herrero, Javier; Notredame, Cedric; Johnson, Audra; Vong, Shinny; Lee, Kristen; Bates, Daniel; Neri, Fidencio; Diegel, Morgan; Canfield, Theresa; Sabo, Peter J.; Wilken, Matthew S.; Reh, Thomas A.; Giste, Erika; Shafer, Anthony; Kutyavin, Tanya; Haugen, Eric; Dunn, Douglas; Reynolds, Alex P.; Neph, Shane; Humbert, Richard; Hansen, R. Scott; De Bruijn, Marella; Selleri, Licia; Rudensky, Alexander; Josefowicz, Steven; Samstein, Robert; Eichler, Evan E.; Orkin, Stuart H.; Levasseur, Dana; Papayannopoulou, Thalia; Chang, Kai-Hsin; Skoultchi, Arthur; Gosh, Srikanta; Disteche, Christine; Treuting, Piper; Wang, Yanli; Weiss, Mitchell J.; Blobel, Gerd A.; Good, Peter J.; Lowdon, Rebecca F.; Adams, Leslie B.; Zhou, Xiao-Qiao; Pazin, Michael J.; Feingold, Elise A.; Wold, Barbara; Taylor, James; Kellis, Manolis; Mortazavi, Ali; Weissman, Sherman M.; Stamatoyannopoulos, John; Snyder, Michael P.; Guigo, Roderic; Gingeras, Thomas R.; Gilbert, David M.; Hardison, Ross C.; Beer, Michael A.; Ren, Bing

2014-01-01

Summary As the premier model organism in biomedical research, the laboratory mouse shares the majority of protein-coding genes with humans, yet the two mammals differ in significant ways. To gain greater insights into both shared and species-specific transcriptional and cellular regulatory programs in the mouse, the Mouse ENCODE Consortium has mapped transcription, DNase I hypersensitivity, transcription factor binding, chromatin modifications, and replication domains throughout the mouse genome in diverse cell and tissue types. By comparing with the human genome, we not only confirm substantial conservation in the newly annotated potential functional sequences, but also find a large degree of divergence of other sequences involved in transcriptional regulation, chromatin state and higher order chromatin organization. Our results illuminate the wide range of evolutionary forces acting on genes and their regulatory regions, and provide a general resource for research into mammalian biology and mechanisms of human diseases. PMID:25409824

A comparative encyclopedia of DNA elements in the mouse genome.

PubMed

Yue, Feng; Cheng, Yong; Breschi, Alessandra; Vierstra, Jeff; Wu, Weisheng; Ryba, Tyrone; Sandstrom, Richard; Ma, Zhihai; Davis, Carrie; Pope, Benjamin D; Shen, Yin; Pervouchine, Dmitri D; Djebali, Sarah; Thurman, Robert E; Kaul, Rajinder; Rynes, Eric; Kirilusha, Anthony; Marinov, Georgi K; Williams, Brian A; Trout, Diane; Amrhein, Henry; Fisher-Aylor, Katherine; Antoshechkin, Igor; DeSalvo, Gilberto; See, Lei-Hoon; Fastuca, Meagan; Drenkow, Jorg; Zaleski, Chris; Dobin, Alex; Prieto, Pablo; Lagarde, Julien; Bussotti, Giovanni; Tanzer, Andrea; Denas, Olgert; Li, Kanwei; Bender, M A; Zhang, Miaohua; Byron, Rachel; Groudine, Mark T; McCleary, David; Pham, Long; Ye, Zhen; Kuan, Samantha; Edsall, Lee; Wu, Yi-Chieh; Rasmussen, Matthew D; Bansal, Mukul S; Kellis, Manolis; Keller, Cheryl A; Morrissey, Christapher S; Mishra, Tejaswini; Jain, Deepti; Dogan, Nergiz; Harris, Robert S; Cayting, Philip; Kawli, Trupti; Boyle, Alan P; Euskirchen, Ghia; Kundaje, Anshul; Lin, Shin; Lin, Yiing; Jansen, Camden; Malladi, Venkat S; Cline, Melissa S; Erickson, Drew T; Kirkup, Vanessa M; Learned, Katrina; Sloan, Cricket A; Rosenbloom, Kate R; Lacerda de Sousa, Beatriz; Beal, Kathryn; Pignatelli, Miguel; Flicek, Paul; Lian, Jin; Kahveci, Tamer; Lee, Dongwon; Kent, W James; Ramalho Santos, Miguel; Herrero, Javier; Notredame, Cedric; Johnson, Audra; Vong, Shinny; Lee, Kristen; Bates, Daniel; Neri, Fidencio; Diegel, Morgan; Canfield, Theresa; Sabo, Peter J; Wilken, Matthew S; Reh, Thomas A; Giste, Erika; Shafer, Anthony; Kutyavin, Tanya; Haugen, Eric; Dunn, Douglas; Reynolds, Alex P; Neph, Shane; Humbert, Richard; Hansen, R Scott; De Bruijn, Marella; Selleri, Licia; Rudensky, Alexander; Josefowicz, Steven; Samstein, Robert; Eichler, Evan E; Orkin, Stuart H; Levasseur, Dana; Papayannopoulou, Thalia; Chang, Kai-Hsin; Skoultchi, Arthur; Gosh, Srikanta; Disteche, Christine; Treuting, Piper; Wang, Yanli; Weiss, Mitchell J; Blobel, Gerd A; Cao, Xiaoyi; Zhong, Sheng; Wang, Ting; Good, Peter J; Lowdon, Rebecca F; Adams, Leslie B; Zhou, Xiao-Qiao; Pazin, Michael J; Feingold, Elise A; Wold, Barbara; Taylor, James; Mortazavi, Ali; Weissman, Sherman M; Stamatoyannopoulos, John A; Snyder, Michael P; Guigo, Roderic; Gingeras, Thomas R; Gilbert, David M; Hardison, Ross C; Beer, Michael A; Ren, Bing

2014-11-20

The laboratory mouse shares the majority of its protein-coding genes with humans, making it the premier model organism in biomedical research, yet the two mammals differ in significant ways. To gain greater insights into both shared and species-specific transcriptional and cellular regulatory programs in the mouse, the Mouse ENCODE Consortium has mapped transcription, DNase I hypersensitivity, transcription factor binding, chromatin modifications and replication domains throughout the mouse genome in diverse cell and tissue types. By comparing with the human genome, we not only confirm substantial conservation in the newly annotated potential functional sequences, but also find a large degree of divergence of sequences involved in transcriptional regulation, chromatin state and higher order chromatin organization. Our results illuminate the wide range of evolutionary forces acting on genes and their regulatory regions, and provide a general resource for research into mammalian biology and mechanisms of human diseases.

A Catalogue of Putative cis-Regulatory Interactions Between Long Non-coding RNAs and Proximal Coding Genes Based on Correlative Analysis Across Diverse Human Tumors.

PubMed

Basu, Swaraj; Larsson, Erik

2018-05-31

Antisense transcripts and other long non-coding RNAs are pervasive in mammalian cells, and some of these molecules have been proposed to regulate proximal protein-coding genes in cis For example, non-coding transcription can contribute to inactivation of tumor suppressor genes in cancer, and antisense transcripts have been implicated in the epigenetic inactivation of imprinted genes. However, our knowledge is still limited and more such regulatory interactions likely await discovery. Here, we make use of available gene expression data from a large compendium of human tumors to generate hypotheses regarding non-coding-to-coding cis -regulatory relationships with emphasis on negative associations, as these are less likely to arise for reasons other than cis -regulation. We document a large number of possible regulatory interactions, including 193 coding/non-coding pairs that show expression patterns compatible with negative cis -regulation. Importantly, by this approach we capture several known cases, and many of the involved coding genes have known roles in cancer. Our study provides a large catalog of putative non-coding/coding cis -regulatory pairs that may serve as a basis for further experimental validation and characterization. Copyright © 2018 Basu and Larsson.

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

PubMed

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

2018-03-22

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

Distinct Developmental Origins Manifest in the Specialized Encoding of Movement by Adult Neurons of the External Globus Pallidus

PubMed Central

Dodson, Paul D.; Larvin, Joseph T.; Duffell, James M.; Garas, Farid N.; Doig, Natalie M.; Kessaris, Nicoletta; Duguid, Ian C.; Bogacz, Rafal; Butt, Simon J.B.; Magill, Peter J.

2015-01-01

Summary Transcriptional codes initiated during brain development are ultimately realized in adulthood as distinct cell types performing specialized roles in behavior. Focusing on the mouse external globus pallidus (GPe), we demonstrate that the potential contributions of two GABAergic GPe cell types to voluntary action are fated from early life to be distinct. Prototypic GPe neurons derive from the medial ganglionic eminence of the embryonic subpallium and express the transcription factor Nkx2-1. These neurons fire at high rates during alert rest, and encode movements through heterogeneous firing rate changes, with many neurons decreasing their activity. In contrast, arkypallidal GPe neurons originate from lateral/caudal ganglionic eminences, express the transcription factor FoxP2, fire at low rates during rest, and encode movements with robust increases in firing. We conclude that developmental diversity positions prototypic and arkypallidal neurons to fulfil distinct roles in behavior via their disparate regulation of GABA release onto different basal ganglia targets. PMID:25843402

MicroRNAs in Palatogenesis and Cleft Palate

PubMed Central

Schoen, Christian; Aschrafi, Armaz; Thonissen, Michelle; Poelmans, Geert; Von den Hoff, Johannes W.; Carels, Carine E. L.

2017-01-01

Palatogenesis requires a precise spatiotemporal regulation of gene expression, which is controlled by an intricate network of transcription factors and their corresponding DNA motifs. Even minor perturbations of this network may cause cleft palate, the most common congenital craniofacial defect in humans. MicroRNAs (miRNAs), a class of small regulatory non-coding RNAs, have elicited strong interest as key regulators of embryological development, and as etiological factors in disease. MiRNAs function as post-transcriptional repressors of gene expression and are therefore able to fine-tune gene regulatory networks. Several miRNAs are already identified to be involved in congenital diseases. Recent evidence from research in zebrafish and mice indicates that miRNAs are key factors in both normal palatogenesis and cleft palate formation. Here, we provide an overview of recently identified molecular mechanisms underlying palatogenesis involving specific miRNAs, and discuss how dysregulation of these miRNAs may result in cleft palate. PMID:28420997

Genome-scale cold stress response regulatory networks in ten Arabidopsis thaliana ecotypes

PubMed Central

2013-01-01

Background Low temperature leads to major crop losses every year. Although several studies have been conducted focusing on diversity of cold tolerance level in multiple phenotypically divergent Arabidopsis thaliana (A. thaliana) ecotypes, genome-scale molecular understanding is still lacking. Results In this study, we report genome-scale transcript response diversity of 10 A. thaliana ecotypes originating from different geographical locations to non-freezing cold stress (10°C). To analyze the transcriptional response diversity, we initially compared transcriptome changes in all 10 ecotypes using Arabidopsis NimbleGen ATH6 microarrays. In total 6061 transcripts were significantly cold regulated (p < 0.01) in 10 ecotypes, including 498 transcription factors and 315 transposable elements. The majority of the transcripts (75%) showed ecotype specific expression pattern. By using sequence data available from Arabidopsis thaliana 1001 genome project, we further investigated sequence polymorphisms in the core cold stress regulon genes. Significant numbers of non-synonymous amino acid changes were observed in the coding region of the CBF regulon genes. Considering the limited knowledge about regulatory interactions between transcription factors and their target genes in the model plant A. thaliana, we have adopted a powerful systems genetics approach- Network Component Analysis (NCA) to construct an in-silico transcriptional regulatory network model during response to cold stress. The resulting regulatory network contained 1,275 nodes and 7,720 connections, with 178 transcription factors and 1,331 target genes. Conclusions A. thaliana ecotypes exhibit considerable variation in transcriptome level responses to non-freezing cold stress treatment. Ecotype specific transcripts and related gene ontology (GO) categories were identified to delineate natural variation of cold stress regulated differential gene expression in the model plant A. thaliana. The predicted regulatory network model was able to identify new ecotype specific transcription factors and their regulatory interactions, which might be crucial for their local geographic adaptation to cold temperature. Additionally, since the approach presented here is general, it could be adapted to study networks regulating biological process in any biological systems. PMID:24148294

Serine/Threonine kinase dependent transcription from the polyhedrin promoter of SpltNPV-I

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mishra, Gourav; Gautam, Hemant K.; Das, Rakha H.

2007-07-06

Polyhedrin (polh) and p10 are the two hyper-expressed very late genes of nucleopolyhedroviruses. Alpha amanitin resistant transcription from Spodoptera litura nucleopolyhedrovirus (SpltNPV-I) polyhedrin promoter was observed with virus infected nuclear extract of NIV-HA-197 cells but not with that from uninfected nuclear extract. Anti-protein kinase-1 (pk1) antibody inhibited the transcription and the inhibition reversed on addition of pk1, however, pk1 mutant protein, K50M having no phosphorylation activity did not overcome the transcription inhibition. Chromatin immuno-precipitation assays with viral anti-pk1 antibody showed the interaction of pk1 with the polh while electrophoretic mobility shift assays indicated the strong binding affinity (K {sub d}more » {approx} 5.5 x 10{sup -11}) of purified pk1 with the polh promoter. These results suggested that the viral coded pk1 acts as a transcription factor in transcribing baculovirus very late genes.« less

GENCODE: the reference human genome annotation for The ENCODE Project.

PubMed

Harrow, Jennifer; Frankish, Adam; Gonzalez, Jose M; Tapanari, Electra; Diekhans, Mark; Kokocinski, Felix; Aken, Bronwen L; Barrell, Daniel; Zadissa, Amonida; Searle, Stephen; Barnes, If; Bignell, Alexandra; Boychenko, Veronika; Hunt, Toby; Kay, Mike; Mukherjee, Gaurab; Rajan, Jeena; Despacio-Reyes, Gloria; Saunders, Gary; Steward, Charles; Harte, Rachel; Lin, Michael; Howald, Cédric; Tanzer, Andrea; Derrien, Thomas; Chrast, Jacqueline; Walters, Nathalie; Balasubramanian, Suganthi; Pei, Baikang; Tress, Michael; Rodriguez, Jose Manuel; Ezkurdia, Iakes; van Baren, Jeltje; Brent, Michael; Haussler, David; Kellis, Manolis; Valencia, Alfonso; Reymond, Alexandre; Gerstein, Mark; Guigó, Roderic; Hubbard, Tim J

2012-09-01

The GENCODE Consortium aims to identify all gene features in the human genome using a combination of computational analysis, manual annotation, and experimental validation. Since the first public release of this annotation data set, few new protein-coding loci have been added, yet the number of alternative splicing transcripts annotated has steadily increased. The GENCODE 7 release contains 20,687 protein-coding and 9640 long noncoding RNA loci and has 33,977 coding transcripts not represented in UCSC genes and RefSeq. It also has the most comprehensive annotation of long noncoding RNA (lncRNA) loci publicly available with the predominant transcript form consisting of two exons. We have examined the completeness of the transcript annotation and found that 35% of transcriptional start sites are supported by CAGE clusters and 62% of protein-coding genes have annotated polyA sites. Over one-third of GENCODE protein-coding genes are supported by peptide hits derived from mass spectrometry spectra submitted to Peptide Atlas. New models derived from the Illumina Body Map 2.0 RNA-seq data identify 3689 new loci not currently in GENCODE, of which 3127 consist of two exon models indicating that they are possibly unannotated long noncoding loci. GENCODE 7 is publicly available from gencodegenes.org and via the Ensembl and UCSC Genome Browsers.

Aging in the Brain: New Roles of Epigenetics in Cognitive Decline.

PubMed

Barter, Jolie D; Foster, Thomas C

2018-06-01

Gene expression in the aging brain depends on transcription signals generated by senescent physiology, interacting with genetic and epigenetic programs. In turn, environmental factors influence epigenetic mechanisms, such that an epigenetic-environmental link may contribute to the accumulation of cellular damage, susceptibility or resilience to stressors, and variability in the trajectory of age-related cognitive decline. Epigenetic mechanisms, DNA methylation and histone modifications, alter chromatin structure and the accessibility of DNA. Furthermore, small non-coding RNA, termed microRNA (miRNA) bind to messenger RNA (mRNA) to regulate translation. In this review, we examine key questions concerning epigenetic mechanisms in regulating the expression of genes associated with brain aging and age-related cognitive decline. In addition, we highlight the interaction of epigenetics with senescent physiology and environmental factors in regulating transcription.

Characterization of stress-responsive lncRNAs in Arabidopsis thaliana by integrating expression, epigenetic and structural features.

PubMed

Di, Chao; Yuan, Jiapei; Wu, Yue; Li, Jingrui; Lin, Huixin; Hu, Long; Zhang, Ting; Qi, Yijun; Gerstein, Mark B; Guo, Yan; Lu, Zhi John

2014-12-01

Recently, in addition to poly(A)+ long non-coding RNAs (lncRNAs), many lncRNAs without poly(A) tails, have been characterized in mammals. However, the non-polyA lncRNAs and their conserved motifs, especially those associated with environmental stresses, have not been fully investigated in plant genomes. We performed poly(A)- RNA-seq for seedlings of Arabidopsis thaliana under four stress conditions, and predicted lncRNA transcripts. We classified the lncRNAs into three confidence levels according to their expression patterns, epigenetic signatures and RNA secondary structures. Then, we further classified the lncRNAs to poly(A)+ and poly(A)- transcripts. Compared with poly(A)+ lncRNAs and coding genes, we found that poly(A)- lncRNAs tend to have shorter transcripts and lower expression levels, and they show significant expression specificity in response to stresses. In addition, their differential expression is significantly enriched in drought condition and depleted in heat condition. Overall, we identified 245 poly(A)+ and 58 poly(A)- lncRNAs that are differentially expressed under various stress stimuli. The differential expression was validated by qRT-PCR, and the signaling pathways involved were supported by specific binding of transcription factors (TFs), phytochrome-interacting factor 4 (PIF4) and PIF5. Moreover, we found many conserved sequence and structural motifs of lncRNAs from different functional groups (e.g. a UUC motif responding to salt and a AU-rich stem-loop responding to cold), indicated that the conserved elements might be responsible for the stress-responsive functions of lncRNAs. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

Barriers to Early Detection of Breast Cancer Among African American Females Over Age of 55

DTIC Science & Technology

2005-02-01

used for data analysis. NUDIST , software for qualitative data analysis will be used for systematic coding. All transcripts, as well as interviewer notes...will be coded in NUDIST . Dr. Smith and Mr. Worts will jointly develop the NUDIST coding system. Each of them will separately code each transcript and...already provided training in NUDIST to Dr. Smith and Mr. Worts. All interviews will be conducted by the Principal Investigator for this study who is

Non-coding, mRNA-like RNAs database Y2K.

PubMed

Erdmann, V A; Szymanski, M; Hochberg, A; Groot, N; Barciszewski, J

2000-01-01

In last few years much data has accumulated on various non-translatable RNA transcripts that are synthesised in different cells. They are lacking in protein coding capacity and it seems that they work mainly or exclusively at the RNA level. All known non-coding RNA transcripts are collected in the database: http://www. man.poznan.pl/5SData/ncRNA/index.html

Non-coding, mRNA-like RNAs database Y2K

PubMed Central

Erdmann, Volker A.; Szymanski, Maciej; Hochberg, Abraham; Groot, Nathan de; Barciszewski, Jan

2000-01-01

In last few years much data has accumulated on various non-translatable RNA transcripts that are synthesised in different cells. They are lacking in protein coding capacity and it seems that they work mainly or exclusively at the RNA level. All known non-coding RNA transcripts are collected in the database: http://www.man.poznan.pl/5SData/ncRNA/index.html PMID:10592224

Decoding the non-coding genome: elucidating genetic risk outside the coding genome.

PubMed

Barr, C L; Misener, V L

2016-01-01

Current evidence emerging from genome-wide association studies indicates that the genetic underpinnings of complex traits are likely attributable to genetic variation that changes gene expression, rather than (or in combination with) variation that changes protein-coding sequences. This is particularly compelling with respect to psychiatric disorders, as genetic changes in regulatory regions may result in differential transcriptional responses to developmental cues and environmental/psychosocial stressors. Until recently, however, the link between transcriptional regulation and psychiatric genetic risk has been understudied. Multiple obstacles have contributed to the paucity of research in this area, including challenges in identifying the positions of remote (distal from the promoter) regulatory elements (e.g. enhancers) and their target genes and the underrepresentation of neural cell types and brain tissues in epigenome projects - the availability of high-quality brain tissues for epigenetic and transcriptome profiling, particularly for the adolescent and developing brain, has been limited. Further challenges have arisen in the prediction and testing of the functional impact of DNA variation with respect to multiple aspects of transcriptional control, including regulatory-element interaction (e.g. between enhancers and promoters), transcription factor binding and DNA methylation. Further, the brain has uncommon DNA-methylation marks with unique genomic distributions not found in other tissues - current evidence suggests the involvement of non-CG methylation and 5-hydroxymethylation in neurodevelopmental processes but much remains unknown. We review here knowledge gaps as well as both technological and resource obstacles that will need to be overcome in order to elucidate the involvement of brain-relevant gene-regulatory variants in genetic risk for psychiatric disorders. © 2015 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

Regulation of a maize HD-ZIP IV transcription factor by a non-conventional RDR2-dependent small RNA.

PubMed

Klein-Cosson, Catherine; Chambrier, Pierre; Rogowsky, Peter M; Vernoud, Vanessa

2015-03-01

Small non-coding RNAs are versatile riboregulators that control gene expression at the transcriptional or post-transcriptional level, governing many facets of plant development. Here we present evidence for the existence of a 24 nt small RNA (named small1) that is complementary to the 3' UTR of OCL1 (Outer Cell Layer1), the founding member of the maize HD-ZIP IV gene family encoding plant-specific transcription factors that are mainly involved in epidermis differentiation and specialization. The biogenesis of small1 depends on DICER-like 3 (DCL3), RNA-dependent RNA polymerase 2 (RDR2) and RNA polymerase IV, components that are usually required for RNA-dependent DNA-methylation. Unexpectedly, GFP sensor experiments in transient and stable transformation systems revealed that small1 may regulate its target at the post-transcriptional level, mainly through translational repression. This translational repression is attenuated in an rdr2 mutant background in which small1 does not accumulate. Our experiments further showed the possible involvement of a secondary stem-loop structure present in the 3' UTR of OCL1 for efficient target repression, suggesting the existence of several regulatory mechanisms affecting OCL1 mRNA stability and translation. © 2015 The Authors The Plant Journal © 2015 John Wiley & Sons Ltd.

Combinatorial influence of environmental parameters on transcription factor activity.

PubMed

Knijnenburg, T A; Wessels, L F A; Reinders, M J T

2008-07-01

Cells receive a wide variety of environmental signals, which are often processed combinatorially to generate specific genetic responses. Changes in transcript levels, as observed across different environmental conditions, can, to a large extent, be attributed to changes in the activity of transcription factors (TFs). However, in unraveling these transcription regulation networks, the actual environmental signals are often not incorporated into the model, simply because they have not been measured. The unquantified heterogeneity of the environmental parameters across microarray experiments frustrates regulatory network inference. We propose an inference algorithm that models the influence of environmental parameters on gene expression. The approach is based on a yeast microarray compendium of chemostat steady-state experiments. Chemostat cultivation enables the accurate control and measurement of many of the key cultivation parameters, such as nutrient concentrations, growth rate and temperature. The observed transcript levels are explained by inferring the activity of TFs in response to combinations of cultivation parameters. The interplay between activated enhancers and repressors that bind a gene promoter determine the possible up- or downregulation of the gene. The model is translated into a linear integer optimization problem. The resulting regulatory network identifies the combinatorial effects of environmental parameters on TF activity and gene expression. The Matlab code is available from the authors upon request. Supplementary data are available at Bioinformatics online.

Deciphering Mineral Homeostasis in Barley Seed Transfer Cells at Transcriptional Level.

PubMed

Darbani, Behrooz; Noeparvar, Shahin; Borg, Søren

2015-01-01

In addition to the micronutrient inadequacy of staple crops for optimal human nutrition, a global downtrend in crop-quality has emerged from intensive breeding for yield. This trend will be aggravated by elevated levels of the greenhouse gas carbon dioxide. Therefore, crop biofortification is inevitable to ensure a sustainable supply of minerals to the large part of human population who is dietary dependent on staple crops. This requires a thorough understanding of plant-mineral interactions due to the complexity of mineral homeostasis. Employing RNA sequencing, we here communicate transfer cell specific effects of excess iron and zinc during grain filling in our model crop plant barley. Responding to alterations in mineral contents, we found a long range of different genes and transcripts. Among them, it is worth to highlight the auxin and ethylene signaling factors Arfs, Abcbs, Cand1, Hps4, Hac1, Ecr1, and Ctr1, diurnal fluctuation components Sdg2, Imb1, Lip1, and PhyC, retroelements, sulfur homeostasis components Amp1, Hmt3, Eil3, and Vip1, mineral trafficking components Med16, Cnnm4, Aha2, Clpc1, and Pcbps, and vacuole organization factors Ymr155W, RabG3F, Vps4, and Cbl3. Our analysis introduces new interactors and signifies a broad spectrum of regulatory levels from chromatin remodeling to intracellular protein sorting mechanisms active in the plant mineral homeostasis. The results highlight the importance of storage proteins in metal ion toxicity-resistance and chelation. Interestingly, the protein sorting and recycling factors Exoc7, Cdc1, Sec23A, and Rab11A contributed to the response as well as the polar distributors of metal-transporters ensuring the directional flow of minerals. Alternative isoform switching was found important for plant adaptation and occurred among transcripts coding for identical proteins as well as transcripts coding for protein isoforms. We also identified differences in the alternative-isoform preference between the treatments, indicating metal-affinity shifts among isoforms of metal transporters. Most important, we found the zinc treatment to impair both photosynthesis and respiration. A wide range of transcriptional changes including stress-related genes and negative feedback loops emphasize the importance to withhold mineral contents below certain cellular levels which otherwise might lead to agronomical impeding side-effects. By illustrating new mechanisms, genes, and transcripts, this report provides a solid platform towards understanding the complex network of plant mineral homeostasis.

A Change in SHATTERPROOF Protein Lies at the Origin of a Fruit Morphological Novelty and a New Strategy for Seed Dispersal in Medicago Genus1[C][W

PubMed Central

Fourquin, Chloé; del Cerro, Carolina; Victoria, Filipe C.; Vialette-Guiraud, Aurélie; de Oliveira, Antonio C.; Ferrándiz, Cristina

2013-01-01

Angiosperms are the most diverse and numerous group of plants, and it is generally accepted that this evolutionary success owes in part to the diversity found in fruits, key for protecting the developing seeds and ensuring seed dispersal. Although studies on the molecular basis of morphological innovations are few, they all illustrate the central role played by transcription factors acting as developmental regulators. Here, we show that a small change in the protein sequence of a MADS-box transcription factor correlates with the origin of a highly modified fruit morphology and the change in seed dispersal strategies that occurred in Medicago, a genus belonging to the large legume family. This protein sequence modification alters the functional properties of the protein, affecting the affinities for other protein partners involved in high-order complexes. Our work illustrates that variation in coding regions can generate evolutionary novelties not based on gene duplication/subfunctionalization but by interactions in complex networks, contributing also to the current debate on the relative importance of changes in regulatory or coding regions of master regulators in generating morphological novelties. PMID:23640757

Protein-DNA binding dynamics predict transcriptional response to nutrients in archaea.

PubMed

Todor, Horia; Sharma, Kriti; Pittman, Adrianne M C; Schmid, Amy K

2013-10-01

Organisms across all three domains of life use gene regulatory networks (GRNs) to integrate varied stimuli into coherent transcriptional responses to environmental pressures. However, inferring GRN topology and regulatory causality remains a central challenge in systems biology. Previous work characterized TrmB as a global metabolic transcription factor in archaeal extremophiles. However, it remains unclear how TrmB dynamically regulates its ∼100 metabolic enzyme-coding gene targets. Using a dynamic perturbation approach, we elucidate the topology of the TrmB metabolic GRN in the model archaeon Halobacterium salinarum. Clustering of dynamic gene expression patterns reveals that TrmB functions alone to regulate central metabolic enzyme-coding genes but cooperates with various regulators to control peripheral metabolic pathways. Using a dynamical model, we predict gene expression patterns for some TrmB-dependent promoters and infer secondary regulators for others. Our data suggest feed-forward gene regulatory topology for cobalamin biosynthesis. In contrast, purine biosynthesis appears to require TrmB-independent regulators. We conclude that TrmB is an important component for mediating metabolic modularity, integrating nutrient status and regulating gene expression dynamics alone and in concert with secondary regulators.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gehrmann, Thies; Pelkmans, Jordi F.; Lugones, Luis G.

Recent genome-wide studies have demonstrated that fungi possess the machinery to alternatively splice pre-mRNA. However, there has not been a systematic categorization of the functional impact of alternative splicing in a fungus. We investigate alternative splicing and its functional consequences in the model mushroom forming fungus Schizophyllum commune. Alternative splicing was demonstrated for 2,285 out of 12,988 expressed genes, resulting in 20% additional transcripts. Intron retentions were the most common alternative splicing events, accounting for 33% of all splicing events, and 43% of the events in coding regions. On the other hand, exon skipping events were rare in coding regionsmore » (1%) but enriched in UTRs where they accounted for 57% of the events. Specific functional groups, including transcription factors, contained alternatively spliced genes. Alternatively spliced transcripts were regulated differently throughout development in 19% of the 2,285 alternatively spliced genes. Notably, 69% of alternatively spliced genes have predicted alternative functionality by loss or gain of functional domains, or by acquiring alternative subcellular locations. S. commune exhibits more alternative splicing than any other studied fungus. Finally, taken together, alternative splicing increases the complexity of the S. commune proteome considerably and provides it with a rich repertoire of alternative functionality that is exploited dynamically.« less

The structure of transcription termination factor Nrd1 reveals an original mode for GUAA recognition

PubMed Central

Franco-Echevarría, Elsa; González-Polo, Noelia; Zorrilla, Silvia; Martínez-Lumbreras, Santiago; Santiveri, Clara M.; Campos-Olivas, Ramón; Sánchez, Mar; Calvo, Olga

2017-01-01

Abstract Transcription termination of non-coding RNAs is regulated in yeast by a complex of three RNA binding proteins: Nrd1, Nab3 and Sen1. Nrd1 is central in this process by interacting with Rbp1 of RNA polymerase II, Trf4 of TRAMP and GUAA/G terminator sequences. We lack structural data for the last of these binding events. We determined the structures of Nrd1 RNA binding domain and its complexes with three GUAA-containing RNAs, characterized RNA binding energetics and tested rationally designed mutants in vivo. The Nrd1 structure shows an RRM domain fused with a second α/β domain that we name split domain (SD), because it is formed by two non-consecutive segments at each side of the RRM. The GUAA interacts with both domains and with a pocket of water molecules, trapped between the two stacking adenines and the SD. Comprehensive binding studies demonstrate for the first time that Nrd1 has a slight preference for GUAA over GUAG and genetic and functional studies suggest that Nrd1 RNA binding domain might play further roles in non-coding RNAs transcription termination. PMID:28973465

Chromatin-bound RNA and the neurobiology of psychiatric disease.

PubMed

Tushir, J S; Akbarian, S

2014-04-04

A large, and still rapidly expanding literature on epigenetic regulation in the nervous system has provided fundamental insights into the dynamic regulation of DNA methylation and post-translational histone modifications in the context of neuronal plasticity in health and disease. Remarkably, however, very little is known about the potential role of chromatin-bound RNAs, including many long non-coding transcripts and various types of small RNAs. Here, we provide an overview on RNA-mediated regulation of chromatin structure and function, with focus on histone lysine methylation and psychiatric disease. Examples of recently discovered chromatin-bound long non-coding RNAs important for neuronal health and function include the brain-derived neurotrophic factor antisense transcript (Bdnf-AS) which regulates expression of the corresponding sense transcript, and LOC389023 which is associated with human-specific histone methylation signatures at the chromosome 2q14.1 neurodevelopmental risk locus by regulating expression of DPP10, an auxillary subunit for voltage-gated K(+) channels. We predict that the exploration of chromatin-bound RNA will significantly advance our current knowledge base in neuroepigenetics and biological psychiatry. Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.

Early Cone Setting in Picea abies acrocona Is Associated with Increased Transcriptional Activity of a MADS Box Transcription Factor1[W][OA

PubMed Central

Uddenberg, Daniel; Reimegård, Johan; Clapham, David; Almqvist, Curt; von Arnold, Sara; Emanuelsson, Olof; Sundström, Jens F.

2013-01-01

Conifers normally go through a long juvenile period, for Norway spruce (Picea abies) around 20 to 25 years, before developing male and female cones. We have grown plants from inbred crosses of a naturally occurring spruce mutant (acrocona). One-fourth of the segregating acrocona plants initiate cones already in their second growth cycle, suggesting control by a single locus. The early cone-setting properties of the acrocona mutant were utilized to identify candidate genes involved in vegetative-to-reproductive phase change in Norway spruce. Poly(A+) RNA samples from apical and basal shoots of cone-setting and non-cone-setting plants were subjected to high-throughput sequencing (RNA-seq). We assembled and investigated 33,383 expressed putative protein-coding acrocona transcripts. Eight transcripts were differentially expressed between selected sample pairs. One of these (Acr42124_1) was significantly up-regulated in apical shoot samples from cone-setting acrocona plants, and the encoded protein belongs to the MADS box gene family of transcription factors. Using quantitative real-time polymerase chain reaction with independently derived plant material, we confirmed that the MADS box gene is up-regulated in both needles and buds of cone-inducing shoots when reproductive identity is determined. Our results constitute important steps for the development of a rapid cycling model system that can be used to study gene function in conifers. In addition, our data suggest the involvement of a MADS box transcription factor in the vegetative-to-reproductive phase change in Norway spruce. PMID:23221834

Deleterious ABCA7 mutations and transcript rescue mechanisms in early onset Alzheimer's disease.

PubMed

De Roeck, Arne; Van den Bossche, Tobi; van der Zee, Julie; Verheijen, Jan; De Coster, Wouter; Van Dongen, Jasper; Dillen, Lubina; Baradaran-Heravi, Yalda; Heeman, Bavo; Sanchez-Valle, Raquel; Lladó, Albert; Nacmias, Benedetta; Sorbi, Sandro; Gelpi, Ellen; Grau-Rivera, Oriol; Gómez-Tortosa, Estrella; Pastor, Pau; Ortega-Cubero, Sara; Pastor, Maria A; Graff, Caroline; Thonberg, Håkan; Benussi, Luisa; Ghidoni, Roberta; Binetti, Giuliano; de Mendonça, Alexandre; Martins, Madalena; Borroni, Barbara; Padovani, Alessandro; Almeida, Maria Rosário; Santana, Isabel; Diehl-Schmid, Janine; Alexopoulos, Panagiotis; Clarimon, Jordi; Lleó, Alberto; Fortea, Juan; Tsolaki, Magda; Koutroumani, Maria; Matěj, Radoslav; Rohan, Zdenek; De Deyn, Peter; Engelborghs, Sebastiaan; Cras, Patrick; Van Broeckhoven, Christine; Sleegers, Kristel

2017-09-01

Premature termination codon (PTC) mutations in the ATP-Binding Cassette, Sub-Family A, Member 7 gene (ABCA7) have recently been identified as intermediate-to-high penetrant risk factor for late-onset Alzheimer's disease (LOAD). High variability, however, is observed in downstream ABCA7 mRNA and protein expression, disease penetrance, and onset age, indicative of unknown modifying factors. Here, we investigated the prevalence and disease penetrance of ABCA7 PTC mutations in a large early onset AD (EOAD)-control cohort, and examined the effect on transcript level with comprehensive third-generation long-read sequencing. We characterized the ABCA7 coding sequence with next-generation sequencing in 928 EOAD patients and 980 matched control individuals. With MetaSKAT rare variant association analysis, we observed a fivefold enrichment (p = 0.0004) of PTC mutations in EOAD patients (3%) versus controls (0.6%). Ten novel PTC mutations were only observed in patients, and PTC mutation carriers in general had an increased familial AD load. In addition, we observed nominal risk reducing trends for three common coding variants. Seven PTC mutations were further analyzed using targeted long-read cDNA sequencing on an Oxford Nanopore MinION platform. PTC-containing transcripts for each investigated PTC mutation were observed at varying proportion (5-41% of the total read count), implying incomplete nonsense-mediated mRNA decay (NMD). Furthermore, we distinguished and phased several previously unknown alternative splicing events (up to 30% of transcripts). In conjunction with PTC mutations, several of these novel ABCA7 isoforms have the potential to rescue deleterious PTC effects. In conclusion, ABCA7 PTC mutations play a substantial role in EOAD, warranting genetic screening of ABCA7 in genetically unexplained patients. Long-read cDNA sequencing revealed both varying degrees of NMD and transcript-modifying events, which may influence ABCA7 dosage, disease severity, and may create opportunities for therapeutic interventions in AD.

YY1 binding association with sex-biased transcription revealed through X-linked transcript levels and allelic binding analyses.

PubMed

Chen, Chih-Yu; Shi, Wenqiang; Balaton, Bradley P; Matthews, Allison M; Li, Yifeng; Arenillas, David J; Mathelier, Anthony; Itoh, Masayoshi; Kawaji, Hideya; Lassmann, Timo; Hayashizaki, Yoshihide; Carninci, Piero; Forrest, Alistair R R; Brown, Carolyn J; Wasserman, Wyeth W

2016-11-18

Sex differences in susceptibility and progression have been reported in numerous diseases. Female cells have two copies of the X chromosome with X-chromosome inactivation imparting mono-allelic gene silencing for dosage compensation. However, a subset of genes, named escapees, escape silencing and are transcribed bi-allelically resulting in sexual dimorphism. Here we conducted in silico analyses of the sexes using human datasets to gain perspectives into such regulation. We identified transcription start sites of escapees (escTSSs) based on higher transcription levels in female cells using FANTOM5 CAGE data. Significant over-representations of YY1 transcription factor binding motif and ChIP-seq peaks around escTSSs highlighted its positive association with escapees. Furthermore, YY1 occupancy is significantly biased towards the inactive X (Xi) at long non-coding RNA loci that are frequent contacts of Xi-specific superloops. Our study suggests a role for YY1 in transcriptional activity on Xi in general through sequence-specific binding, and its involvement at superloop anchors.

YY1 binding association with sex-biased transcription revealed through X-linked transcript levels and allelic binding analyses

PubMed Central

Chen, Chih-yu; Shi, Wenqiang; Balaton, Bradley P.; Matthews, Allison M.; Li, Yifeng; Arenillas, David J.; Mathelier, Anthony; Itoh, Masayoshi; Kawaji, Hideya; Lassmann, Timo; Hayashizaki, Yoshihide; Carninci, Piero; Forrest, Alistair R. R.; Brown, Carolyn J.; Wasserman, Wyeth W.

2016-01-01

Sex differences in susceptibility and progression have been reported in numerous diseases. Female cells have two copies of the X chromosome with X-chromosome inactivation imparting mono-allelic gene silencing for dosage compensation. However, a subset of genes, named escapees, escape silencing and are transcribed bi-allelically resulting in sexual dimorphism. Here we conducted in silico analyses of the sexes using human datasets to gain perspectives into such regulation. We identified transcription start sites of escapees (escTSSs) based on higher transcription levels in female cells using FANTOM5 CAGE data. Significant over-representations of YY1 transcription factor binding motif and ChIP-seq peaks around escTSSs highlighted its positive association with escapees. Furthermore, YY1 occupancy is significantly biased towards the inactive X (Xi) at long non-coding RNA loci that are frequent contacts of Xi-specific superloops. Our study suggests a role for YY1 in transcriptional activity on Xi in general through sequence-specific binding, and its involvement at superloop anchors. PMID:27857184

Biological significance of long non-coding RNA FTX expression in human colorectal cancer

PubMed Central

Guo, Xiao-Bo; Hua, Zhu; Li, Chen; Peng, Li-Pan; Wang, Jing-Shen; Wang, Bo; Zhi, Qiao-Ming

2015-01-01

The purpose of this study was to determine the expression of long non-coding RNA (lncRNA) FTX and analyze its prognostic and biological significance in colorectal cancer (CRC). A quantitative reverse transcription PCR was performed to detect the expression of long non-coding RNA FTX in 35 pairs of colorectal cancer and corresponding noncancerous tissues. The expression of long non-coding RNA FTX was detected in 187 colorectal cancer tissues and its correlations with clinicopathological factors of patients were examined. Univariate and multivariate analyses were performed to analyze the prognostic significance of Long Non-coding RNA FTX expression. The effects of long non-coding RNA FTX expression on malignant phenotypes of colorectal cancer cells and its possible biological significances were further determined. Long non-coding RNA FTX was significantly upregulated in colorectal cancer tissues, and low long non-coding RNA FTX expression was significantly correlated with differentiation grade, lymph vascular invasion, and clinical stage. Patients with high long non-coding RNA FTX showed poorer overall survival than those with low long non-coding RNA FTX. Multivariate analyses indicated that status of long non-coding RNA FTX was an independent prognostic factor for patients. Functional analyses showed that upregulation of long non-coding RNA FTX significantly promoted growth, migration, invasion, and increased colony formation in colorectal cancer cells. Therefore, long non-coding RNA FTX may be a potential biomarker for predicting the survival of colorectal cancer patients and might be a molecular target for treatment of human colorectal cancer. PMID:26629053

Variations in the non-coding transcriptome as a driver of inter-strain divergence and physiological adaptation in bacteria

PubMed Central

Kopf, Matthias; Klähn, Stephan; Scholz, Ingeborg; Hess, Wolfgang R.; Voß, Björn

2015-01-01

In all studied organisms, a substantial portion of the transcriptome consists of non-coding RNAs that frequently execute regulatory functions. Here, we have compared the primary transcriptomes of the cyanobacteria Synechocystis sp. PCC 6714 and PCC 6803 under 10 different conditions. These strains share 2854 protein-coding genes and a 16S rRNA identity of 99.4%, indicating their close relatedness. Conserved major transcriptional start sites (TSSs) give rise to non-coding transcripts within the sigB gene, from the 5′UTRs of cmpA and isiA, and 168 loci in antisense orientation. Distinct differences include single nucleotide polymorphisms rendering promoters inactive in one of the strains, e.g., for cmpR and for the asRNA PsbA2R. Based on the genome-wide mapped location, regulation and classification of TSSs, non-coding transcripts were identified as the most dynamic component of the transcriptome. We identified a class of mRNAs that originate by read-through from an sRNA that accumulates as a discrete and abundant transcript while also serving as the 5′UTR. Such an sRNA/mRNA structure, which we name ‘actuaton’, represents another way for bacteria to remodel their transcriptional network. Our findings support the hypothesis that variations in the non-coding transcriptome constitute a major evolutionary element of inter-strain divergence and capability for physiological adaptation. PMID:25902393

Microprocessor mediates transcriptional termination in long noncoding microRNA genes

PubMed Central

Dhir, Ashish; Dhir, Somdutta; Proudfoot, Nick J.; Jopling, Catherine L.

2015-01-01

MicroRNA (miRNA) play a major role in the post-transcriptional regulation of gene expression. Mammalian miRNA biogenesis begins with co-transcriptional cleavage of RNA polymerase II (Pol II) transcripts by the Microprocessor complex. While most miRNA are located within introns of protein coding genes, a substantial minority of miRNA originate from long non coding (lnc) RNA where transcript processing is largely uncharacterized. We show, by detailed characterization of liver-specific lnc-pri-miR-122 and genome-wide analysis in human cell lines, that most lnc-pri-miRNA do not use the canonical cleavage and polyadenylation (CPA) pathway, but instead use Microprocessor cleavage to terminate transcription. This Microprocessor inactivation leads to extensive transcriptional readthrough of lnc-pri-miRNA and transcriptional interference with downstream genes. Consequently we define a novel RNase III-mediated, polyadenylation-independent mechanism of Pol II transcription termination in mammalian cells. PMID:25730776

Snail1 transcription factor controls telomere transcription and integrity

PubMed Central

Mazzolini, Rocco; Gonzàlez, Núria; Garcia-Garijo, Andrea; Millanes-Romero, Alba; Peiró, Sandra; Smith, Susan

2018-01-01

Abstract Besides controlling epithelial-to-mesenchymal transition (EMT) and cell invasion, the Snail1 transcriptional factor also provides cells with cancer stem cell features. Since telomere maintenance is essential for stemness, we have examined the control of telomere integrity by Snail1. Fluorescence in situ hybridization (FISH) analysis indicates that Snail1-depleted mouse mesenchymal stem cells (MSC) have both a dramatic increase of telomere alterations and shorter telomeres. Remarkably, Snail1-deficient MSC present higher levels of both telomerase activity and the long non-coding RNA called telomeric repeat-containing RNA (TERRA), an RNA that controls telomere integrity. Accordingly, Snail1 expression downregulates expression of the telomerase gene (TERT) as well as of TERRA 2q, 11q and 18q. TERRA and TERT are transiently downregulated during TGFβ-induced EMT in NMuMG cells, correlating with Snail1 expression. Global transcriptome analysis indicates that ectopic expression of TERRA affects the transcription of some genes induced during EMT, such as fibronectin, whereas that of TERT does not modify those genes. We propose that Snail1 repression of TERRA is required not only for telomere maintenance but also for the expression of a subset of mesenchymal genes. PMID:29059385

The Unexpected Tuners: Are LncRNAs Regulating Host Translation during Infections?

PubMed Central

Knap, Primoz; Tebaldi, Toma; Di Leva, Francesca; Biagioli, Marta; Dalla Serra, Mauro; Viero, Gabriella

2017-01-01

Pathogenic bacteria produce powerful virulent factors, such as pore-forming toxins, that promote their survival and cause serious damage to the host. Host cells reply to membrane stresses and ionic imbalance by modifying gene expression at the epigenetic, transcriptional and translational level, to recover from the toxin attack. The fact that the majority of the human transcriptome encodes for non-coding RNAs (ncRNAs) raises the question: do host cells deploy non-coding transcripts to rapidly control the most energy-consuming process in cells—i.e., host translation—to counteract the infection? Here, we discuss the intriguing possibility that membrane-damaging toxins induce, in the host, the expression of toxin-specific long non-coding RNAs (lncRNAs), which act as sponges for other molecules, encoding small peptides or binding target mRNAs to depress their translation efficiency. Unravelling the function of host-produced lncRNAs upon bacterial infection or membrane damage requires an improved understanding of host lncRNA expression patterns, their association with polysomes and their function during this stress. This field of investigation holds a unique opportunity to reveal unpredicted scenarios and novel approaches to counteract antibiotic-resistant infections. PMID:29469820

Rapid evolution of cis-regulatory sequences via local point mutations

NASA Technical Reports Server (NTRS)

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

2001-01-01

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

Long non-coding RNA and Polycomb: an intricate partnership in cancer biology.

PubMed

Achour, Cyrinne; Aguilo, Francesca

2018-06-01

High-throughput analyses have revealed that the vast majority of the transcriptome does not code for proteins. These non-translated transcripts, when larger than 200 nucleotides, are termed long non-coding RNAs (lncRNAs), and play fundamental roles in diverse cellular processes. LncRNAs are subject to dynamic chemical modification, adding another layer of complexity to our understanding of the potential roles that lncRNAs play in health and disease. Many lncRNAs regulate transcriptional programs by influencing the epigenetic state through direct interactions with chromatin-modifying proteins. Among these proteins, Polycomb repressive complexes 1 and 2 (PRC1 and PRC2) have been shown to be recruited by lncRNAs to silence target genes. Aberrant expression, deficiency or mutation of both lncRNA and Polycomb have been associated with numerous human diseases, including cancer. In this review, we have highlighted recent findings regarding the concerted mechanism of action of Polycomb group proteins (PcG), acting together with some classically defined lncRNAs including X-inactive specific transcript ( XIST ), antisense non-coding RNA in the INK4 locus ( ANRIL ), metastasis associated lung adenocarcinoma transcript 1 ( MALAT1 ), and HOX transcript antisense RNA ( HOTAIR ).

Epigenetics of Peripheral B-Cell Differentiation and the Antibody Response

PubMed Central

Zan, Hong; Casali, Paolo

2015-01-01

Epigenetic modifications, such as histone post-translational modifications, DNA methylation, and alteration of gene expression by non-coding RNAs, including microRNAs (miRNAs) and long non-coding RNAs (lncRNAs), are heritable changes that are independent from the genomic DNA sequence. These regulate gene activities and, therefore, cellular functions. Epigenetic modifications act in concert with transcription factors and play critical roles in B cell development and differentiation, thereby modulating antibody responses to foreign- and self-antigens. Upon antigen encounter by mature B cells in the periphery, alterations of these lymphocytes epigenetic landscape are induced by the same stimuli that drive the antibody response. Such alterations instruct B cells to undergo immunoglobulin (Ig) class switch DNA recombination (CSR) and somatic hypermutation (SHM), as well as differentiation to memory B cells or long-lived plasma cells for the immune memory. Inducible histone modifications, together with DNA methylation and miRNAs modulate the transcriptome, particularly the expression of activation-induced cytidine deaminase, which is essential for CSR and SHM, and factors central to plasma cell differentiation, such as B lymphocyte-induced maturation protein-1. These inducible B cell-intrinsic epigenetic marks guide the maturation of antibody responses. Combinatorial histone modifications also function as histone codes to target CSR and, possibly, SHM machinery to the Ig loci by recruiting specific adaptors that can stabilize CSR/SHM factors. In addition, lncRNAs, such as recently reported lncRNA-CSR and an lncRNA generated through transcription of the S region that form G-quadruplex structures, are also important for CSR targeting. Epigenetic dysregulation in B cells, including the aberrant expression of non-coding RNAs and alterations of histone modifications and DNA methylation, can result in aberrant antibody responses to foreign antigens, such as those on microbial pathogens, and generation of pathogenic autoantibodies, IgE in allergic reactions, as well as B cell neoplasia. Epigenetic marks would be attractive targets for new therapeutics for autoimmune and allergic diseases, and B cell malignancies. PMID:26697022

Tudor-staphylococcal nuclease regulates the expression and biological function of alkylglycerone phosphate synthase via nuclear factor-κB and microRNA-127 in human glioma U87MG cells.

PubMed

Zhang, Yongqiang; Jia, Jun; Li, Ying; Chen, Yan-Ge; Huang, Huan; Qiao, Yang; Zhu, Yu

2018-06-01

Glioma is one of the malignant tumor types detrimental to human health; therefore, it is important to find novel targets and therapeutics for this tumor. The downregulated expression of Tudor-staphylococcal nuclease (SN) and alkylglycerone phosphate synthase (AGPS) can decrease cancer malignancy, and the overexpression of them can the increase viability and migration potential of various tumor cell types; however, the role of AGPS in the proliferation and migration of glioma, and the association of Tudor-SN and AGPS in human glioma is not clear. In the present study, it was determined that AGPS silencing suppressed the proliferation and migration potential of glioma U87MG cells, and suppressed the expression of the circular RNAs circ-ubiquitin-associated protein 2, circ-zinc finger protein 292 and circ-homeodomain-interacting protein kinase 3, and the long non-coding RNAs H19 imprinted maternally expressed transcript (non-protein coding), colon cancer-associated transcript 1 (non-protein coding) and hepatocellular carcinoma upregulated long non-coding RNA. Furthermore, Tudor-SN silencing suppressed the expression of AGPS; however, nuclear factor (NF)-κB and microRNA (miR)-127 retrieval experiments partially reduced the expression of AGPS. Additionally, it was determined that Tudor-SN silencing suppressed the activity of the mechanistic target of rapamycin (mTOR) signaling pathway, and NF-κB and miR-127 retrieval experiments partially reduced the activity of mTOR. Therefore, it was considered that NF-κB and miR-127 may be the mediators of Tudor-SN-regulated AGPS via the mTOR signaling pathway. These results improve on our knowledge of the mechanisms underlying Tudor-SN and AGPS in human glioma.

Cap 'n' collar C regulates genes responsible for imidacloprid resistance in the Colorado potato beetle, Leptinotarsa decemlineata.

PubMed

Gaddelapati, Sharath Chandra; Kalsi, Megha; Roy, Amit; Palli, Subba Reddy

2018-08-01

The Colorado potato beetle (CPB), Leptinotarsa decemlineata developed resistance to imidacloprid after exposure to this insecticide for multiple generations. Our previous studies showed that xenobiotic transcription factor, cap 'n' collar isoform C (CncC) regulates the expression of multiple cytochrome P450 genes, which play essential roles in resistance to plant allelochemicals and insecticides. In this study, we sought to obtain a comprehensive picture of the genes regulated by CncC in imidacloprid-resistant CPB. We performed sequencing of RNA isolated from imidacloprid-resistant CPB treated with dsRNA targeting CncC or gene coding for green fluorescent protein (control). Comparative transcriptome analysis showed that CncC regulated the expression of 1798 genes, out of which 1499 genes were downregulated in CncC knockdown beetles. Interestingly, expression of 79% of imidacloprid induced P450 genes requires CncC. We performed quantitative real-time PCR to verify the reduction in the expression of 20 genes including those coding for detoxification enzymes (P450s, glutathione S-transferases, and esterases) and ABC transporters. The genes coding for ABC transporters are induced in insecticide resistant CPB and require CncC for their expression. Knockdown of genes coding for ABC transporters simultaneously or individually caused an increase in imidacloprid-induced mortality in resistant beetles confirming their contribution to insecticide resistance. These studies identified CncC as a transcription factor involved in regulation of genes responsible for imidacloprid resistance. Small molecule inhibitors of CncC or suppression of CncC by RNAi could provide effective synergists for pest control or management of insecticide resistance. Copyright © 2018 Elsevier Ltd. All rights reserved.

Analysis of miRNAs targeting transcription factors in Persicaria minor induced by Fusarium oxysporum

NASA Astrophysics Data System (ADS)

Samad, Abdul Fatah A.; Ali, Nazaruddin Muhammad; Ismail, Ismanizan; Murad, Abdul Munir Abdul

2016-11-01

A recent discovery showed small non-coding RNA known as microRNA has a crucial role in plant development and plant survival in extreme condition. In the past few years, researchers have managed to identify the various families of transcription factors that play a crucial role in regulating plant development and plant responses to stresses. This study focuses on the expression pattern of miRNA targeted transcription factor under biotic stress in a plant rich with secondary metabolite, Persicaria minor. A pathogenic fungus, Fusarium oxysporum was used in the biotic stress treatment since the previous study revealed this fungus could trigger plant defense system. Two small RNA libraries were constructed which consist of control and treated samples. In order to identify the potential target, psRobot target prediction software was used for each miRNA that shows significant change due to the infection. The result showed miR156b/c, miR172a, miR319, miR858, and miR894 were found to be targeting a wide range of transcription factors that involve in plant development and plant response towards stresses. The expression of miR156b/c and miR172 were up-regulated while the expression of miR319, miR858, and miR894 was found to be down-regulated. These results may provide a certain level of networking between those two regulatory molecules in plant genetic system under biotic stress.

Intracellular Bioinorganic Chemistry and Cross Talk Among Different -Omics.

PubMed

Mendola, Diego La; Giacomelli, Chiara; Rizzarelli, Enrico

2016-01-01

The description of the cell life needs not only the knowledge of its genome and proteome, but also of the location of the metal ions and their different complex species in the subcellular compartments, that is of metallome. The cross-talk among these players of the omics' world secures the cellular homeostasis by means of a complex network, the alteration of which may give rise to many diseases. Copper and zinc ions levels regulate protein expression and metal-responsive transcription factors and in many pathologies metal dyshomeostasis induces to aberrant expression of different factors. microRNAs, a class of a small non-coding RNA molecules, act as RNA silencing and post-transcriptional regulators of gene expression contributing also to metal regulatory activity. The aim of the present review is to present how metals dyshomeostasis can be cause of diseases, involving different and specific metal chaperones, metal transporters, metalloproteins, small molecules and metal-sensing transcription factors. Two distinct classes of pathologies, cancer and osteoarthritis, are discussed starting from the metallostasis (metal homeostasis) and turning up to miRNAs regulation. The understanding of post-translational regulation, driven by metal ions sensing, may help to identify more specific targets and drugs to pathologies in which metal ions are involved.

Signaling coupled epigenomic regulation of gene expression.

PubMed

Kumar, R; Deivendran, S; Santhoshkumar, T R; Pillai, M R

2017-10-26

Inheritance of genomic information independent of the DNA sequence, the epigenetics, as well as gene transcription are profoundly shaped by serine/threonine and tyrosine signaling kinases and components of the chromatin remodeling complexes. To precisely respond to a changing external milieu, human cells efficiently translate upstream signals into post-translational modifications (PTMs) on histones and coregulators such as corepressors, coactivators, DNA-binding factors and PTM modifying enzymes. Because a protein with multiple residues for putative PTMs is expected to undergo more than one PTM in cells stimulated with growth factors, the outcome of combinational PTM codes on histones and coregulators is profoundly shaped by regulatory interplays between PTMs. The genomic functions of signaling kinases in cancer cells are manifested by the downstream effectors of cytoplasmic signaling cascades as well as translocation of the cytoplasmic signaling kinases to the nucleus. Signaling-mediated phosphorylation of histones serves as a regulatory switch for other PTMs, and connects chromatin remodeling complexes into gene transcription and gene activity. Here, we will discuss the recent advances in signaling-dependent epigenomic regulation of gene transcription using a few representative cancer-relevant serine/threonine and tyrosine kinases and their interplay with chromatin remodeling factors in cancer cells.

Connecting the dots: chromatin and alternative splicing in EMT

PubMed Central

Warns, Jessica A.; Davie, James R.; Dhasarathy, Archana

2015-01-01

Nature has devised sophisticated cellular machinery to process mRNA transcripts produced by RNA Polymerase II, removing intronic regions and connecting exons together, to produce mature RNAs. This process, known as splicing, is very closely linked to transcription. Alternative splicing, or the ability to produce different combinations of exons that are spliced together from the same genomic template, is a fundamental means of regulating protein complexity. Similar to transcription, both constitutive and alternative splicing can be regulated by chromatin and its associated factors in response to various signal transduction pathways activated by external stimuli. This regulation can vary between different cell types, and interference with these pathways can lead to changes in splicing, often resulting in aberrant cellular states and disease. The epithelial to mesenchymal transition (EMT), which leads to cancer metastasis, is influenced by alternative splicing events of chromatin remodelers and epigenetic factors such as DNA methylation and non-coding RNAs. In this review, we will discuss the role of epigenetic factors including chromatin, chromatin remodelers, DNA methyltransferases and microRNAs in the context of alternative splicing, and discuss their potential involvement in alternative splicing during the EMT process. PMID:26291837

The Transcriptome of Exophiala dermatitidis during Ex-vivo Skin Model Infection

PubMed Central

Poyntner, Caroline; Blasi, Barbara; Arcalis, Elsa; Mirastschijski, Ursula; Sterflinger, Katja; Tafer, Hakim

2016-01-01

The black yeast Exophiala dermatitidis is a widespread polyextremophile and human pathogen, that is found in extreme natural habitats and man-made environments such as dishwashers. It can cause various diseases ranging from phaeohyphomycosis and systemic infections, with fatality rates reaching 40%. While the number of cases in immunocompromised patients are increasing, knowledge of the infections, virulence factors and host response is still scarce. In this study, for the first time, an artificial infection of an ex-vivo skin model with Exophiala dermatitidis was monitored microscopically and transcriptomically. Results show that Exophiala dermatitidis is able to actively grow and penetrate the skin. The analysis of the genomic and RNA-sequencing data delivers a rich and complex transcriptome where circular RNAs, fusion transcripts, long non-coding RNAs and antisense transcripts are found. Changes in transcription strongly affect pathways related to nutrients acquisition, energy metabolism, cell wall, morphological switch, and known virulence factors. The L-Tyrosine melanin pathway is specifically upregulated during infection. Moreover the production of secondary metabolites, especially alkaloids, is increased. Our study is the first that gives an insight into the complexity of the transcriptome of Exophiala dermatitidis during artificial skin infections and reveals new virulence factors. PMID:27822460

Genomic organization of human fetal specific P-450IIIA7 (cytochrome P-450HFLa)-related gene(s) and interaction of transcriptional regulatory factor with its DNA element in the 5' flanking region.

PubMed

Itoh, S; Yanagimoto, T; Tagawa, S; Hashimoto, H; Kitamura, R; Nakajima, Y; Okochi, T; Fujimoto, S; Uchino, J; Kamataki, T

1992-03-24

P-450IIIA7 is a form of cytochrome P-450 which was isolated from human fetal livers and termed P-450HFLa. This form has been clarified to be expressed during fetal life specifically (Komori, M., Nishio, K., Kitada, M., Shiramatsu, K., Muroya, K., Soma, M., Nagashima, K. and Kamataki, T. (1990) Biochemistry 29, 4430-4433). In the present study, we isolated five independent clones which probably corresponded to the human P-450IIIA7 gene. These clones were completely sequenced, all exons, exon-intron junctions and the 5' flanking region from the cap site to-869. Although the sequences in the coding region were completely identical to P-450IIIA7, it is possible that genomic fragments sequenced in this study encode portions of other P-450IIIA7-related genes since we could not obtain a complete overlapping set of genomic clones. Within its 5' flanking sequence, the putative binding sites of several transcriptional regulatory factors existed. Among them, it was shown that a basic transcription element binding factor (BTEB) actually interacted with the 5' flanking region of this gene.

Small non coding RNAs in adipocyte biology and obesity.

PubMed

Amri, Ez-Zoubir; Scheideler, Marcel

2017-11-15

Obesity has reached epidemic proportions world-wide and constitutes a substantial risk factor for hypertension, type 2 diabetes, cardiovascular diseases and certain cancers. So far, regulation of energy intake by dietary and pharmacological treatments has met limited success. The main interest of current research is focused on understanding the role of different pathways involved in adipose tissue function and modulation of its mass. Whole-genome sequencing studies revealed that the majority of the human genome is transcribed, with thousands of non-protein-coding RNAs (ncRNA), which comprise small and long ncRNAs. ncRNAs regulate gene expression at the transcriptional and post-transcriptional level. Numerous studies described the involvement of ncRNAs in the pathogenesis of many diseases including obesity and associated metabolic disorders. ncRNAs represent potential diagnostic biomarkers and promising therapeutic targets. In this review, we focused on small ncRNAs involved in the formation and function of adipocytes and obesity. Copyright © 2017 Elsevier B.V. All rights reserved.

Brd2 gene disruption causes ‘metabolically healthy’ obesity: Epigenetic and chromatin-based mechanisms that uncouple obesity from Type 2 diabetes

PubMed Central

Wang, Fangnian; Deeney, Jude T.; Denis, Gerald V.

2014-01-01

Disturbed body energy balance can lead to obesity and obesity-driven diseases such as Type 2 diabetes, which have reached an epidemic level. Evidence indicates that obesity induced inflammation is a major cause of insulin resistance and Type 2 diabetes. Environmental factors, such as nutrients, affect body energy balance through epigenetic or chromatin-based mechanisms. As a bromodomain and external domain family transcription regulator, Brd2 regulates expression of many genes through interpretation of chromatin codes, and participates in the regulation of body energy balance and immune function. In the severely obese state, Brd2 knockdown in mice prevented obesity-induced inflammatory responses, protected animals from Type 2 diabetes, and thus uncoupled obesity from diabetes. Brd2 provides an important model for investigation of the function of transcription regulators and the development of obesity and diabetes; it also provides a possible target to treat obesity and diabetes through modulation of the function of a chromatin code reader. PMID:23374712

NRF2: Translating the Redox Code.

PubMed

Tummala, Krishna S; Kottakis, Filippos; Bardeesy, Nabeel

2016-10-01

Cancer requires mechanisms to mitigate reactive oxygen species (ROS) generated during rapid growth, such as induction of the antioxidant transcription factor, Nrf2. However, the targets of ROS-mediated cytotoxicity are unclear. Recent studies in pancreatic cancer show that redox control by Nrf2 prevents cysteine oxidation of the mRNA translational machinery, thereby supporting efficient protein synthesis. Copyright © 2016 Elsevier Ltd. All rights reserved.

Binding Sites Analyser (BiSA): Software for Genomic Binding Sites Archiving and Overlap Analysis

PubMed Central

Khushi, Matloob; Liddle, Christopher; Clarke, Christine L.; Graham, J. Dinny

2014-01-01

Genome-wide mapping of transcription factor binding and histone modification reveals complex patterns of interactions. Identifying overlaps in binding patterns by different factors is a major objective of genomic studies, but existing methods to archive large numbers of datasets in a personalised database lack sophistication and utility. Therefore we have developed transcription factor DNA binding site analyser software (BiSA), for archiving of binding regions and easy identification of overlap with or proximity to other regions of interest. Analysis results can be restricted by chromosome or base pair overlap between regions or maximum distance between binding peaks. BiSA is capable of reporting overlapping regions that share common base pairs; regions that are nearby; regions that are not overlapping; and average region sizes. BiSA can identify genes located near binding regions of interest, genomic features near a gene or locus of interest and statistical significance of overlapping regions can also be reported. Overlapping results can be visualized as Venn diagrams. A major strength of BiSA is that it is supported by a comprehensive database of publicly available transcription factor binding sites and histone modifications, which can be directly compared to user data. The documentation and source code are available on http://bisa.sourceforge.net PMID:24533055

In silico methods for co-transcriptional RNA secondary structure prediction and for investigating alternative RNA structure expression.

PubMed

Meyer, Irmtraud M

2017-05-01

RNA transcripts are the primary products of active genes in any living organism, including many viruses. Their cellular destiny not only depends on primary sequence signals, but can also be determined by RNA structure. Recent experimental evidence shows that many transcripts can be assigned more than a single functional RNA structure throughout their cellular life and that structure formation happens co-transcriptionally, i.e. as the transcript is synthesised in the cell. Moreover, functional RNA structures are not limited to non-coding transcripts, but can also feature in coding transcripts. The picture that now emerges is that RNA structures constitute an additional layer of information that can be encoded in any RNA transcript (and on top of other layers of information such as protein-context) in order to exert a wide range of functional roles. Moreover, different encoded RNA structures can be expressed at different stages of a transcript's life in order to alter the transcript's behaviour depending on its actual cellular context. Similar to the concept of alternative splicing for protein-coding genes, where a single transcript can yield different proteins depending on cellular context, it is thus appropriate to propose the notion of alternative RNA structure expression for any given transcript. This review introduces several computational strategies that my group developed to detect different aspects of RNA structure expression in vivo. Two aspects are of particular interest to us: (1) RNA secondary structure features that emerge during co-transcriptional folding and (2) functional RNA structure features that are expressed at different times of a transcript's life and potentially mutually exclusive. Copyright © 2017. Published by Elsevier Inc.

Sequence variations of the partially dominant DELLA gene Rht-B1c in wheat and their functional impacts

PubMed Central

Ma, Zhengqiang

2013-01-01

Rht-B1c, allelic to the DELLA protein-encoding gene Rht-B1a, is a natural mutation documented in common wheat (Triticum aestivum). It confers variation to a number of traits related to cell and plant morphology, seed dormancy, and photosynthesis. The present study was conducted to examine the sequence variations of Rht-B1c and their functional impacts. The results showed that Rht-B1c was partially dominant or co-dominant for plant height, and exhibited an increased dwarfing effect. At the sequence level, Rht-B1c differed from Rht-B1a by one 2kb Veju retrotransposon insertion, three coding region single nucleotide polymorphisms (SNPs), one 197bp insertion, and four SNPs in the 1kb upstream sequence. Haplotype investigations, association analyses, transient expression assays, and expression profiling showed that the Veju insertion was primarily responsible for the extreme dwarfing effect. It was found that the Veju insertion changed processing of the Rht-B1c transcripts and resulted in DELLA motif primary structure disruption. Expression assays showed that Rht-B1c caused reduction of total Rht-1 transcript levels, and up-regulation of GATA-like transcription factors and genes positively regulated by these factors, suggesting that one way in which Rht-1 proteins affect plant growth and development is through GATA-like transcription factor regulation. PMID:23918966

The Aspergillus fumigatus conidial melanin production is regulated by the bifunctional bHLH DevR and MADS-box RlmA transcription factors.

PubMed

Valiante, Vito; Baldin, Clara; Hortschansky, Peter; Jain, Radhika; Thywißen, Andreas; Straßburger, Maria; Shelest, Ekaterina; Heinekamp, Thorsten; Brakhage, Axel A

2016-10-01

Melanins play a crucial role in defending organisms against external stressors. In several pathogenic fungi, including the human pathogen Aspergillus fumigatus, melanin production was shown to contribute to virulence. A. fumigatus produces two different types of melanins, i.e., pyomelanin and dihydroxynaphthalene (DHN)-melanin. DHN-melanin forms the gray-green pigment characteristic for conidia, playing an important role in immune evasion of conidia and thus for fungal virulence. The DHN-melanin biosynthesis pathway is encoded by six genes organized in a cluster with the polyketide synthase gene pksP as a core element. Here, cross-species promoter analysis identified specific DNA binding sites in the DHN-melanin biosynthesis genes pksP-arp1 intergenic region that can be recognized by bHLH and MADS-box transcriptional regulators. Independent deletion of two genes coding for the transcription factors DevR (bHLH) and RlmA (MADS-box) interfered with sporulation and reduced the expression of the DHN-melanin gene cluster. In vitro and in vivo experiments proved that these transcription factors cooperatively regulate pksP expression acting both as repressors and activators in a mutually exclusive manner. The dual role executed by each regulator depends on specific DNA motifs recognized in the pksP promoter region. © 2016 John Wiley & Sons Ltd.

Effect of bar-code technology on the safety of medication administration.

PubMed

Poon, Eric G; Keohane, Carol A; Yoon, Catherine S; Ditmore, Matthew; Bane, Anne; Levtzion-Korach, Osnat; Moniz, Thomas; Rothschild, Jeffrey M; Kachalia, Allen B; Hayes, Judy; Churchill, William W; Lipsitz, Stuart; Whittemore, Anthony D; Bates, David W; Gandhi, Tejal K

2010-05-06

Serious medication errors are common in hospitals and often occur during order transcription or administration of medication. To help prevent such errors, technology has been developed to verify medications by incorporating bar-code verification technology within an electronic medication-administration system (bar-code eMAR). We conducted a before-and-after, quasi-experimental study in an academic medical center that was implementing the bar-code eMAR. We assessed rates of errors in order transcription and medication administration on units before and after implementation of the bar-code eMAR. Errors that involved early or late administration of medications were classified as timing errors and all others as nontiming errors. Two clinicians reviewed the errors to determine their potential to harm patients and classified those that could be harmful as potential adverse drug events. We observed 14,041 medication administrations and reviewed 3082 order transcriptions. Observers noted 776 nontiming errors in medication administration on units that did not use the bar-code eMAR (an 11.5% error rate) versus 495 such errors on units that did use it (a 6.8% error rate)--a 41.4% relative reduction in errors (P<0.001). The rate of potential adverse drug events (other than those associated with timing errors) fell from 3.1% without the use of the bar-code eMAR to 1.6% with its use, representing a 50.8% relative reduction (P<0.001). The rate of timing errors in medication administration fell by 27.3% (P<0.001), but the rate of potential adverse drug events associated with timing errors did not change significantly. Transcription errors occurred at a rate of 6.1% on units that did not use the bar-code eMAR but were completely eliminated on units that did use it. Use of the bar-code eMAR substantially reduced the rate of errors in order transcription and in medication administration as well as potential adverse drug events, although it did not eliminate such errors. Our data show that the bar-code eMAR is an important intervention to improve medication safety. (ClinicalTrials.gov number, NCT00243373.) 2010 Massachusetts Medical Society

BioCichlid: central dogma-based 3D visualization system of time-course microarray data on a hierarchical biological network.

PubMed

Ishiwata, Ryosuke R; Morioka, Masaki S; Ogishima, Soichi; Tanaka, Hiroshi

2009-02-15

BioCichlid is a 3D visualization system of time-course microarray data on molecular networks, aiming at interpretation of gene expression data by transcriptional relationships based on the central dogma with physical and genetic interactions. BioCichlid visualizes both physical (protein) and genetic (regulatory) network layers, and provides animation of time-course gene expression data on the genetic network layer. Transcriptional regulations are represented to bridge the physical network (transcription factors) and genetic network (regulated genes) layers, thus integrating promoter analysis into the pathway mapping. BioCichlid enhances the interpretation of microarray data and allows for revealing the underlying mechanisms causing differential gene expressions. BioCichlid is freely available and can be accessed at http://newton.tmd.ac.jp/. Source codes for both biocichlid server and client are also available.

Mediator complex dependent regulation of cardiac development and disease.

PubMed

Grueter, Chad E

2013-06-01

Cardiovascular disease (CVD) is a leading cause of morbidity and mortality. The risk factors for CVD include environmental and genetic components. Human mutations in genes involved in most aspects of cardiovascular function have been identified, many of which are involved in transcriptional regulation. The Mediator complex serves as a pivotal transcriptional regulator that functions to integrate diverse cellular signals by multiple mechanisms including recruiting RNA polymerase II, chromatin modifying proteins and non-coding RNAs to promoters in a context dependent manner. This review discusses components of the Mediator complex and the contribution of the Mediator complex to normal and pathological cardiac development and function. Enhanced understanding of the role of this core transcriptional regulatory complex in the heart will help us gain further insights into CVD. Copyright © 2013. Production and hosting by Elsevier Ltd.

Expression of Fushi tarazu factor 1 homolog and Pit-1 genes in the pituitaries of pre-spawning chum and sockeye salmon.

PubMed

Higa, M; Ando, H; Urano, A

2001-06-01

Fushi tarazu factor-1 (FTZ-F1) and Pit-1 are major pituitary transcription factors, controlling expression of genes coding for gonadotropin (GTH) subunits and growth hormone/prolactin/somatolactin family hormone, respectively. As a first step to investigate physiological factors regulating gene expression of these transcription factors, we determined their mRNA levels in the pituitaries of chum salmon (Oncorhynchus keta) at different stages of sexual maturation. FTZ-F1 gene expression was increased in males at the stage before spermiation, where the levels of GTH alpha and IIbeta subunit mRNAs were elevated. Pit-1 mRNA showed maximum levels at the final stage of sexual maturation in both sexes, when expression of somatolactin gene peaked. To clarify whether gonadotropin-releasing hormone (GnRH) is involved in these increases in FTZ-F1 and Pit-1 gene expression, we examined effects of GnRH analog (GnRHa) administration on their gene expression in maturing sockeye salmon (Oncorhynchus nerka). GnRHa stimulated Pit-1 gene expression in females only, but failed to stimulate FTZ-F1 gene expression in both sexes. The up-regulated expression of FTZ-F1 and Pit-1 genes at the pre-spawning stages suggest that the two transcription factors have roles in sexual maturation of salmonids. Physiological factors regulating gene expression of FTZ-F1 and Pit-1 are discussed in this review.

Hypoxia regulates alternative splicing of HIF and non-HIF target genes.

PubMed

Sena, Johnny A; Wang, Liyi; Heasley, Lynn E; Hu, Cheng-Jun

2014-09-01

Hypoxia is a common characteristic of many solid tumors. The hypoxic microenvironment stabilizes hypoxia-inducible transcription factor 1α (HIF1α) and 2α (HIF2α/EPAS1) to activate gene transcription, which promotes tumor cell survival. The majority of human genes are alternatively spliced, producing RNA isoforms that code for functionally distinct proteins. Thus, an effective hypoxia response requires increased HIF target gene expression as well as proper RNA splicing of these HIF-dependent transcripts. However, it is unclear if and how hypoxia regulates RNA splicing of HIF targets. This study determined the effects of hypoxia on alternative splicing (AS) of HIF and non-HIF target genes in hepatocellular carcinoma cells and characterized the role of HIF in regulating AS of HIF-induced genes. The results indicate that hypoxia generally promotes exon inclusion for hypoxia-induced, but reduces exon inclusion for hypoxia-reduced genes. Mechanistically, HIF activity, but not hypoxia per se is found to be necessary and sufficient to increase exon inclusion of several HIF targets, including pyruvate dehydrogenase kinase 1 (PDK1). PDK1 splicing reporters confirm that transcriptional activation by HIF is sufficient to increase exon inclusion of PDK1 splicing reporter. In contrast, transcriptional activation of a PDK1 minigene by other transcription factors in the absence of endogenous HIF target gene activation fails to alter PDK1 RNA splicing. This study demonstrates a novel function of HIF in regulating RNA splicing of HIF target genes. ©2014 American Association for Cancer Research.

Dynamic Modeling of GAIT System Reveals Transcriptome Expansion and Translational Trickle Control Device

PubMed Central

Yao, Peng; Potdar, Alka A.; Arif, Abul; Ray, Partho Sarothi; Mukhopadhyay, Rupak; Willard, Belinda; Xu, Yichi; Yan, Jun; Saidel, Gerald M.; Fox, Paul L.

2012-01-01

SUMMARY Post-transcriptional regulatory mechanisms superimpose “fine-tuning” control upon “on-off” switches characteristic of gene transcription. We have exploited computational modeling with experimental validation to resolve an anomalous relationship between mRNA expression and protein synthesis. Differential GAIT (Gamma-interferon Activated Inhibitor of Translation) complex activation repressed VEGF-A synthesis to a low, constant rate despite high, variable VEGFA mRNA expression. Dynamic model simulations indicated the presence of an unidentified, inhibitory GAIT element-interacting factor. We discovered a truncated form of glutamyl-prolyl tRNA synthetase (EPRS), the GAIT constituent that binds the 3’-UTR GAIT element in target transcripts. The truncated protein, EPRSN1, prevents binding of functional GAIT complex. EPRSN1 mRNA is generated by a remarkable polyadenylation-directed conversion of a Tyr codon in the EPRS coding sequence to a stop codon (PAY*). By low-level protection of GAIT element-bearing transcripts, EPRSN1 imposes a robust “translational trickle” of target protein expression. Genome-wide analysis shows PAY* generates multiple truncated transcripts thereby contributing to transcriptome expansion. PMID:22386318

Combinatorial influence of environmental parameters on transcription factor activity

PubMed Central

Knijnenburg, T.A.; Wessels, L.F.A.; Reinders, M.J.T.

2008-01-01

Motivation: Cells receive a wide variety of environmental signals, which are often processed combinatorially to generate specific genetic responses. Changes in transcript levels, as observed across different environmental conditions, can, to a large extent, be attributed to changes in the activity of transcription factors (TFs). However, in unraveling these transcription regulation networks, the actual environmental signals are often not incorporated into the model, simply because they have not been measured. The unquantified heterogeneity of the environmental parameters across microarray experiments frustrates regulatory network inference. Results: We propose an inference algorithm that models the influence of environmental parameters on gene expression. The approach is based on a yeast microarray compendium of chemostat steady-state experiments. Chemostat cultivation enables the accurate control and measurement of many of the key cultivation parameters, such as nutrient concentrations, growth rate and temperature. The observed transcript levels are explained by inferring the activity of TFs in response to combinations of cultivation parameters. The interplay between activated enhancers and repressors that bind a gene promoter determine the possible up- or downregulation of the gene. The model is translated into a linear integer optimization problem. The resulting regulatory network identifies the combinatorial effects of environmental parameters on TF activity and gene expression. Availability: The Matlab code is available from the authors upon request. Contact: t.a.knijnenburg@tudelft.nl Supplementary information: Supplementary data are available at Bioinformatics online. PMID:18586711

Gene expression profile of human Down syndrome leukocytes.

PubMed

Malagó, Wilson; Sommer, César A; Del Cistia Andrade, Camillo; Soares-Costa, Andrea; Abrao Possik, Patricia; Cassago, Alexandre; Santejo Silveira, Henrique C; Henrique-Silva, Flavio

2005-08-01

Identification of differences in the gene expression patterns of Down syndrome and normal leukocytes. We constructed the first Down syndrome leukocyte serial analysis of gene expression (SAGE) library from a 28 year-old patient. This library was analyzed and compared with a normal leukocyte SAGE library using the eSAGE software. Reverse transcriptase polymerase chain reaction (RT-PCR) was used to validate the results. We found that a large number of unidentified transcripts were overexpressed in Down syndrome leukocytes and some transcripts coding for growth factors (e.g. interleukin 8, IL-8), ribosomaproteins (e.g. L13a, L29, and L37), and transcription factors (e.g., Jun B, Jun D, and C/EBP beta) were underexpressed. The SAGE data were successfully validated for the genes IL-8, CXCR4, BCL2A1, L13a, L29, L37, and GTF3A using RT-PCR. Our analysis identified significant changes in the expression pattern of Down syndrome leukocytes compared with normal ones, including key regulators of growth and proliferation, ribosomal proteins, and a large number of overexpressed transcripts that were not matched in UniGene clusters and that may represent novel genes related to Down syndrome. This study offers a new insight into transcriptional changes in Down syndrome leukocytes and indicates candidate genes for further investigations into the molecular mechanism of Down syndrome pathology.

Context-Dependent Piano Music Transcription With Convolutional Sparse Coding

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cogliati, Andrea; Duan, Zhiyao; Wohlberg, Brendt

This study presents a novel approach to automatic transcription of piano music in a context-dependent setting. This approach employs convolutional sparse coding to approximate the music waveform as the summation of piano note waveforms (dictionary elements) convolved with their temporal activations (onset transcription). The piano note waveforms are pre-recorded for the specific piano to be transcribed in the specific environment. During transcription, the note waveforms are fixed and their temporal activations are estimated and post-processed to obtain the pitch and onset transcription. This approach works in the time domain, models temporal evolution of piano notes, and estimates pitches and onsetsmore » simultaneously in the same framework. Finally, experiments show that it significantly outperforms a state-of-the-art music transcription method trained in the same context-dependent setting, in both transcription accuracy and time precision, in various scenarios including synthetic, anechoic, noisy, and reverberant environments.« less

Context-Dependent Piano Music Transcription With Convolutional Sparse Coding

DOE PAGES

Cogliati, Andrea; Duan, Zhiyao; Wohlberg, Brendt

2016-08-04

This study presents a novel approach to automatic transcription of piano music in a context-dependent setting. This approach employs convolutional sparse coding to approximate the music waveform as the summation of piano note waveforms (dictionary elements) convolved with their temporal activations (onset transcription). The piano note waveforms are pre-recorded for the specific piano to be transcribed in the specific environment. During transcription, the note waveforms are fixed and their temporal activations are estimated and post-processed to obtain the pitch and onset transcription. This approach works in the time domain, models temporal evolution of piano notes, and estimates pitches and onsetsmore » simultaneously in the same framework. Finally, experiments show that it significantly outperforms a state-of-the-art music transcription method trained in the same context-dependent setting, in both transcription accuracy and time precision, in various scenarios including synthetic, anechoic, noisy, and reverberant environments.« less

The Snail repressor recruits EZH2 to specific genomic sites through the enrollment of the lncRNA HOTAIR in epithelial-to-mesenchymal transition

PubMed Central

Battistelli, C; Cicchini, C; Santangelo, L; Tramontano, A; Grassi, L; Gonzalez, F J; de Nonno, V; Grassi, G; Amicone, L; Tripodi, M

2017-01-01

The transcription factor Snail is a master regulator of cellular identity and epithelial-to-mesenchymal transition (EMT) directly repressing a broad repertoire of epithelial genes. How chromatin modifiers instrumental to its activity are recruited to Snail-specific binding sites is unclear. Here we report that the long non-coding RNA (lncRNA) HOTAIR (for HOX Transcript Antisense Intergenic RNA) mediates a physical interaction between Snail and enhancer of zeste homolog 2 (EZH2), an enzymatic subunit of the polycomb-repressive complex 2 and the main writer of chromatin-repressive marks. The Snail-repressive activity, here monitored on genes with a pivotal function in epithelial and hepatic morphogenesis, differentiation and cell-type identity, depends on the formation of a tripartite Snail/HOTAIR/EZH2 complex. These results demonstrate an lncRNA-mediated mechanism by which a transcriptional factor conveys a general chromatin modifier to specific genes, thereby allowing the execution of hepatocyte transdifferentiation; moreover, they highlight HOTAIR as a crucial player in the Snail-mediated EMT. PMID:27452518

Long Non-coding RNA, PANDA, Contributes to the Stabilization of p53 Tumor Suppressor Protein.

PubMed

Kotake, Yojiro; Kitagawa, Kyoko; Ohhata, Tatsuya; Sakai, Satoshi; Uchida, Chiharu; Niida, Hiroyuki; Naemura, Madoka; Kitagawa, Masatoshi

2016-04-01

P21-associated noncoding RNA DNA damage-activated (PANDA) is induced in response to DNA damage and represses apoptosis by inhibiting the function of nuclear transcription factor Y subunit alpha (NF-YA) transcription factor. Herein, we report that PANDA affects regulation of p53 tumor-suppressor protein. U2OS cells were transfected with PANDA siRNAs. At 72 h post-transfection, cells were subjected to immunoblotting and quantitative reverse transcription-polymerase chain reaction. Depletion of PANDA was associated with decreased levels of p53 protein, but not p53 mRNA. The stability of p53 protein was markedly reduced by PANDA silencing. Degradation of p53 protein by silencing PANDA was prevented by treatment of MG132, a proteasome inhibitor. Moreover, depletion of PANDA prevented accumulation of p53 protein, as a result of DNA damage, induced by the genotoxic agent etoposide. These results suggest that PANDA stabilizes p53 protein in response to DNA damage, and provide new insight into the regulatory mechanisms of p53. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

The G-Box Transcriptional Regulatory Code in Arabidopsis1[OPEN

PubMed Central

Shepherd, Samuel J.K.; Brestovitsky, Anna; Dickinson, Patrick; Biswas, Surojit

2017-01-01

Plants have significantly more transcription factor (TF) families than animals and fungi, and plant TF families tend to contain more genes; these expansions are linked to adaptation to environmental stressors. Many TF family members bind to similar or identical sequence motifs, such as G-boxes (CACGTG), so it is difficult to predict regulatory relationships. We determined that the flanking sequences near G-boxes help determine in vitro specificity but that this is insufficient to predict the transcription pattern of genes near G-boxes. Therefore, we constructed a gene regulatory network that identifies the set of bZIPs and bHLHs that are most predictive of the expression of genes downstream of perfect G-boxes. This network accurately predicts transcriptional patterns and reconstructs known regulatory subnetworks. Finally, we present Ara-BOX-cis (araboxcis.org), a Web site that provides interactive visualizations of the G-box regulatory network, a useful resource for generating predictions for gene regulatory relations. PMID:28864470

Differentially-Expressed Pseudogenes in HIV-1 Infection.

PubMed

Gupta, Aditi; Brown, C Titus; Zheng, Yong-Hui; Adami, Christoph

2015-09-29

Not all pseudogenes are transcriptionally silent as previously thought. Pseudogene transcripts, although not translated, contribute to the non-coding RNA pool of the cell that regulates the expression of other genes. Pseudogene transcripts can also directly compete with the parent gene transcripts for mRNA stability and other cell factors, modulating their expression levels. Tissue-specific and cancer-specific differential expression of these "functional" pseudogenes has been reported. To ascertain potential pseudogene:gene interactions in HIV-1 infection, we analyzed transcriptomes from infected and uninfected T-cells and found that 21 pseudogenes are differentially expressed in HIV-1 infection. This is interesting because parent genes of one-third of these differentially-expressed pseudogenes are implicated in HIV-1 life cycle, and parent genes of half of these pseudogenes are involved in different viral infections. Our bioinformatics analysis identifies candidate pseudogene:gene interactions that may be of significance in HIV-1 infection. Experimental validation of these interactions would establish that retroviruses exploit this newly-discovered layer of host gene expression regulation for their own benefit.

Transcription initiation complex structures elucidate DNA opening.

PubMed

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.

Widespread antisense transcription of Populus genome under drought.

PubMed

Yuan, Yinan; Chen, Su

2018-06-06

Antisense transcription is widespread in many genomes and plays important regulatory roles in gene expression. The objective of our study was to investigate the extent and functional relevance of antisense transcription in forest trees. We employed Populus, a model tree species, to probe the antisense transcriptional response of tree genome under drought, through stranded RNA-seq analysis. We detected nearly 48% of annotated Populus gene loci with antisense transcripts and 44% of them with co-transcription from both DNA strands. Global distribution of reads pattern across annotated gene regions uncovered that antisense transcription was enriched in untranslated regions while sense reads were predominantly mapped in coding exons. We further detected 1185 drought-responsive sense and antisense gene loci and identified a strong positive correlation between the expression of antisense and sense transcripts. Additionally, we assessed the antisense expression in introns and found a strong correlation between intronic expression and exonic expression, confirming antisense transcription of introns contributes to transcriptional activity of Populus genome under drought. Finally, we functionally characterized drought-responsive sense-antisense transcript pairs through gene ontology analysis and discovered that functional groups including transcription factors and histones were concordantly regulated at both sense and antisense transcriptional level. Overall, our study demonstrated the extensive occurrence of antisense transcripts of Populus genes under drought and provided insights into genome structure, regulation pattern and functional significance of drought-responsive antisense genes in forest trees. Datasets generated in this study serve as a foundation for future genetic analysis to improve our understanding of gene regulation by antisense transcription.

Constraints on the evolution of a doublesex target gene arising from doublesex’s pleiotropic deployment

PubMed Central

Luo, Shengzhan D.; Baker, Bruce S.

2015-01-01

“Regulatory evolution,” that is, changes in a gene’s expression pattern through changes at its regulatory sequence, rather than changes at the coding sequence of the gene or changes of the upstream transcription factors, has been increasingly recognized as a pervasive evolution mechanism. Many somatic sexually dimorphic features of Drosophila melanogaster are the results of gene expression regulated by the doublesex (dsx) gene, which encodes sex-specific transcription factors (DSXF in females and DSXM in males). Rapid changes in such sexually dimorphic features are likely a result of changes at the regulatory sequence of the target genes. We focused on the Flavin-containing monooxygenase-2 (Fmo-2) gene, a likely direct dsx target, to elucidate how sexually dimorphic expression and its evolution are brought about. We found that dsx is deployed to regulate the Fmo-2 transcription both in the midgut and in fat body cells of the spermatheca (a female-specific tissue), through a canonical DSX-binding site in the Fmo-2 regulatory sequence. In the melanogaster group, Fmo-2 transcription in the midgut has evolved rapidly, in contrast to the conserved spermathecal transcription. We identified two cis-regulatory modules (CRM-p and CRM-d) that direct sexually monomorphic or dimorphic Fmo-2 transcription, respectively, in the midguts of these species. Changes of Fmo-2 transcription in the midgut from sexually dimorphic to sexually monomorphic in some species are caused by the loss of CRM-d function, but not the loss of the canonical DSX-binding site. Thus, conferring transcriptional regulation on a CRM level allows the regulation to evolve rapidly in one tissue while evading evolutionary constraints posed by other tissues. PMID:25675536

PAR-CLIP data indicate that Nrd1-Nab3-dependent transcription termination regulates expression of hundreds of protein coding genes in yeast

PubMed Central

2014-01-01

Background Nrd1 and Nab3 are essential sequence-specific yeast RNA binding proteins that function as a heterodimer in the processing and degradation of diverse classes of RNAs. These proteins also regulate several mRNA coding genes; however, it remains unclear exactly what percentage of the mRNA component of the transcriptome these proteins control. To address this question, we used the pyCRAC software package developed in our laboratory to analyze CRAC and PAR-CLIP data for Nrd1-Nab3-RNA interactions. Results We generated high-resolution maps of Nrd1-Nab3-RNA interactions, from which we have uncovered hundreds of new Nrd1-Nab3 mRNA targets, representing between 20 and 30% of protein-coding transcripts. Although Nrd1 and Nab3 showed a preference for binding near 5′ ends of relatively short transcripts, they bound transcripts throughout coding sequences and 3′ UTRs. Moreover, our data for Nrd1-Nab3 binding to 3′ UTRs was consistent with a role for these proteins in the termination of transcription. Our data also support a tight integration of Nrd1-Nab3 with the nutrient response pathway. Finally, we provide experimental evidence for some of our predictions, using northern blot and RT-PCR assays. Conclusions Collectively, our data support the notion that Nrd1 and Nab3 function is tightly integrated with the nutrient response and indicate a role for these proteins in the regulation of many mRNA coding genes. Further, we provide evidence to support the hypothesis that Nrd1-Nab3 represents a failsafe termination mechanism in instances of readthrough transcription. PMID:24393166

Transcriptional mapping of the ribosomal RNA region of mouse L-cell mitochondrial DNA.

PubMed Central

Nagley, P; Clayton, D A

1980-01-01

The map positions in mouse mitochondrial DNA of the two ribosomal RNA genes and adjacent genes coding several small transcripts have been determined precisely by application of a procedure in which DNA-RNA hybrids have been subjected to digestion by S1 nuclease under conditions of varying severity. Digestion of the DNA-RNA hybrids with S1 nuclease yielded a series of species which were shown to contain ribosomal RNA molecules together with adjacent transcripts hybridized conjointly to a continuous segment of mitochondrial DNA. There is one small transcript about 60 bases long whose gene adjoins the sequences coding the 5'-end of the small ribosomal RNA (950 bases) and which lies approximately 200 nucleotides from the D-loop origin of heavy strand mitochondrial DNA synthesis. An 80-base transcript lies between the small and large ribosomal RNA genes, and genes for two further short transcript (each about 80 bases in length) abut the sequences coding the 3'-end of the large ribosomal RNA (approximately 1500 bases). The ability to isolate a discrete DNA-RNA hybrid species approximately 2700 base pairs in length containing all these transcripts suggests that there can be few nucleotides in this region of mouse mitochondrial DNA which are not represented as stable RNA species. Images PMID:6253898

Mediator directs co-transcriptional heterochromatin assembly by RNA interference-dependent and -independent pathways.

PubMed

Oya, Eriko; Kato, Hiroaki; Chikashige, Yuji; Tsutsumi, Chihiro; Hiraoka, Yasushi; Murakami, Yota

2013-01-01

Heterochromatin at the pericentromeric repeats in fission yeast is assembled and spread by an RNAi-dependent mechanism, which is coupled with the transcription of non-coding RNA from the repeats by RNA polymerase II. In addition, Rrp6, a component of the nuclear exosome, also contributes to heterochromatin assembly and is coupled with non-coding RNA transcription. The multi-subunit complex Mediator, which directs initiation of RNA polymerase II-dependent transcription, has recently been suggested to function after initiation in processes such as elongation of transcription and splicing. However, the role of Mediator in the regulation of chromatin structure is not well understood. We investigated the role of Mediator in pericentromeric heterochromatin formation and found that deletion of specific subunits of the head domain of Mediator compromised heterochromatin structure. The Mediator head domain was required for Rrp6-dependent heterochromatin nucleation at the pericentromere and for RNAi-dependent spreading of heterochromatin into the neighboring region. In the latter process, Mediator appeared to contribute to efficient processing of siRNA from transcribed non-coding RNA, which was required for efficient spreading of heterochromatin. Furthermore, the head domain directed efficient transcription in heterochromatin. These results reveal a pivotal role for Mediator in multiple steps of transcription-coupled formation of pericentromeric heterochromatin. This observation further extends the role of Mediator to co-transcriptional chromatin regulation.

Emerging Roles of Small Epstein-Barr Virus Derived Non-Coding RNAs in Epithelial Malignancy

PubMed Central

Lung, Raymond Wai-Ming; Tong, Joanna Hung-Man; To, Ka-Fai

2013-01-01

Latent Epstein-Barr virus (EBV) infection is an etiological factor in the progression of several human epithelial malignancies such as nasopharyngeal carcinoma (NPC) and a subset of gastric carcinoma. Reports have shown that EBV produces several viral oncoproteins, yet their pathological roles in carcinogenesis are not fully elucidated. Studies on the recently discovered of EBV-encoded microRNAs (ebv-miRNAs) showed that these small molecules function as post-transcriptional gene regulators and may play a role in the carcinogenesis process. In NPC and EBV positive gastric carcinoma (EBVaGC), 22 viral miRNAs which are located in the long alternative splicing EBV transcripts, named BamH1 A rightward transcripts (BARTs), are abundantly expressed. The importance of several miR-BARTs in carcinogenesis has recently been demonstrated. These novel findings enhance our understanding of the oncogenic properties of EBV and may lead to a more effective design of therapeutic regimens to combat EBV-associated malignancies. This article will review the pathological roles of miR-BARTs in modulating the expression of cancer-related genes in both host and viral genomes. The expression of other small non-coding RNAs in NPC and the expression pattern of miR-BARTs in rare EBV-associated epithelial cancers will also be discussed. PMID:23979421

Schizophyllum commune has an extensive and functional alternative splicing repertoire

PubMed Central

Gehrmann, Thies; Pelkmans, Jordi F.; Lugones, Luis G.; Wösten, Han A. B.; Abeel, Thomas; Reinders, Marcel J. T.

2016-01-01

Recent genome-wide studies have demonstrated that fungi possess the machinery to alternatively splice pre-mRNA. However, there has not been a systematic categorization of the functional impact of alternative splicing in a fungus. We investigate alternative splicing and its functional consequences in the model mushroom forming fungus Schizophyllum commune. Alternative splicing was demonstrated for 2,285 out of 12,988 expressed genes, resulting in 20% additional transcripts. Intron retentions were the most common alternative splicing events, accounting for 33% of all splicing events, and 43% of the events in coding regions. On the other hand, exon skipping events were rare in coding regions (1%) but enriched in UTRs where they accounted for 57% of the events. Specific functional groups, including transcription factors, contained alternatively spliced genes. Alternatively spliced transcripts were regulated differently throughout development in 19% of the 2,285 alternatively spliced genes. Notably, 69% of alternatively spliced genes have predicted alternative functionality by loss or gain of functional domains, or by acquiring alternative subcellular locations. S. commune exhibits more alternative splicing than any other studied fungus. Taken together, alternative splicing increases the complexity of the S. commune proteome considerably and provides it with a rich repertoire of alternative functionality that is exploited dynamically. PMID:27659065

Schizophyllum commune has an extensive and functional alternative splicing repertoire

DOE PAGES

Gehrmann, Thies; Pelkmans, Jordi F.; Lugones, Luis G.; ...

2016-09-23

Recent genome-wide studies have demonstrated that fungi possess the machinery to alternatively splice pre-mRNA. However, there has not been a systematic categorization of the functional impact of alternative splicing in a fungus. We investigate alternative splicing and its functional consequences in the model mushroom forming fungus Schizophyllum commune. Alternative splicing was demonstrated for 2,285 out of 12,988 expressed genes, resulting in 20% additional transcripts. Intron retentions were the most common alternative splicing events, accounting for 33% of all splicing events, and 43% of the events in coding regions. On the other hand, exon skipping events were rare in coding regionsmore » (1%) but enriched in UTRs where they accounted for 57% of the events. Specific functional groups, including transcription factors, contained alternatively spliced genes. Alternatively spliced transcripts were regulated differently throughout development in 19% of the 2,285 alternatively spliced genes. Notably, 69% of alternatively spliced genes have predicted alternative functionality by loss or gain of functional domains, or by acquiring alternative subcellular locations. S. commune exhibits more alternative splicing than any other studied fungus. Finally, taken together, alternative splicing increases the complexity of the S. commune proteome considerably and provides it with a rich repertoire of alternative functionality that is exploited dynamically.« less

RhoA Regulation of Cardiomyocyte Differentiation

PubMed Central

Kaarbø, Mari; Crane, Denis I.; Murrell, Wayne G.

2013-01-01

Earlier findings from our laboratory implicated RhoA in heart developmental processes. To investigate factors that potentially regulate RhoA expression, RhoA gene organisation and promoter activity were analysed. Comparative analysis indicated strict conservation of both gene organisation and coding sequence of the chick, mouse, and human RhoA genes. Bioinformatics analysis of the derived promoter region of mouse RhoA identified putative consensus sequence binding sites for several transcription factors involved in heart formation and organogenesis generally. Using luciferase reporter assays, RhoA promoter activity was shown to increase in mouse-derived P19CL6 cells that were induced to differentiate into cardiomyocytes. Overexpression of a dominant negative mutant of mouse RhoA (mRhoAN19) blocked this cardiomyocyte differentiation of P19CL6 cells and led to the accumulation of the cardiac transcription factors SRF and GATA4 and the early cardiac marker cardiac α-actin. Taken together, these findings indicate a fundamental role for RhoA in the differentiation of cardiomyocytes. PMID:23935420

A repertoire of the dominant transcripts from the salivary glands of the blood-sucking bug, Triatoma dimidiata, a vector of Chagas disease

PubMed Central

Kato, Hirotomo; Jochim, Ryan C.; Gomez, Eduardo A.; Sakoda, Ryo; Iwata, Hiroyuki; Valenzuela, Jesus G.; Hashiguchi, Yoshihisa

2010-01-01

Triatoma (T.) dimidiata is a hematophagous Hemiptera and a main vector of Chagas disease. The saliva of this and other blood-sucking insects contains potent pharmacologically active components that assist them in counteracting the host hemostatic and inflammatory systems during blood feeding. To describe the repertoire of potential bioactive salivary molecules from this insect, a number of randomly selected transcripts from the salivary gland cDNA library of T. dimidiata were sequenced and analyzed. This analysis showed that 77.5% of the isolated transcripts coded for putative secreted proteins, and 89.9% of these coded for variants of the lipocalin family proteins. The most abundant transcript was a homologue of procalin, the major allergen of T. protracta saliva, and contributed more than 50% of the transcripts coding for putative secreted proteins, suggesting that it may play an important role in the blood-feeding process. Other salivary transcripts encoding lipocalin family proteins had homology to triabin (a thrombin inhibitor), triafestin (an inhibitor of kallikrein–kinin system), pallidipin (an inhibitor of collagen-induced platelet aggregation) and others with unknown function. PMID:19900580

Problem-Solving Test: The Role of a Micro-RNA in the Regulation of "fos" Gene Expression

ERIC Educational Resources Information Center

Szeberenyi, Jozsef

2009-01-01

The "fos" proto-oncogene codes for a component of the AP1 transcription factor, an important regulator of gene expression and cell proliferation. Dysregulation of AP1 function may lead to the malignant transformation of the cell. The present test describes an experiment in which the role of a micro-RNA (miR-7b) in the regulation of "fos" gene…

Short Exogenous Peptides Regulate Expression of CLE, KNOX1, and GRF Family Genes in Nicotiana tabacum.

PubMed

Fedoreyeva, L I; Dilovarova, T A; Ashapkin, V V; Martirosyan, Yu Ts; Khavinson, V Kh; Kharchenko, P N; Vanyushin, B F

2017-04-01

Exogenous short biologically active peptides epitalon (Ala-Glu-Asp-Gly), bronchogen (Ala-Glu-Asp-Leu), and vilon (Lys-Glu) at concentrations 10 -7 -10 -9  M significantly influence growth, development, and differentiation of tobacco (Nicotiana tabacum) callus cultures. Epitalon and bronchogen, in particular, both increase growth of calluses and stimulate formation and growth of leaves in plant regenerants. Because the regulatory activity of the short peptides appears at low peptide concentrations, their action to some extent is like that of the activity of phytohormones, and it seems to have signaling character and epigenetic nature. The investigated peptides modulate in tobacco cells the expression of genes including genes responsible for tissue formation and cell differentiation. These peptides differently modulate expression of CLE family genes coding for known endogenous regulatory peptides, the KNOX1 genes (transcription factor genes) and GRF (growth regulatory factor) genes coding for respective DNA-binding proteins such as topoisomerases, nucleases, and others. Thus, at the level of transcription, plants have a system of short peptide regulation of formation of long-known peptide regulators of growth and development. The peptides studied here may be related to a new generation of plant growth regulators. They can be used in the experimental botany, plant molecular biology, biotechnology, and practical agronomy.

Role of Alternative Polyadenylation during Adipogenic Differentiation: An In Silico Approach

PubMed Central

Spangenberg, Lucía; Correa, Alejandro; Dallagiovanna, Bruno; Naya, Hugo

2013-01-01

Post-transcriptional regulation of stem cell differentiation is far from being completely understood. Changes in protein levels are not fully correlated with corresponding changes in mRNAs; the observed differences might be partially explained by post-transcriptional regulation mechanisms, such as alternative polyadenylation. This would involve changes in protein binding, transcript usage, miRNAs and other non-coding RNAs. In the present work we analyzed the distribution of alternative transcripts during adipogenic differentiation and the potential role of miRNAs in post-transcriptional regulation. Our in silico analysis suggests a modest, consistent, bias in 3′UTR lengths during differentiation enabling a fine-tuned transcript regulation via small non-coding RNAs. Including these effects in the analyses partially accounts for the observed discrepancies in relative abundance of protein and mRNA. PMID:24143171

Hepatic Long Intergenic Noncoding RNAs: High Promoter Conservation and Dynamic, Sex-Dependent Transcriptional Regulation by Growth Hormone

PubMed Central

Melia, Tisha; Hao, Pengying; Yilmaz, Feyza

2015-01-01

Long intergenic noncoding RNAs (lincRNAs) are increasingly recognized as key chromatin regulators, yet few studies have characterized lincRNAs in a single tissue under diverse conditions. Here, we analyzed 45 mouse liver RNA sequencing (RNA-Seq) data sets collected under diverse conditions to systematically characterize 4,961 liver lincRNAs, 59% of them novel, with regard to gene structures, species conservation, chromatin accessibility, transcription factor binding, and epigenetic states. To investigate the potential for functionality, we focused on the responses of the liver lincRNAs to growth hormone stimulation, which imparts clinically relevant sex differences to hepatic metabolism and liver disease susceptibility. Sex-biased expression characterized 247 liver lincRNAs, with many being nuclear RNA enriched and regulated by growth hormone. The sex-biased lincRNA genes are enriched for nearby and correspondingly sex-biased accessible chromatin regions, as well as sex-biased binding sites for growth hormone-regulated transcriptional activators (STAT5, hepatocyte nuclear factor 6 [HNF6], FOXA1, and FOXA2) and transcriptional repressors (CUX2 and BCL6). Repression of female-specific lincRNAs in male liver, but not that of male-specific lincRNAs in female liver, was associated with enrichment of H3K27me3-associated inactive states and poised (bivalent) enhancer states. Strikingly, we found that liver-specific lincRNA gene promoters are more highly species conserved and have a significantly higher frequency of proximal binding by liver transcription factors than liver-specific protein-coding gene promoters. Orthologs for many liver lincRNAs were identified in one or more supraprimates, including two rat lincRNAs showing the same growth hormone-regulated, sex-biased expression as their mouse counterparts. This integrative analysis of liver lincRNA chromatin states, transcription factor occupancy, and growth hormone regulation provides novel insights into the expression of sex-specific lincRNAs and their potential for regulation of sex differences in liver physiology and disease. PMID:26459762

Barriers Influencing Linkage to Hypertension Care in Kenya: Qualitative Analysis from the LARK Hypertension Study.

PubMed

Naanyu, Violet; Vedanthan, Rajesh; Kamano, Jemima H; Rotich, Jackson K; Lagat, Kennedy K; Kiptoo, Peninah; Kofler, Claire; Mutai, Kennedy K; Bloomfield, Gerald S; Menya, Diana; Kimaiyo, Sylvester; Fuster, Valentin; Horowitz, Carol R; Inui, Thomas S

2016-03-01

Hypertension, the leading global risk factor for mortality, is characterized by low treatment and control rates in low- and middle-income countries. Poor linkage to hypertension care contributes to poor outcomes for patients. However, specific factors influencing linkage to hypertension care are not well known. To evaluate factors influencing linkage to hypertension care in rural western Kenya. Qualitative research study using a modified Health Belief Model that incorporates the impact of emotional and environmental factors on behavior. Mabaraza (traditional community assembly) participants (n = 242) responded to an open invitation to residents in their respective communities. Focus groups, formed by purposive sampling, consisted of hypertensive individuals, at-large community members, and community health workers (n = 169). We performed content analysis of the transcripts with NVivo 10 software, using both deductive and inductive codes. We used a two-round Delphi method to rank the barriers identified in the content analysis. We selected factors using triangulation of frequency of codes and themes from the transcripts, in addition to the results of the Delphi exercise. Sociodemographic characteristics of participants were summarized using descriptive statistics. We identified 27 barriers to linkage to hypertension care, grouped into individual (cognitive and emotional) and environmental factors. Cognitive factors included the asymptomatic nature of hypertension and limited information. Emotional factors included fear of being a burden to the family and fear of being screened for stigmatized diseases such as HIV. Environmental factors were divided into physical (e.g. distance), socioeconomic (e.g. poverty), and health system factors (e.g. popularity of alternative therapies). The Delphi results were generally consistent with the findings from the content analysis. Individual and environmental factors are barriers to linkage to hypertension care in rural western Kenya. Our analysis provides new insights and methodological approaches that may be relevant to other low-resource settings worldwide.

Long Non-Coding RNAs: A Novel Paradigm for Toxicology

PubMed Central

Dempsey, Joseph L.; Cui, Julia Yue

2017-01-01

Long non-coding RNAs (lncRNAs) are over 200 nucleotides in length and are transcribed from the mammalian genome in a tissue-specific and developmentally regulated pattern. There is growing recognition that lncRNAs are novel biomarkers and/or key regulators of toxicological responses in humans and animal models. Lacking protein-coding capacity, the numerous types of lncRNAs possess a myriad of transcriptional regulatory functions that include cis and trans gene expression, transcription factor activity, chromatin remodeling, imprinting, and enhancer up-regulation. LncRNAs also influence mRNA processing, post-transcriptional regulation, and protein trafficking. Dysregulation of lncRNAs has been implicated in various human health outcomes such as various cancers, Alzheimer’s disease, cardiovascular disease, autoimmune diseases, as well as intermediary metabolism such as glucose, lipid, and bile acid homeostasis. Interestingly, emerging evidence in the literature over the past five years has shown that lncRNA regulation is impacted by exposures to various chemicals such as polycyclic aromatic hydrocarbons, benzene, cadmium, chlorpyrifos-methyl, bisphenol A, phthalates, phenols, and bile acids. Recent technological advancements, including next-generation sequencing technologies and novel computational algorithms, have enabled the profiling and functional characterizations of lncRNAs on a genomic scale. In this review, we summarize the biogenesis and general biological functions of lncRNAs, highlight the important roles of lncRNAs in human diseases and especially during the toxicological responses to various xenobiotics, evaluate current methods for identifying aberrant lncRNA expression and molecular target interactions, and discuss the potential to implement these tools to address fundamental questions in toxicology. PMID:27864543

microRNA in Cerebral Spinal Fluid as Biomarkers of Alzheimer’s Disease Risk After Brain Injury

DTIC Science & Technology

2016-08-01

protein processing is a key feature of AD. MiRNAs are small non- coding RNA that regulate mRNA transcription, and may be a significant cause of protein...non- coding RNA that regulate mRNA transcription, and may be a significant cause of protein dysregulation. Our investigative team has generated

The Cell Cycle Regulator CCDC6 Is a Key Target of RNA-Binding Protein EWS

PubMed Central

Duggimpudi, Sujitha; Larsson, Erik; Nabhani, Schafiq; Borkhardt, Arndt; Hoell, Jessica I

2015-01-01

Genetic translocation of EWSR1 to ETS transcription factor coding region is considered as primary cause for Ewing sarcoma. Previous studies focused on the biology of chimeric transcription factors formed due to this translocation. However, the physiological consequences of heterozygous EWSR1 loss in these tumors have largely remained elusive. Previously, we have identified various mRNAs bound to EWS using PAR-CLIP. In this study, we demonstrate CCDC6, a known cell cycle regulator protein, as a novel target regulated by EWS. siRNA mediated down regulation of EWS caused an elevated apoptosis in cells in a CCDC6-dependant manner. This effect was rescued upon re-expression of CCDC6. This study provides evidence for a novel functional link through which wild-type EWS operates in a target-dependant manner in Ewing sarcoma. PMID:25751255

Deciphering Mineral Homeostasis in Barley Seed Transfer Cells at Transcriptional Level

PubMed Central

Borg, Søren

2015-01-01

In addition to the micronutrient inadequacy of staple crops for optimal human nutrition, a global downtrend in crop-quality has emerged from intensive breeding for yield. This trend will be aggravated by elevated levels of the greenhouse gas carbon dioxide. Therefore, crop biofortification is inevitable to ensure a sustainable supply of minerals to the large part of human population who is dietary dependent on staple crops. This requires a thorough understanding of plant-mineral interactions due to the complexity of mineral homeostasis. Employing RNA sequencing, we here communicate transfer cell specific effects of excess iron and zinc during grain filling in our model crop plant barley. Responding to alterations in mineral contents, we found a long range of different genes and transcripts. Among them, it is worth to highlight the auxin and ethylene signaling factors Arfs, Abcbs, Cand1, Hps4, Hac1, Ecr1, and Ctr1, diurnal fluctuation components Sdg2, Imb1, Lip1, and PhyC, retroelements, sulfur homeostasis components Amp1, Hmt3, Eil3, and Vip1, mineral trafficking components Med16, Cnnm4, Aha2, Clpc1, and Pcbps, and vacuole organization factors Ymr155W, RabG3F, Vps4, and Cbl3. Our analysis introduces new interactors and signifies a broad spectrum of regulatory levels from chromatin remodeling to intracellular protein sorting mechanisms active in the plant mineral homeostasis. The results highlight the importance of storage proteins in metal ion toxicity-resistance and chelation. Interestingly, the protein sorting and recycling factors Exoc7, Cdc1, Sec23A, and Rab11A contributed to the response as well as the polar distributors of metal-transporters ensuring the directional flow of minerals. Alternative isoform switching was found important for plant adaptation and occurred among transcripts coding for identical proteins as well as transcripts coding for protein isoforms. We also identified differences in the alternative-isoform preference between the treatments, indicating metal-affinity shifts among isoforms of metal transporters. Most important, we found the zinc treatment to impair both photosynthesis and respiration. A wide range of transcriptional changes including stress-related genes and negative feedback loops emphasize the importance to withhold mineral contents below certain cellular levels which otherwise might lead to agronomical impeding side-effects. By illustrating new mechanisms, genes, and transcripts, this report provides a solid platform towards understanding the complex network of plant mineral homeostasis. PMID:26536247

Prediction of plant lncRNA by ensemble machine learning classifiers.

PubMed

Simopoulos, Caitlin M A; Weretilnyk, Elizabeth A; Golding, G Brian

2018-05-02

In plants, long non-protein coding RNAs are believed to have essential roles in development and stress responses. However, relative to advances on discerning biological roles for long non-protein coding RNAs in animal systems, this RNA class in plants is largely understudied. With comparatively few validated plant long non-coding RNAs, research on this potentially critical class of RNA is hindered by a lack of appropriate prediction tools and databases. Supervised learning models trained on data sets of mostly non-validated, non-coding transcripts have been previously used to identify this enigmatic RNA class with applications largely focused on animal systems. Our approach uses a training set comprised only of empirically validated long non-protein coding RNAs from plant, animal, and viral sources to predict and rank candidate long non-protein coding gene products for future functional validation. Individual stochastic gradient boosting and random forest classifiers trained on only empirically validated long non-protein coding RNAs were constructed. In order to use the strengths of multiple classifiers, we combined multiple models into a single stacking meta-learner. This ensemble approach benefits from the diversity of several learners to effectively identify putative plant long non-coding RNAs from transcript sequence features. When the predicted genes identified by the ensemble classifier were compared to those listed in GreeNC, an established plant long non-coding RNA database, overlap for predicted genes from Arabidopsis thaliana, Oryza sativa and Eutrema salsugineum ranged from 51 to 83% with the highest agreement in Eutrema salsugineum. Most of the highest ranking predictions from Arabidopsis thaliana were annotated as potential natural antisense genes, pseudogenes, transposable elements, or simply computationally predicted hypothetical protein. Due to the nature of this tool, the model can be updated as new long non-protein coding transcripts are identified and functionally verified. This ensemble classifier is an accurate tool that can be used to rank long non-protein coding RNA predictions for use in conjunction with gene expression studies. Selection of plant transcripts with a high potential for regulatory roles as long non-protein coding RNAs will advance research in the elucidation of long non-protein coding RNA function.

A purified truncated form of yeast Gal4 expressed in Escherichia coli and used to functionalize poly(lactic acid) nanoparticle surface is transcriptionally active in cellulo.

PubMed

Legaz, Sophie; Exposito, Jean-Yves; Borel, Agnès; Candusso, Marie-Pierre; Megy, Simon; Montserret, Roland; Lahaye, Vincent; Terzian, Christophe; Verrier, Bernard

2015-09-01

Gal4/UAS system is a powerful tool for the analysis of numerous biological processes. Gal4 is a large yeast transcription factor that activates genes including UAS sequences in their promoter. Here, we have synthesized a minimal form of Gal4 DNA sequence coding for the binding and dimerization regions, but also part of the transcriptional activation domain. This truncated Gal4 protein was expressed as inclusion bodies in Escherichia coli. A structured and active form of this recombinant protein was purified and used to cover poly(lactic acid) (PLA) nanoparticles. In cellulo, these Gal4-vehicles were able to activate the expression of a Green Fluorescent Protein (GFP) gene under the control of UAS sequences, demonstrating that the decorated Gal4 variant can be delivery into cells where it still retains its transcription factor capacities. Thus, we have produced in E. coli and purified a short active form of Gal4 that retains its functions at the surface of PLA-nanoparticles in cellular assay. These decorated Gal4-nanoparticles will be useful to decipher their tissue distribution and their potential after ingestion or injection in UAS-GFP recombinant animal models. Copyright © 2015 Elsevier Inc. All rights reserved.

Snail1 transcription factor controls telomere transcription and integrity.

PubMed

Mazzolini, Rocco; Gonzàlez, Núria; Garcia-Garijo, Andrea; Millanes-Romero, Alba; Peiró, Sandra; Smith, Susan; García de Herreros, Antonio; Canudas, Sílvia

2018-01-09

Besides controlling epithelial-to-mesenchymal transition (EMT) and cell invasion, the Snail1 transcriptional factor also provides cells with cancer stem cell features. Since telomere maintenance is essential for stemness, we have examined the control of telomere integrity by Snail1. Fluorescence in situ hybridization (FISH) analysis indicates that Snail1-depleted mouse mesenchymal stem cells (MSC) have both a dramatic increase of telomere alterations and shorter telomeres. Remarkably, Snail1-deficient MSC present higher levels of both telomerase activity and the long non-coding RNA called telomeric repeat-containing RNA (TERRA), an RNA that controls telomere integrity. Accordingly, Snail1 expression downregulates expression of the telomerase gene (TERT) as well as of TERRA 2q, 11q and 18q. TERRA and TERT are transiently downregulated during TGFβ-induced EMT in NMuMG cells, correlating with Snail1 expression. Global transcriptome analysis indicates that ectopic expression of TERRA affects the transcription of some genes induced during EMT, such as fibronectin, whereas that of TERT does not modify those genes. We propose that Snail1 repression of TERRA is required not only for telomere maintenance but also for the expression of a subset of mesenchymal genes. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

cDNA cloning and characterization of mouse DTEF-1 and ETF, members of the TEA/ATTS family of transcription factors.

PubMed

Yockey, C E; Shimizu, N

1998-02-01

Members of the TEA/ATTS family of transcription factors have been found in most representative eukaryotic organisms. In vertebrates, the TEA family contains at least four members, which share overlapping DNA-binding specificity and have similar transcriptional activation properties. In this article, we describe the cDNA cloning and characterization of the murine TEA proteins DTEF-1 (mDTEF-1) and ETF. Using in situ hybridization analysis of mouse embryos, we found that mDTEF-1 and ETF transcript distributions substantially overlap. ETF is expressed throughout the embryo except in the myocardium early in development, whereas late in development, it is enriched in lung and neuroectoderm. Mouse DTEF-1 is expressed at a much lower level throughout development and is substantially enriched in ectoderm and skin, as well as in the developing pituitary at midgestation. Northern blot analysis of adult mouse tissue total RNA showed that both ETF and mDTEF-1 are abundant in uterus and lung relative to other tissues. Using gel mobility shift assays and GAL4-fusion protein analysis, we demonstrated that the full coding sequences of ETF and mDTEF-1 encode M-CAT/GT-IIC-binding proteins containing activation domains.

The Mediator complex of Caenorhabditis elegans: insights into the developmental and physiological roles of a conserved transcriptional coregulator

PubMed Central

Grants, Jennifer M.; Goh, Grace Y. S.; Taubert, Stefan

2015-01-01

The Mediator multiprotein complex (‘Mediator’) is an important transcriptional coregulator that is evolutionarily conserved throughout eukaryotes. Although some Mediator subunits are essential for the transcription of all protein-coding genes, others influence the expression of only subsets of genes and participate selectively in cellular signaling pathways. Here, we review the current knowledge of Mediator subunit function in the nematode Caenorhabditis elegans, a metazoan in which established and emerging genetic technologies facilitate the study of developmental and physiological regulation in vivo. In this nematode, unbiased genetic screens have revealed critical roles for Mediator components in core developmental pathways such as epidermal growth factor (EGF) and Wnt/β-catenin signaling. More recently, important roles for C. elegans Mediator subunits have emerged in the regulation of lipid metabolism and of systemic stress responses, engaging conserved transcription factors such as nuclear hormone receptors (NHRs). We emphasize instances where similar functions for individual Mediator subunits exist in mammals, highlighting parallels between Mediator subunit action in nematode development and in human cancer biology. We also discuss a parallel between the association of the Mediator subunit MED12 with several human disorders and the role of its C. elegans ortholog mdt-12 as a regulatory hub that interacts with numerous signaling pathways. PMID:25634893

Transposon-driven transcription is a conserved feature of vertebrate spermatogenesis and transcript evolution.

PubMed

Davis, Matthew P; Carrieri, Claudia; Saini, Harpreet K; van Dongen, Stijn; Leonardi, Tommaso; Bussotti, Giovanni; Monahan, Jack M; Auchynnikava, Tania; Bitetti, Angelo; Rappsilber, Juri; Allshire, Robin C; Shkumatava, Alena; O'Carroll, Dónal; Enright, Anton J

2017-07-01

Spermatogenesis is associated with major and unique changes to chromosomes and chromatin. Here, we sought to understand the impact of these changes on spermatogenic transcriptomes. We show that long terminal repeats (LTRs) of specific mouse endogenous retroviruses (ERVs) drive the expression of many long non-coding transcripts (lncRNA). This process occurs post-mitotically predominantly in spermatocytes and round spermatids. We demonstrate that this transposon-driven lncRNA expression is a conserved feature of vertebrate spermatogenesis. We propose that transposon promoters are a mechanism by which the genome can explore novel transcriptional substrates, increasing evolutionary plasticity and allowing for the genesis of novel coding and non-coding genes. Accordingly, we show that a small fraction of these novel ERV-driven transcripts encode short open reading frames that produce detectable peptides. Finally, we find that distinct ERV elements from the same subfamilies act as differentially activated promoters in a tissue-specific context. In summary, we demonstrate that LTRs can act as tissue-specific promoters and contribute to post-mitotic spermatogenic transcriptome diversity. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

Transcription and translation of the sigG gene is tuned for proper execution of the switch from early to late gene expression in the developing Bacillus subtilis spore

PubMed Central

Mearls, Elizabeth B.; Jackter, Jacquelin; Colquhoun, Jennifer M.; Matthews, Allison J.; Fenton, Colleen

2018-01-01

A cascade of alternative sigma factors directs developmental gene expression during spore formation by the bacterium Bacillus subtilis. As the spore develops, a tightly regulated switch occurs in which the early-acting sigma factor σF is replaced by the late-acting sigma factor σG. The gene encoding σG (sigG) is transcribed by σF and by σG itself in an autoregulatory loop; yet σG activity is not detected until σF-dependent gene expression is complete. This separation in σF and σG activities has been suggested to be due at least in part to a poorly understood intercellular checkpoint pathway that delays sigG expression by σF. Here we report the results of a careful examination of sigG expression during sporulation. Unexpectedly, our findings argue against the existence of a regulatory mechanism to delay sigG transcription by σF and instead support a model in which sigG is transcribed by σF with normal timing, but at levels that are very low. This low-level expression of sigG is the consequence of several intrinsic features of the sigG regulatory and coding sequence—promoter spacing, secondary structure potential of the mRNA, and start codon identity—that dampen its transcription and translation. Especially notable is the presence of a conserved hairpin in the 5’ leader sequence of the sigG mRNA that occludes the ribosome-binding site, reducing translation by up to 4-fold. Finally, we demonstrate that misexpression of sigG from regulatory and coding sequences lacking these features triggers premature σG activity in the forespore during sporulation, as well as inappropriate σG activity during vegetative growth. Altogether, these data indicate that transcription and translation of the sigG gene is tuned to prevent vegetative expression of σG and to ensure the precise timing of the switch from σF to σG in the developing spore. PMID:29702640

Hypoxia-induced long non-coding RNA Malat1 is dispensable for renal ischemia/reperfusion-injury.

PubMed

Kölling, Malte; Genschel, Celina; Kaucsar, Tamas; Hübner, Anika; Rong, Song; Schmitt, Roland; Sörensen-Zender, Inga; Haddad, George; Kistler, Andreas; Seeger, Harald; Kielstein, Jan T; Fliser, Danilo; Haller, Hermann; Wüthrich, Rudolf; Zörnig, Martin; Thum, Thomas; Lorenzen, Johan

2018-02-21

Renal ischemia-reperfusion (I/R) injury is a major cause of acute kidney injury (AKI). Non-coding RNAs are crucially involved in its pathophysiology. We identified hypoxia-induced long non-coding RNA Malat1 (Metastasis Associated Lung Adenocarcinoma Transcript 1) to be upregulated in renal I/R injury. We here elucidated the functional role of Malat1 in vitro and its potential contribution to kidney injury in vivo. Malat1 was upregulated in kidney biopsies and plasma of patients with AKI, in murine hypoxic kidney tissue as well as in cultured and ex vivo sorted hypoxic endothelial cells and tubular epithelial cells. Malat1 was transcriptionally activated by hypoxia-inducible factor 1-α. In vitro, Malat1 inhibition reduced proliferation and the number of endothelial cells in the S-phase of the cell cycle. In vivo, Malat1 knockout and wildtype mice showed similar degrees of outer medullary tubular epithelial injury, proliferation, capillary rarefaction, inflammation and fibrosis, survival and kidney function. Small-RNA sequencing and whole genome expression analysis revealed only minor changes between ischemic Malat1 knockout and wildtype mice. Contrary to previous studies, which suggested a prominent role of Malat1 in the induction of disease, we did not confirm an in vivo role of Malat1 concerning renal I/R-injury.

Genome Analysis Reveals Interplay between 5′UTR Introns and Nuclear mRNA Export for Secretory and Mitochondrial Genes

PubMed Central

Cenik, Can; Chua, Hon Nian; Zhang, Hui; Tarnawsky, Stefan P.; Akef, Abdalla; Derti, Adnan; Tasan, Murat; Moore, Melissa J.; Palazzo, Alexander F.; Roth, Frederick P.

2011-01-01

In higher eukaryotes, messenger RNAs (mRNAs) are exported from the nucleus to the cytoplasm via factors deposited near the 5′ end of the transcript during splicing. The signal sequence coding region (SSCR) can support an alternative mRNA export (ALREX) pathway that does not require splicing. However, most SSCR–containing genes also have introns, so the interplay between these export mechanisms remains unclear. Here we support a model in which the furthest upstream element in a given transcript, be it an intron or an ALREX–promoting SSCR, dictates the mRNA export pathway used. We also experimentally demonstrate that nuclear-encoded mitochondrial genes can use the ALREX pathway. Thus, ALREX can also be supported by nucleotide signals within mitochondrial-targeting sequence coding regions (MSCRs). Finally, we identified and experimentally verified novel motifs associated with the ALREX pathway that are shared by both SSCRs and MSCRs. Our results show strong correlation between 5′ untranslated region (5′UTR) intron presence/absence and sequence features at the beginning of the coding region. They also suggest that genes encoding secretory and mitochondrial proteins share a common regulatory mechanism at the level of mRNA export. PMID:21533221

Genome-wide transcriptional profiling of human glioblastoma cells in response to ITE treatment.

PubMed

Kang, Bo; Zhou, Yanwen; Zheng, Min; Wang, Ying-Jie

2015-09-01

A ligand-activated transcription factor aryl hydrocarbon receptor (AhR) is recently revealed to play a key role in embryogenesis and tumorigenesis (Feng et al. [1], Safe et al. [2]) and 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) (Song et al. [3]) is an endogenous AhR ligand that possesses anti-tumor activity. In order to gain insights into how ITE acts via the AhR in embryogenesis and tumorigenesis, we analyzed the genome-wide transcriptional profiles of the following three groups of cells: the human glioblastoma U87 parental cells, U87 tumor sphere cells treated with vehicle (DMSO) and U87 tumor sphere cells treated with ITE. Here, we provide the details of the sample gathering strategy and show the quality controls and the analyses associated with our gene array data deposited into the Gene Expression Omnibus (GEO) under the accession code of GSE67986.

Regulation of nucleolus assembly by non-coding RNA polymerase II transcripts

PubMed Central

Caudron-Herger, Maïwen; Pankert, Teresa; Rippe, Karsten

2016-01-01

ABSTRACT The nucleolus is a nuclear subcompartment for tightly regulated rRNA production and ribosome subunit biogenesis. It also acts as a cellular stress sensor and can release enriched factors in response to cellular stimuli. Accordingly, the content and structure of the nucleolus change dynamically, which is particularly evident during cell cycle progression: the nucleolus completely disassembles during mitosis and reassembles in interphase. Although the mechanisms that drive nucleolar (re)organization have been the subject of a number of studies, they are only partly understood. Recently, we identified Alu element-containing RNA polymerase II transcripts (aluRNAs) as important for nucleolar structure and rRNA synthesis. Integrating these findings with studies on the liquid droplet-like nature of the nucleolus leads us to propose a model on how RNA polymerase II transcripts could regulate the assembly of the nucleolus in response to external stimuli and during cell cycle progression. PMID:27416361

Regulation of nucleolus assembly by non-coding RNA polymerase II transcripts.

PubMed

Caudron-Herger, Maïwen; Pankert, Teresa; Rippe, Karsten

2016-05-03

The nucleolus is a nuclear subcompartment for tightly regulated rRNA production and ribosome subunit biogenesis. It also acts as a cellular stress sensor and can release enriched factors in response to cellular stimuli. Accordingly, the content and structure of the nucleolus change dynamically, which is particularly evident during cell cycle progression: the nucleolus completely disassembles during mitosis and reassembles in interphase. Although the mechanisms that drive nucleolar (re)organization have been the subject of a number of studies, they are only partly understood. Recently, we identified Alu element-containing RNA polymerase II transcripts (aluRNAs) as important for nucleolar structure and rRNA synthesis. Integrating these findings with studies on the liquid droplet-like nature of the nucleolus leads us to propose a model on how RNA polymerase II transcripts could regulate the assembly of the nucleolus in response to external stimuli and during cell cycle progression.

Spinal neurons require Islet1 for subtype-specific differentiation of electrical excitability

PubMed Central

2014-01-01

Background In the spinal cord, stereotypic patterns of transcription factor expression uniquely identify neuronal subtypes. These transcription factors function combinatorially to regulate gene expression. Consequently, a single transcription factor may regulate divergent development programs by participation in different combinatorial codes. One such factor, the LIM-homeodomain transcription factor Islet1, is expressed in the vertebrate spinal cord. In mouse, chick and zebrafish, motor and sensory neurons require Islet1 for specification of biochemical and morphological signatures. Little is known, however, about the role that Islet1 might play for development of electrical membrane properties in vertebrates. Here we test for a role of Islet1 in differentiation of excitable membrane properties of zebrafish spinal neurons. Results We focus our studies on the role of Islet1 in two populations of early born zebrafish spinal neurons: ventral caudal primary motor neurons (CaPs) and dorsal sensory Rohon-Beard cells (RBs). We take advantage of transgenic lines that express green fluorescent protein (GFP) to identify CaPs, RBs and several classes of interneurons for electrophysiological study. Upon knock-down of Islet1, cells occupying CaP-like and RB-like positions continue to express GFP. With respect to voltage-dependent currents, CaP-like and RB-like neurons have novel repertoires that distinguish them from control CaPs and RBs, and, in some respects, resemble those of neighboring interneurons. The action potentials fired by CaP-like and RB-like neurons also have significantly different properties compared to those elicited from control CaPs and RBs. Conclusions Overall, our findings suggest that, for both ventral motor and dorsal sensory neurons, Islet1 directs differentiation programs that ultimately specify electrical membrane as well as morphological properties that act together to sculpt neuron identity. PMID:25149090

Expression of drought tolerance genes in tropical upland rice cultivars (Oryza sativa).

PubMed

Silveira, R D D; Abreu, F R M; Mamidi, S; McClean, P E; Vianello, R P; Lanna, A C; Carneiro, N P; Brondani, C

2015-07-27

Gene expression related to drought response in the leaf tissues of two Brazilian upland cultivars, the drought-tolerant Douradão and the drought-sensitive Primavera, was analyzed. RNA-seq identified 27,618 transcripts in the Douradão cultivar, with 24,090 (87.2%) homologous to the rice database, and 27,221 transcripts in the Primavera cultivar, with 23,663 (86.9%) homologous to the rice database. Gene-expression analysis between control and water-deficient treatments revealed 493 and 1154 differentially expressed genes in Douradão and Primavera cultivars, respectively. Genes exclusively expressed under drought were identified for Douradão, including two genes of particular interest coding for the protein peroxidase precursor, which is involved in three distinct metabolic pathways. Comparisons between the two drought-exposed cultivars revealed 2314 genes were differentially expressed (978 upregulated, 1336 downregulated in Douradão). Six genes distributed across 4 different transcription factor families (bHLH, MYB, NAC, and WRKY) were identified, all of which were upregulated in Douradão compared to Primavera during drought. Most of the genes identified in Douradão activate metabolic pathways responsible for production of secondary metabolites and genes coding for enzymatically active signaling receptors. Quantitative PCR validation showed that most gene expression was in agreement with computational prediction of these transcripts. The transcripts identified here will define molecular markers for identification of Cis-acting elements to search for allelic variants of these genes through analysis of polymorphic SNPs in GenBank accessions of upland rice, aiming to develop cultivars with the best combination of these alleles, resulting in materials with high yield potential in the event of drought during the reproductive phase.

Transcriptomes of six mutants in the Sen1 pathway reveal combinatorial control of transcription termination across the Saccharomyces cerevisiae genome

PubMed Central

Carver, Melissa N.; Müller, Ulrika; Bekiranov, Stefan; Auble, David T.

2017-01-01

Transcriptome studies on eukaryotic cells have revealed an unexpected abundance and diversity of noncoding RNAs synthesized by RNA polymerase II (Pol II), some of which influence the expression of protein-coding genes. Yet, much less is known about biogenesis of Pol II non-coding RNA than mRNAs. In the budding yeast Saccharomyces cerevisiae, initiation of non-coding transcripts by Pol II appears to be similar to that of mRNAs, but a distinct pathway is utilized for termination of most non-coding RNAs: the Sen1-dependent or “NNS” pathway. Here, we examine the effect on the S. cerevisiae transcriptome of conditional mutations in the genes encoding six different essential proteins that influence Sen1-dependent termination: Sen1, Nrd1, Nab3, Ssu72, Rpb11, and Hrp1. We observe surprisingly diverse effects on transcript abundance for the different proteins that cannot be explained simply by differing severity of the mutations. Rather, we infer from our results that termination of Pol II transcription of non-coding RNA genes is subject to complex combinatorial control that likely involves proteins beyond those studied here. Furthermore, we identify new targets and functions of Sen1-dependent termination, including a role in repression of meiotic genes in vegetative cells. In combination with other recent whole-genome studies on termination of non-coding RNAs, our results provide promising directions for further investigation. PMID:28665995

Identification of novel non-coding small RNAs from Streptococcus pneumoniae TIGR4 using high-resolution genome tiling arrays

PubMed Central

2010-01-01

Background The identification of non-coding transcripts in human, mouse, and Escherichia coli has revealed their widespread occurrence and functional importance in both eukaryotic and prokaryotic life. In prokaryotes, studies have shown that non-coding transcripts participate in a broad range of cellular functions like gene regulation, stress and virulence. However, very little is known about non-coding transcripts in Streptococcus pneumoniae (pneumococcus), an obligate human respiratory pathogen responsible for significant worldwide morbidity and mortality. Tiling microarrays enable genome wide mRNA profiling as well as identification of novel transcripts at a high-resolution. Results Here, we describe a high-resolution transcription map of the S. pneumoniae clinical isolate TIGR4 using genomic tiling arrays. Our results indicate that approximately 66% of the genome is expressed under our experimental conditions. We identified a total of 50 non-coding small RNAs (sRNAs) from the intergenic regions, of which 36 had no predicted function. Half of the identified sRNA sequences were found to be unique to S. pneumoniae genome. We identified eight overrepresented sequence motifs among sRNA sequences that correspond to sRNAs in different functional categories. Tiling arrays also identified approximately 202 operon structures in the genome. Conclusions In summary, the pneumococcal operon structures and novel sRNAs identified in this study enhance our understanding of the complexity and extent of the pneumococcal 'expressed' genome. Furthermore, the results of this study open up new avenues of research for understanding the complex RNA regulatory network governing S. pneumoniae physiology and virulence. PMID:20525227

Human dental pulp stem cells cultured in serum-free supplemented medium

PubMed Central

Bonnamain, Virginie; Thinard, Reynald; Sergent-Tanguy, Solène; Huet, Pascal; Bienvenu, Géraldine; Naveilhan, Philippe; Farges, Jean-Christophe; Alliot-Licht, Brigitte

2013-01-01

Growing evidence show that human dental pulp stem cells (DPSCs) could provide a source of adult stem cells for the treatment of neurodegenerative pathologies. In this study, DPSCs were expanded and cultured with a protocol generally used for the culture of neural stem/progenitor cells. Methodology: DPSC cultures were established from third molars. The pulp tissue was enzymatically digested and cultured in serum-supplemented basal medium for 12 h. Adherent (ADH) and non-adherent (non-ADH) cell populations were separated according to their differential adhesion to plastic and then cultured in serum-free defined N2 medium with epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF). Both ADH and non-ADH populations were analyzed by FACS and/or PCR. Results: FACS analysis of ADH-DPSCs revealed the expression of the mesenchymal cell marker CD90, the neuronal marker CD56, the transferrin receptor CD71, and the chemokine receptor CXCR3, whereas hematopoietic stem cells markers CD45, CD133, and CD34 were not expressed. ADH-DPSCs expressed transcripts coding for the Nestin gene, whereas expression levels of genes coding for the neuronal markers β-III tubulin and NF-M, and the oligodendrocyte marker PLP-1 were donor dependent. ADH-DPSCs did not express the transcripts for GFAP, an astrocyte marker. Cells of the non-ADH population that grew as spheroids expressed Nestin, β-III tubulin, NF-M and PLP-1 transcripts. DPSCs that migrated out of the spheroids exhibited an odontoblast-like morphology and expressed a higher level of DSPP and osteocalcin transcripts than ADH-DPSCs. Conclusion: Collectively, these data indicate that human DPSCs can be expanded and cultured in serum-free supplemented medium with EGF and bFGF. ADH-DPSCs and non-ADH populations contained neuronal and/or oligodendrocyte progenitors at different stages of commitment and, interestingly, cells from spheroid structures seem to be more engaged into the odontoblastic lineage than the ADH-DPSCs. PMID:24376422

The Intersection of the Extrinsic Hedgehog and WNT/Wingless Signals with the Intrinsic Hox Code Underpins Branching Pattern and Tube Shape Diversity in the Drosophila Airways

PubMed Central

Matsuda, Ryo; Hosono, Chie; Saigo, Kaoru; Samakovlis, Christos

2015-01-01

The tubular networks of the Drosophila respiratory system and our vasculature show distinct branching patterns and tube shapes in different body regions. These local variations are crucial for organ function and organismal fitness. Organotypic patterns and tube geometries in branched networks are typically controlled by variations of extrinsic signaling but the impact of intrinsic factors on branch patterns and shapes is not well explored. Here, we show that the intersection of extrinsic hedgehog(hh) and WNT/wingless (wg) signaling with the tube-intrinsic Hox code of distinct segments specifies the tube pattern and shape of the Drosophila airways. In the cephalic part of the airways, hh signaling induces expression of the transcription factor (TF) knirps (kni) in the anterior dorsal trunk (DTa1). kni represses the expression of another TF spalt major (salm), making DTa1 a narrow and long tube. In DTa branches of more posterior metameres, Bithorax Complex (BX-C) Hox genes autonomously divert hh signaling from inducing kni, thereby allowing DTa branches to develop as salm-dependent thick and short tubes. Moreover, the differential expression of BX-C genes is partly responsible for the anterior-to-posterior gradual increase of the DT tube diameter through regulating the expression level of Salm, a transcriptional target of WNT/wg signaling. Thus, our results highlight how tube intrinsic differential competence can diversify tube morphology without changing availabilities of extrinsic factors. PMID:25615601

Regulatory effects of cotranscriptional RNA structure formation and transitions.

PubMed

Liu, Sheng-Rui; Hu, Chun-Gen; Zhang, Jin-Zhi

2016-09-01

RNAs, which play significant roles in many fundamental biological processes of life, fold into sophisticated and precise structures. RNA folding is a dynamic and intricate process, which conformation transition of coding and noncoding RNAs form the primary elements of genetic regulation. The cellular environment contains various intrinsic and extrinsic factors that potentially affect RNA folding in vivo, and experimental and theoretical evidence increasingly indicates that the highly flexible features of the RNA structure are affected by these factors, which include the flanking sequence context, physiochemical conditions, cis RNA-RNA interactions, and RNA interactions with other molecules. Furthermore, distinct RNA structures have been identified that govern almost all steps of biological processes in cells, including transcriptional activation and termination, transcriptional mutagenesis, 5'-capping, splicing, 3'-polyadenylation, mRNA export and localization, and translation. Here, we briefly summarize the dynamic and complex features of RNA folding along with a wide variety of intrinsic and extrinsic factors that affect RNA folding. We then provide several examples to elaborate RNA structure-mediated regulation at the transcriptional and posttranscriptional levels. Finally, we illustrate the regulatory roles of RNA structure and discuss advances pertaining to RNA structure in plants. WIREs RNA 2016, 7:562-574. doi: 10.1002/wrna.1350 For further resources related to this article, please visit the WIREs website. © 2016 Wiley Periodicals, Inc.

Long non-coding RNA colon cancer-associated transcript 1 functions as a competing endogenous RNA to regulate cyclin-dependent kinase 1 expression by sponging miR-490-3p in hepatocellular carcinoma progression.

PubMed

Dou, Chunqing; Sun, Liyuan; Jin, Xin; Han, Mingming; Zhang, Bao; Li, Tao

2017-04-01

Hepatocellular carcinoma is an aggressive neoplasm and is one of the most common human cancers. Recently, long non-coding RNAs have been demonstrated to participate in pathogenesis of many diseases including the progression in several cancers. In this study, we found that the long non-coding RNA colon cancer-associated transcript 1 was upregulated in hepatocellular carcinoma tissues (p < 0.05), and high colon cancer-associated transcript 1 expression level was positively associated with tumor volume (p < 0.05) and American Joint Committee on Cancer stage (p < 0.05) in hepatocellular carcinoma patients. Luciferase reporter assays and RNA-pulldown assays showed that colon cancer-associated transcript 1 is a target of miR-490-3p. Real-time quantitative polymerase chain reaction and Western blot analysis indicated that colon cancer-associated transcript 1 regulated cyclin-dependent kinase 1 expression as a competing endogenous RNA by sponging miR-490-3p in hepatocellular carcinoma cells. Furthermore, colon cancer-associated transcript 1 silencing decreased hepatocellular carcinoma cells proliferation and invasion and overexpression promoted cell proliferation and invasion in vitro. These data demonstrated that the colon cancer-associated transcript 1/miR-490-3p/cyclin-dependent kinase 1 regulatory pathway promotes the progression of hepatocellular carcinoma. Inhibition of colon cancer-associated transcript 1 expression may be a novel therapeutic strategy for hepatocellular carcinoma.

The pineapple AcMADS1 promoter confers high level expression in tomato and arabidopsis flowering and fruiting tissues, but AcMADS1 does not complement the tomato LeMADS-RIN (rin) mutant

USDA-ARS?s Scientific Manuscript database

A previous EST study identified a MADS box transcription factor coding sequence, AcMADS1, that is strongly induced during non-climacteric pineapple fruit ripening. Phylogenetic analyses place the AcMADS1 protein in the same superclade as LeMADS-RIN, a master regulator of fruit ripening upstream of e...

Nuclear cereblon modulates transcriptional activity of Ikaros and regulates its downstream target, enkephalin, in human neuroblastoma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wada, Takeyoshi; Asahi, Toru; Research Organization for Nano & Life Innovation, Waseda University #03C309, TWIns, 2-2 Wakamatsu, Shinjuku, Tokyo, 162-8480

2016-08-26

The gene coding cereblon (CRBN) was originally identified in genetic linkage analysis of mild autosomal recessive nonsyndromic intellectual disability. CRBN has broad localization in both the cytoplasm and nucleus. However, the significance of nuclear CRBN remains unknown. In the present study, we aimed to elucidate the role of CRBN in the nucleus. First, we generated a series of CRBN deletion mutants and determined the regions responsible for the nuclear localization. Only CRBN protein lacking the N-terminal region was localized outside of the nucleus, suggesting that the N-terminal region is important for its nuclear localization. CRBN was also identified as amore » thalidomide-binding protein and component of the cullin-4-containing E3 ubiquitin ligase complex. Thalidomide has been reported to be involved in the regulation of the transcription factor Ikaros by CRBN-mediated degradation. To investigate the nuclear functions of CRBN, we performed co-immunoprecipitation experiments and evaluated the binding of CRBN to Ikaros. As a result, we found that CRBN was associated with Ikaros protein, and the N-terminal region of CRBN was required for Ikaros binding. In luciferase reporter gene experiments, CRBN modulated transcriptional activity of Ikaros. Furthermore, we found that CRBN modulated Ikaros-mediated transcriptional repression of the proenkephalin gene by binding to its promoter region. These results suggest that CRBN binds to Ikaros via its N-terminal region and regulates transcriptional activities of Ikaros and its downstream target, enkephalin. - Highlights: • We found that CRBN is a nucleocytoplasmic shutting protein and identified the key domain for nucleocytoplasmic shuttling. • CRBN associates with the transcription factor Ikaros via the N-terminal domain. • CRBN modulates Ikaros-mediated transcriptional regulation and its downstream target, enkephalin.« less

Molecular cloning and identification of the transcriptional regulatory domain of the goat neurokinin B gene TAC3.

PubMed

Suetomi, Yuta; Matsuda, Fuko; Uenoyama, Yoshihisa; Maeda, Kei-ichiro; Tsukamura, Hiroko; Ohkura, Satoshi

2013-10-01

Neurokinin B (NKB), encoded by TAC3, is thought to be an important accelerator of pulsatile gonadotropin-releasing hormone release. This study aimed to clarify the transcriptional regulatory mechanism of goat TAC3. First, we determined the full-length mRNA sequence of goat TAC3 from the hypothalamus to be 820 b, including a 381 b coding region, with the putative transcription start site located 143-b upstream of the start codon. The deduced amino acid sequence of NKB, which is produced from preproNKB, was completely conserved among goat, cattle, and human. Next, we cloned 5'-upstream region of goat TAC3 up to 3400 b from the translation initiation site, and this region was highly homologous with cattle TAC3 (89%). We used this goat TAC3 5'-upstream region to perform luciferase assays. We created a luciferase reporter vector containing DNA constructs from -2706, -1837, -834, -335, or -197 to +166 bp (the putative transcription start site was designated as +1) of goat TAC3 and these were transiently transfected into mouse hypothalamus-derived N7 cells and human neuroblastoma-derived SK-N-AS cells. The luciferase activity gradually increased with the deletion of the 5'-upstream region, suggesting that the transcriptional suppressive region is located between -2706 and -336 bp and that the core promoter exists downstream of -197 bp. Estradiol treatment did not lead to significant suppression of luciferase activity of any constructs, suggesting the existence of other factor(s) that regulate goat TAC3 transcription.

Transcription start site associated RNAs (TSSaRNAs) are ubiquitous in all domains of life.

PubMed

Zaramela, Livia S; Vêncio, Ricardo Z N; ten-Caten, Felipe; Baliga, Nitin S; Koide, Tie

2014-01-01

A plethora of non-coding RNAs has been discovered using high-resolution transcriptomics tools, indicating that transcriptional and post-transcriptional regulation is much more complex than previously appreciated. Small RNAs associated with transcription start sites of annotated coding regions (TSSaRNAs) are pervasive in both eukaryotes and bacteria. Here, we provide evidence for existence of TSSaRNAs in several archaeal transcriptomes including: Halobacterium salinarum, Pyrococcus furiosus, Methanococcus maripaludis, and Sulfolobus solfataricus. We validated TSSaRNAs from the model archaeon Halobacterium salinarum NRC-1 by deep sequencing two independent small-RNA enriched (RNA-seq) and a primary-transcript enriched (dRNA-seq) strand-specific libraries. We identified 652 transcripts, of which 179 were shown to be primary transcripts (∼7% of the annotated genome). Distinct growth-associated expression patterns between TSSaRNAs and their cognate genes were observed, indicating a possible role in environmental responses that may result from RNA polymerase with varying pausing rhythms. This work shows that TSSaRNAs are ubiquitous across all domains of life.

AphasiaBank: a resource for clinicians.

PubMed

Forbes, Margaret M; Fromm, Davida; Macwhinney, Brian

2012-08-01

AphasiaBank is a shared, multimedia database containing videos and transcriptions of ~180 aphasic individuals and 140 nonaphasic controls performing a uniform set of discourse tasks. The language in the videos is transcribed in Codes for the Human Analysis of Transcripts (CHAT) format and coded for analysis with Computerized Language ANalysis (CLAN) programs, which can perform a wide variety of language analyses. The database and the CLAN programs are freely available to aphasia researchers and clinicians for educational, clinical, and scholarly uses. This article describes the database, suggests some ways in which clinicians and clinician researchers might find these materials useful, and introduces a new language analysis program, EVAL, designed to streamline the transcription and coding processes, while still producing an extensive and useful language profile. Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.

Conserved expression of transposon-derived non-coding transcripts in primate stem cells.

PubMed

Ramsay, LeeAnn; Marchetto, Maria C; Caron, Maxime; Chen, Shu-Huang; Busche, Stephan; Kwan, Tony; Pastinen, Tomi; Gage, Fred H; Bourque, Guillaume

2017-02-28

A significant portion of expressed non-coding RNAs in human cells is derived from transposable elements (TEs). Moreover, it has been shown that various long non-coding RNAs (lncRNAs), which come from the human endogenous retrovirus subfamily H (HERVH), are not only expressed but required for pluripotency in human embryonic stem cells (hESCs). To identify additional TE-derived functional non-coding transcripts, we generated RNA-seq data from induced pluripotent stem cells (iPSCs) of four primate species (human, chimpanzee, gorilla, and rhesus) and searched for transcripts whose expression was conserved. We observed that about 30% of TE instances expressed in human iPSCs had orthologous TE instances that were also expressed in chimpanzee and gorilla. Notably, our analysis revealed a number of repeat families with highly conserved expression profiles including HERVH but also MER53, which is known to be the source of a placental-specific family of microRNAs (miRNAs). We also identified a number of repeat families from all classes of TEs, including MLT1-type and Tigger families, that contributed a significant amount of sequence to primate lncRNAs whose expression was conserved. Together, these results describe TE families and TE-derived lncRNAs whose conserved expression patterns can be used to identify what are likely functional TE-derived non-coding transcripts in primate iPSCs.

Long non-coding RNA produced by RNA polymerase V determines boundaries of heterochromatin

PubMed Central

Böhmdorfer, Gudrun; Sethuraman, Shriya; Rowley, M Jordan; Krzyszton, Michal; Rothi, M Hafiz; Bouzit, Lilia; Wierzbicki, Andrzej T

2016-01-01

RNA-mediated transcriptional gene silencing is a conserved process where small RNAs target transposons and other sequences for repression by establishing chromatin modifications. A central element of this process are long non-coding RNAs (lncRNA), which in Arabidopsis thaliana are produced by a specialized RNA polymerase known as Pol V. Here we show that non-coding transcription by Pol V is controlled by preexisting chromatin modifications located within the transcribed regions. Most Pol V transcripts are associated with AGO4 but are not sliced by AGO4. Pol V-dependent DNA methylation is established on both strands of DNA and is tightly restricted to Pol V-transcribed regions. This indicates that chromatin modifications are established in close proximity to Pol V. Finally, Pol V transcription is preferentially enriched on edges of silenced transposable elements, where Pol V transcribes into TEs. We propose that Pol V may play an important role in the determination of heterochromatin boundaries. DOI: http://dx.doi.org/10.7554/eLife.19092.001 PMID:27779094

Diversity of transcripts and transcript processing forms in plastids of the dinoflagellate alga Karenia mikimotoi.

PubMed

Dorrell, Richard G; Hinksman, George A; Howe, Christopher J

2016-02-01

Plastids produce a vast diversity of transcripts. These include mature transcripts containing coding sequences, and their processing precursors, as well as transcripts that lack direct coding functions, such as antisense transcripts. Although plastid transcriptomes have been characterised for many plant species, less is known about the transcripts produced in other plastid lineages. We characterised the transcripts produced in the fucoxanthin-containing plastids of the dinoflagellate alga Karenia mikimotoi. This plastid lineage, acquired through tertiary endosymbiosis, utilises transcript processing pathways that are very different from those found in plants and green algae, including 3' poly(U) tail addition, and extensive substitutional editing of transcript sequences. We have sequenced the plastid transcriptome of K. mikimotoi, and have detected evidence for divergent evolution of fucoxanthin plastid genomes. We have additionally characterised polycistronic and monocistronic transcripts from two plastid loci, psbD-tRNA (Met)-ycf4 and rpl36-rps13-rps11. We find evidence for a range of transcripts produced from each locus that differ in terms of editing state, 5' end cleavage position, and poly(U) tail addition. Finally, we identify antisense transcripts in K. mikimotoi, which appear to undergo different processing events from the corresponding sense transcripts. Overall, our study provides insights into the diversity of transcripts and processing intermediates found in plastid lineages across the eukaryotes.

groHMM: a computational tool for identifying unannotated and cell type-specific transcription units from global run-on sequencing data.

PubMed

Chae, Minho; Danko, Charles G; Kraus, W Lee

2015-07-16

Global run-on coupled with deep sequencing (GRO-seq) provides extensive information on the location and function of coding and non-coding transcripts, including primary microRNAs (miRNAs), long non-coding RNAs (lncRNAs), and enhancer RNAs (eRNAs), as well as yet undiscovered classes of transcripts. However, few computational tools tailored toward this new type of sequencing data are available, limiting the applicability of GRO-seq data for identifying novel transcription units. Here, we present groHMM, a computational tool in R, which defines the boundaries of transcription units de novo using a two state hidden-Markov model (HMM). A systematic comparison of the performance between groHMM and two existing peak-calling methods tuned to identify broad regions (SICER and HOMER) favorably supports our approach on existing GRO-seq data from MCF-7 breast cancer cells. To demonstrate the broader utility of our approach, we have used groHMM to annotate a diverse array of transcription units (i.e., primary transcripts) from four GRO-seq data sets derived from cells representing a variety of different human tissue types, including non-transformed cells (cardiomyocytes and lung fibroblasts) and transformed cells (LNCaP and MCF-7 cancer cells), as well as non-mammalian cells (from flies and worms). As an example of the utility of groHMM and its application to questions about the transcriptome, we show how groHMM can be used to analyze cell type-specific enhancers as defined by newly annotated enhancer transcripts. Our results show that groHMM can reveal new insights into cell type-specific transcription by identifying novel transcription units, and serve as a complete and useful tool for evaluating functional genomic elements in cells.

Complexity of CNC transcription factors as revealed by gene targeting of the Nrf3 locus.

PubMed

Derjuga, Anna; Gourley, Tania S; Holm, Teresa M; Heng, Henry H Q; Shivdasani, Ramesh A; Ahmed, Rafi; Andrews, Nancy C; Blank, Volker

2004-04-01

Cap'n'collar (CNC) family basic leucine zipper transcription factors play crucial roles in the regulation of mammalian gene expression and development. To determine the in vivo function of the CNC protein Nrf3 (NF-E2-related factor 3), we generated mice deficient in this transcription factor. We performed targeted disruption of two Nrf3 exons coding for CNC homology, basic DNA-binding, and leucine zipper dimerization domains. Nrf3 null mice developed normally and revealed no obvious phenotypic differences compared to wild-type animals. Nrf3(-/-) mice were fertile, and gross anatomy as well as behavior appeared normal. The mice showed normal age progression and did not show any apparent additional phenotype during their life span. We observed no differences in various blood parameters and chemistry values. We infected wild-type and Nrf3(-/-) mice with acute lymphocytic choriomeningitis virus and found no differences in these animals with respect to their number of virus-specific CD8 and CD4 T cells as well as their B-lymphocyte response. To determine whether the mild phenotype of Nrf3 null animals is due to functional redundancy, we generated mice deficient in multiple CNC factors. Contrary to our expectations, an absence of Nrf3 does not seem to cause additional lethality in compound Nrf3(-/-)/Nrf2(-/-) and Nrf3(-/-)/p45(-/-) mice. We hypothesize that the role of Nrf3 in vivo may become apparent only after appropriate challenge to the mice.

Decoding the ubiquitous role of microRNAs in neurogenesis.

PubMed

Nampoothiri, Sreekala S; Rajanikant, G K

2017-04-01

Neurogenesis generates fledgling neurons that mature to form an intricate neuronal circuitry. The delusion on adult neurogenesis was far resolved in the past decade and became one of the largely explored domains to identify multifaceted mechanisms bridging neurodevelopment and neuropathology. Neurogenesis encompasses multiple processes including neural stem cell proliferation, neuronal differentiation, and cell fate determination. Each neurogenic process is specifically governed by manifold signaling pathways, several growth factors, coding, and non-coding RNAs. A class of small non-coding RNAs, microRNAs (miRNAs), is ubiquitously expressed in the brain and has emerged to be potent regulators of neurogenesis. It functions by fine-tuning the expression of specific neurogenic gene targets at the post-transcriptional level and modulates the development of mature neurons from neural progenitor cells. Besides the commonly discussed intrinsic factors, the neuronal morphogenesis is also under the control of several extrinsic temporal cues, which in turn are regulated by miRNAs. This review enlightens on dicer controlled switch from neurogenesis to gliogenesis, miRNA regulation of neuronal maturation and the differential expression of miRNAs in response to various extrinsic cues affecting neurogenesis.

Over Expression of Long Non-Coding RNA PANDA Promotes Hepatocellular Carcinoma by Inhibiting Senescence Associated Inflammatory Factor IL8.

PubMed

Peng, Chuanhui; Hu, Wendi; Weng, Xiaoyu; Tong, Rongliang; Cheng, Shaobing; Ding, Chaofeng; Xiao, Heng; Lv, Zhen; Xie, Haiyang; Zhou, Lin; Wu, Jian; Zheng, Shusen

2017-06-23

It has been reported that long non-coding RNA PANDA was disregulated in varieties types of tumor, but its expression level and biological role in hepatocellular carcinoma (HCC) remains contradictory. We detected PANDA expression in two independent cohorts (48 HCC patients following liver transplantation and 84 HCC patients following liver resection), and found that PANDA was down-regulated in HCC. Thereafter we explored its function in cancer biology by inversing its low expression. Surprisingly, overexpression of PANDA promoted HCC proliferation and carcinogenesis in vitro and in vivo. Mechanistically, PANDA repressed transcriptional activity of senescence associated inflammatory factor IL8, which leaded to inhibition of cellular senescence. Therefore, our research help to better understand the complex role of PANDA in HCC, and suggest more thoughtful strategies should be applied before it can be treated as a potential therapeutic target.

The effects of potato virus Y-derived virus small interfering RNAs of three biologically distinct strains on potato (Solanum tuberosum) transcriptome.

PubMed

Moyo, Lindani; Ramesh, Shunmugiah V; Kappagantu, Madhu; Mitter, Neena; Sathuvalli, Vidyasagar; Pappu, Hanu R

2017-07-17

Potato virus Y (PVY) is one of the most economically important pathogen of potato that is present as biologically distinct strains. The virus-derived small interfering RNAs (vsiRNAs) from potato cv. Russet Burbank individually infected with PVY-N, PVY-NTN and PVY-O strains were recently characterized. Plant defense RNA-silencing mechanisms deployed against viruses produce vsiRNAs to degrade homologous viral transcripts. Based on sequence complementarity, the vsiRNAs can potentially degrade host RNA transcripts raising the prospect of vsiRNAs as pathogenicity determinants in virus-host interactions. This study investigated the global effects of PVY vsiRNAs on the host potato transcriptome. The strain-specific vsiRNAs of PVY, expressed in high copy number, were analyzed in silico for their proclivity to target potato coding and non-coding RNAs using psRobot and psRNATarget algorithms. Functional annotation of target coding transcripts was carried out to predict physiological effects of the vsiRNAs on the potato cv. Russet Burbank. The downregulation of selected target coding transcripts was further validated using qRT-PCR. The vsiRNAs derived from biologically distinct strains of PVY displayed diversity in terms of absolute number, copy number and hotspots for siRNAs on their respective genomes. The vsiRNAs populations were derived with a high frequency from 6 K1, P1 and Hc-Pro for PVY-N, P1, Hc-Pro and P3 for PVY-NTN, and P1, 3' UTR and NIa for PVY-O genomic regions. The number of vsiRNAs that displayed interaction with potato coding transcripts and number of putative coding target transcripts were comparable between PVY-N and PVY-O, and were relatively higher for PVY-NTN. The most abundant target non-coding RNA transcripts for the strain specific PVY-derived vsiRNAs were found to be MIR821, 28S rRNA,18S rRNA, snoR71, tRNA-Met and U5. Functional annotation and qRT-PCR validation suggested that the vsiRNAs target genes involved in plant hormone signaling, genetic information processing, plant-pathogen interactions, plant defense and stress response processes in potato. The findings suggested that the PVY-derived vsiRNAs could act as a pathogenicity determinant and as a counter-defense strategy to host RNA silencing in PVY-potato interactions. The broad range of host genes targeted by PVY vsiRNAs in infected potato suggests a diverse role for vsiRNAs that includes suppression of host stress responses and developmental processes. The interactome scenario is the first report on the interaction between one of the most important Potyvirus genome-derived siRNAs and the potato transcripts.

Human RNA polymerase II associated factor 1 complex promotes tumorigenesis by activating c-MYC transcription in non-small cell lung cancer

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhi, Xiuyi; Giroux-Leprieur, Etienne; Respiratory Diseases and Thoracic Oncology Department, Ambroise Pare Hospital – APHP, Versailles Saint Quentin en Yvelines University, 9 Avenue Charles de Gaulle, 92100, Boulogne-Billancourt

2015-10-02

Human RNA polymerase II (RNAPII)-associated factor 1 complex (hPAF1C) plays a crucial role in protein-coding gene transcription. Overexpression of hPAF1C has been implicated in the initiation and progression of various human cancers. However, the molecular pathways involved in tumorigenesis through hPAF1C remain to be elucidated. The current study suggested hPAF1C expression as a prognostic biomarker for early stage non-small cell lung cancer (NSCLC) and patients with low hPAF1C expression levels had significantly better overall survival. Furthermore, the expression of hPAF1C was found to be positively correlated with c-MYC expression in patient tumor samples and in cancer cell lines. Mechanistic studiesmore » indicated that hPAF1C could promote lung cancer cell proliferation through regulating c-MYC transcription. These results demonstrated the prognostic value of hPAF1C in early-stage NSCLC and the role of hPAF1C in the transcriptional regulation of c-MYC oncogene during NSCLC tumorigenesis. - Highlights: • hPAF1C expression is a prognostic biomarker for early stage non-small cell lung cancer. • The expression of hPAF1C was positively correlated with c-MYC in tumor samples of patients and in several NSCLC cell lines. • hPAF1C could promote lung cancer cell proliferation through regulating c-MYC transcription.« less

DNA methylation of the GC box in the promoter region mediates isolation rearing-induced suppression of srd5a1 transcription in the prefrontal cortex.

PubMed

Araki, Ryota; Nishida, Shoji; Hiraki, Yosuke; Matsumoto, Kinzo; Yabe, Takeshi

2015-10-08

The levels of allopregnanolone (ALLO), a neurosteroid, in brain and serum are related to severity of depression and anxiety. Steroid 5α-reductase type I is the rate-limiting enzyme in ALLO biosynthesis and plays an important role in control of the ALLO level in mammalian brain. In this study, we examined an epigenetic mechanism for transcriptional regulation of srd5a1, which codes for steroid 5α-reductase type I, using isolation-reared mice. The mRNA level of srd5a1 was decreased in the prefrontal cortex (PFC) in isolation-reared mice. Rearing in social isolation increased methylation of cytosines at -82 and -12 bp downstream of the transcription start site, which are located in a GC box element in the promoter region of srd5a1. Binding of Sp1, a ubiquitous transcription factor, to the GC box was decreased in the promoter region of srd5a1 in the PFC in isolation-reared mice. Site-specific methylation at cytosine -12 of a srd5a1 promoter-luciferase reporter construct, but not that of cytosine -82, downregulated the promoter activity of srd5a1. These findings suggest that transcription of srd5a1 in brain is regulated by environmental factor-induced cytosine methylation in the promoter region. This finding could contribute to development of antidepressant and anxiolytic agents. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Long non-coding RNA MALAT1 interacts with transcription factor Foxo1 to regulate SIRT1 transcription in high glucose-induced HK-2 cells injury.

PubMed

Zhou, Ling; Xu, De-Yu; Sha, Wen-Gang; Shen, Lei; Lu, Guo-Yuan

2018-06-18

Tubular injury is considered as a crucial pathological feature of diabetic nephropathy. LncRNA MALAT1 is involved in diabetic complications. Hence the role of MALAT1 in high glucose-induced renal tubular epithelial cells (HK-2) injury deserves investigation. The diabetic mice model was established with streptozotocin (STZ) injection. The expression of NEAT1, SIRT1, and Foxo1 mRNA and protein was determined with qRT-PCR and western blot, respectively. The serum creatinine and urinary albumin were examined by enzyme linked immunosorbent assay (ELISA). Interaction between MALAT1 and Foxo1 was detected with RIP and RNA pull-down assay, respectively. Dual luciferase reporter assay was used to evaluate the binding between Foxo1 and SIRT1. LncRNA MALAT1 was up-regulated in kidney tissues of diabetic mice and in HK-2 cells treated with high glucose, while the expression of SIRT1 was decreased. Interaction between MALAT1 and Foxo1 was observed in HK-2 cells and the interaction was promoted by high glucose treatment. Foxo1 activated SIRT1 transcription by binding to its promoter, and MALAT1 repressed SIRT1 expression through targeting Foxo1. LncRNA MALAT1 interacts with transcription factor Foxo1 to represses SIRT1 transcription in high glucose incubated HK-2 cells, which promotes high glucose-induced HK-2 cells injury. Copyright © 2018. Published by Elsevier Inc.

Candida albicans Swi/Snf and Mediator Complexes Differentially Regulate Mrr1-Induced MDR1 Expression and Fluconazole Resistance.

PubMed

Liu, Zhongle; Myers, Lawrence C

2017-11-01

Long-term azole treatment of patients with chronic Candida albicans infections can lead to drug resistance. Gain-of-function (GOF) mutations in the transcription factor Mrr1 and the consequent transcriptional activation of MDR1 , a drug efflux coding gene, is a common pathway by which this human fungal pathogen acquires fluconazole resistance. This work elucidates the previously unknown downstream transcription mechanisms utilized by hyperactive Mrr1. We identified the Swi/Snf chromatin remodeling complex as a key coactivator for Mrr1, which is required to maintain basal and induced open chromatin, and Mrr1 occupancy, at the MDR1 promoter. Deletion of snf2 , the catalytic subunit of Swi/Snf, largely abrogates the increases in MDR1 expression and fluconazole MIC observed in MRR1 GOF mutant strains. Mediator positively and negatively regulates key Mrr1 target promoters. Deletion of the Mediator tail module med3 subunit reduces, but does not eliminate, the increased MDR1 expression and fluconazole MIC conferred by MRR1 GOF mutations. Eliminating the kinase activity of the Mediator Ssn3 subunit suppresses the decreased MDR1 expression and fluconazole MIC of the snf2 null mutation in MRR1 GOF strains. Ssn3 deletion also suppresses MDR1 promoter histone displacement defects in snf2 null mutants. The combination of this work with studies on other hyperactive zinc cluster transcription factors that confer azole resistance in fungal pathogens reveals a complex picture where the induction of drug efflux pump expression requires the coordination of multiple coactivators. The observed variations in transcription factor and target promoter dependence of this process may make the search for azole sensitivity-restoring small molecules more complicated. Copyright © 2017 American Society for Microbiology.

Predicting Phonetic Transcription Agreement: Insights from Research in Infant Vocalizations

ERIC Educational Resources Information Center

Ramsdell, Heather L.; Oller, D. Kimbrough; Ethington, Corinna A.

2007-01-01

The purpose of this study is to provide new perspectives on correlates of phonetic transcription agreement. Our research focuses on phonetic transcription and coding of infant vocalizations. The findings are presumed to be broadly applicable to other difficult cases of transcription, such as found in severe disorders of speech, which similarly…

Quantitative Profiling of Peptides from RNAs classified as non-coding

PubMed Central

Prabakaran, Sudhakaran; Hemberg, Martin; Chauhan, Ruchi; Winter, Dominic; Tweedie-Cullen, Ry Y.; Dittrich, Christian; Hong, Elizabeth; Gunawardena, Jeremy; Steen, Hanno; Kreiman, Gabriel; Steen, Judith A.

2014-01-01

Only a small fraction of the mammalian genome codes for messenger RNAs destined to be translated into proteins, and it is generally assumed that a large portion of transcribed sequences - including introns and several classes of non-coding RNAs (ncRNAs) do not give rise to peptide products. A systematic examination of translation and physiological regulation of ncRNAs has not been conducted. Here, we use computational methods to identify the products of non-canonical translation in mouse neurons by analyzing unannotated transcripts in combination with proteomic data. This study supports the existence of non-canonical translation products from both intragenic and extragenic genomic regions, including peptides derived from anti-sense transcripts and introns. Moreover, the studied novel translation products exhibit temporal regulation similar to that of proteins known to be involved in neuronal activity processes. These observations highlight a potentially large and complex set of biologically regulated translational events from transcripts formerly thought to lack coding potential. PMID:25403355

Speech processing using conditional observable maximum likelihood continuity mapping

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hogden, John; Nix, David

A computer implemented method enables the recognition of speech and speech characteristics. Parameters are initialized of first probability density functions that map between the symbols in the vocabulary of one or more sequences of speech codes that represent speech sounds and a continuity map. Parameters are also initialized of second probability density functions that map between the elements in the vocabulary of one or more desired sequences of speech transcription symbols and the continuity map. The parameters of the probability density functions are then trained to maximize the probabilities of the desired sequences of speech-transcription symbols. A new sequence ofmore » speech codes is then input to the continuity map having the trained first and second probability function parameters. A smooth path is identified on the continuity map that has the maximum probability for the new sequence of speech codes. The probability of each speech transcription symbol for each input speech code can then be output.« less

Characterization of New Otic Enhancers of the Pou3f4 Gene Reveal Distinct Signaling Pathway Regulation and Spatio-Temporal Patterns

PubMed Central

Robert-Moreno, Àlex; Naranjo, Silvia; de la Calle-Mustienes, Elisa; Gómez-Skarmeta, José Luis; Alsina, Berta

2010-01-01

POU3F4 is a member of the POU-homedomain transcription factor family with a prominent role in inner ear development. Mutations in the human POU3F4 coding unit leads to X-linked deafness type 3 (DFN3), characterized by conductive hearing loss and progressive sensorineural deafness. Microdeletions found 1 Mb 5′ upstream of the coding region also displayed the same phenotype, suggesting that cis-regulatory elements might be present in that region. Indeed, we and others have recently identified several enhancers at the 1 Mb 5′ upstream interval of the pou3f4 locus. Here we characterize the spatio-temporal patterns of these regulatory elements in zebrafish transgenic lines. We show that the most distal enhancer (HCNR 81675) is activated earlier and drives GFP reporter expression initially to a broad ear domain to progressively restrict to the sensory patches. The proximal enhancer (HCNR 82478) is switched later during development and promotes expression, among in other tissues, in sensory patches from its onset. The third enhancer (HCNR 81728) is also active at later stages in the otic mesenchyme and in the otic epithelium. We also characterize the signaling pathways regulating these enhancers. While HCNR 81675 is regulated by very early signals of retinoic acid, HCNR 82478 is regulated by Fgf activity at a later stage and the HCNR 81728 enhancer is under the control of Hh signaling. Finally, we show that Sox2 and Pax2 transcription factors are bound to HCNR 81675 genomic region during otic development and specific mutations to these transcription factor binding sites abrogates HCNR 81675 enhancer activity. Altogether, our results suggest that pou3f4 expression in inner ear might be under the control of distinct regulatory elements that fine-tune the spatio-temporal activity of this gene and provides novel data on the signaling mechanisms controlling pou3f4 function. PMID:21209840

Conservation of σ28-Dependent Non-Coding RNA Paralogs and Predicted σ54-Dependent Targets in Thermophilic Campylobacter Species

PubMed Central

Le, My Thanh; van Veldhuizen, Mart; Porcelli, Ida; Bongaerts, Roy J.; Gaskin, Duncan J. H.; Pearson, Bruce M.; van Vliet, Arnoud H. M.

2015-01-01

Assembly of flagella requires strict hierarchical and temporal control via flagellar sigma and anti-sigma factors, regulatory proteins and the assembly complex itself, but to date non-coding RNAs (ncRNAs) have not been described to regulate genes directly involved in flagellar assembly. In this study we have investigated the possible role of two ncRNA paralogs (CjNC1, CjNC4) in flagellar assembly and gene regulation of the diarrhoeal pathogen Campylobacter jejuni. CjNC1 and CjNC4 are 37/44 nt identical and predicted to target the 5' untranslated region (5' UTR) of genes transcribed from the flagellar sigma factor σ54. Orthologs of the σ54-dependent 5' UTRs and ncRNAs are present in the genomes of other thermophilic Campylobacter species, and transcription of CjNC1 and CNC4 is dependent on the flagellar sigma factor σ28. Surprisingly, inactivation and overexpression of CjNC1 and CjNC4 did not affect growth, motility or flagella-associated phenotypes such as autoagglutination. However, CjNC1 and CjNC4 were able to mediate sequence-dependent, but Hfq-independent, partial repression of fluorescence of predicted target 5' UTRs in an Escherichia coli-based GFP reporter gene system. This hints towards a subtle role for the CjNC1 and CjNC4 ncRNAs in post-transcriptional gene regulation in thermophilic Campylobacter species, and suggests that the currently used phenotypic methodologies are insufficiently sensitive to detect such subtle phenotypes. The lack of a role of Hfq in the E. coli GFP-based system indicates that the CjNC1 and CjNC4 ncRNAs may mediate post-transcriptional gene regulation in ways that do not conform to the paradigms obtained from the Enterobacteriaceae. PMID:26512728

Network analysis for the visualization and analysis of qualitative data.

PubMed

Pokorny, Jennifer J; Norman, Alex; Zanesco, Anthony P; Bauer-Wu, Susan; Sahdra, Baljinder K; Saron, Clifford D

2018-03-01

We present a novel manner in which to visualize the coding of qualitative data that enables representation and analysis of connections between codes using graph theory and network analysis. Network graphs are created from codes applied to a transcript or audio file using the code names and their chronological location. The resulting network is a representation of the coding data that characterizes the interrelations of codes. This approach enables quantification of qualitative codes using network analysis and facilitates examination of associations of network indices with other quantitative variables using common statistical procedures. Here, as a proof of concept, we applied this method to a set of interview transcripts that had been coded in 2 different ways and the resultant network graphs were examined. The creation of network graphs allows researchers an opportunity to view and share their qualitative data in an innovative way that may provide new insights and enhance transparency of the analytical process by which they reach their conclusions. (PsycINFO Database Record (c) 2018 APA, all rights reserved).

Nonproteolytic Roles of 19S ATPases in Transcription of CIITApIV Genes

PubMed Central

Maganti, Nagini; Moody, Tomika D.; Truax, Agnieszka D.; Thakkar, Meghna; Spring, Alexander M.; Germann, Markus W.; Greer, Susanna F.

2014-01-01

Accumulating evidence shows the 26S proteasome is involved in the regulation of gene expression. We and others have demonstrated that proteasome components bind to sites of gene transcription, regulate covalent modifications to histones, and are involved in the assembly of activator complexes in mammalian cells. The mechanisms by which the proteasome influences transcription remain unclear, although prior observations suggest both proteolytic and non-proteolytic activities. Here, we define novel, non-proteolytic, roles for each of the three 19S heterodimers, represented by the 19S ATPases Sug1, S7, and S6a, in mammalian gene expression using the inflammatory gene CIITApIV. These 19S ATPases are recruited to induced CIITApIV promoters and also associate with CIITA coding regions. Additionally, these ATPases interact with elongation factor PTEFb complex members CDK9 and Hexim-1 and with Ser5 phosphorylated RNA Pol II. Both the generation of transcripts from CIITApIV and efficient recruitment of RNA Pol II to CIITApIV are negatively impacted by siRNA mediated knockdown of these 19S ATPases. Together, these results define novel roles for 19S ATPases in mammalian gene expression and indicate roles for these ATPases in promoting transcription processes. PMID:24625964

Constitutive Expression of a Transcription Termination Factor by a Repressed Prophage: Promoters for Transcribing the Phage HK022 nun Gene

PubMed Central

King, Rodney A.; Madsen, Peter L.; Weisberg, Robert A.

2000-01-01

Lysogens of phage HK022 are resistant to infection by phage λ. Lambda resistance is caused by the action of the HK022 Nun protein, which prematurely terminates early λ transcripts. We report here that transcription of the nun gene initiates at a constitutive prophage promoter, PNun, located just upstream of the protein coding sequence. The 5′ end of the transcript was determined by primer extension analysis of RNA isolated from HK022 lysogens or RNA made in vitro by transcribing a template containing the promoter with purified Escherichia coli RNA polymerase. Inactivation of PNun by mutation greatly reduced Nun activity and Nun antigen in an HK022 lysogen. However, a low level of residual activity was detected, suggesting that a secondary promoter also contributes to nun expression. We found one possible secondary promoter, PNun′, just upstream of PNun. Neither promoter is likely to increase the expression of other phage genes in a lysogen because their transcripts should be terminated downstream of nun. We estimate that HK022 lysogens in stationary phase contain several hundred molecules of Nun per cell and that cells in exponential phase probably contain fewer. PMID:10629193

Regulation of neural macroRNAs by the transcriptional repressor REST

PubMed Central

Johnson, Rory; Teh, Christina Hui-Leng; Jia, Hui; Vanisri, Ravi Raj; Pandey, Tridansh; Lu, Zhong-Hao; Buckley, Noel J.; Stanton, Lawrence W.; Lipovich, Leonard

2009-01-01

The essential transcriptional repressor REST (repressor element 1-silencing transcription factor) plays central roles in development and human disease by regulating a large cohort of neural genes. These have conventionally fallen into the class of known, protein-coding genes; recently, however, several noncoding microRNA genes were identified as REST targets. Given the widespread transcription of messenger RNA-like, noncoding RNAs (“macroRNAs”), some of which are functional and implicated in disease in mammalian genomes, we sought to determine whether this class of noncoding RNAs can also be regulated by REST. By applying a new, unbiased target gene annotation pipeline to computationally discovered REST binding sites, we find that 23% of mammalian REST genomic binding sites are within 10 kb of a macroRNA gene. These putative target genes were overlooked by previous studies. Focusing on a set of 18 candidate macroRNA targets from mouse, we experimentally demonstrate that two are regulated by REST in neural stem cells. Flanking protein-coding genes are, at most, weakly repressed, suggesting specific targeting of the macroRNAs by REST. Similar to the majority of known REST target genes, both of these macroRNAs are induced during nervous system development and have neurally restricted expression profiles in adult mouse. We observe a similar phenomenon in human: the DiGeorge syndrome-associated noncoding RNA, DGCR5, is repressed by REST through a proximal upstream binding site. Therefore neural macroRNAs represent an additional component of the REST regulatory network. These macroRNAs are new candidates for understanding the role of REST in neuronal development, neurodegeneration, and cancer. PMID:19050060

Regulation of neural macroRNAs by the transcriptional repressor REST.

PubMed

Johnson, Rory; Teh, Christina Hui-Leng; Jia, Hui; Vanisri, Ravi Raj; Pandey, Tridansh; Lu, Zhong-Hao; Buckley, Noel J; Stanton, Lawrence W; Lipovich, Leonard

2009-01-01

The essential transcriptional repressor REST (repressor element 1-silencing transcription factor) plays central roles in development and human disease by regulating a large cohort of neural genes. These have conventionally fallen into the class of known, protein-coding genes; recently, however, several noncoding microRNA genes were identified as REST targets. Given the widespread transcription of messenger RNA-like, noncoding RNAs ("macroRNAs"), some of which are functional and implicated in disease in mammalian genomes, we sought to determine whether this class of noncoding RNAs can also be regulated by REST. By applying a new, unbiased target gene annotation pipeline to computationally discovered REST binding sites, we find that 23% of mammalian REST genomic binding sites are within 10 kb of a macroRNA gene. These putative target genes were overlooked by previous studies. Focusing on a set of 18 candidate macroRNA targets from mouse, we experimentally demonstrate that two are regulated by REST in neural stem cells. Flanking protein-coding genes are, at most, weakly repressed, suggesting specific targeting of the macroRNAs by REST. Similar to the majority of known REST target genes, both of these macroRNAs are induced during nervous system development and have neurally restricted expression profiles in adult mouse. We observe a similar phenomenon in human: the DiGeorge syndrome-associated noncoding RNA, DGCR5, is repressed by REST through a proximal upstream binding site. Therefore neural macroRNAs represent an additional component of the REST regulatory network. These macroRNAs are new candidates for understanding the role of REST in neuronal development, neurodegeneration, and cancer.

Activating RNAs associate with Mediator to enhance chromatin architecture and transcription.

PubMed

Lai, Fan; Orom, Ulf A; Cesaroni, Matteo; Beringer, Malte; Taatjes, Dylan J; Blobel, Gerd A; Shiekhattar, Ramin

2013-02-28

Recent advances in genomic research have revealed the existence of a large number of transcripts devoid of protein-coding potential in multiple organisms. Although the functional role for long non-coding RNAs (lncRNAs) has been best defined in epigenetic phenomena such as X-chromosome inactivation and imprinting, different classes of lncRNAs may have varied biological functions. We and others have identified a class of lncRNAs, termed ncRNA-activating (ncRNA-a), that function to activate their neighbouring genes using a cis-mediated mechanism. To define the precise mode by which such enhancer-like RNAs function, we depleted factors with known roles in transcriptional activation and assessed their role in RNA-dependent activation. Here we report that depletion of the components of the co-activator complex, Mediator, specifically and potently diminished the ncRNA-induced activation of transcription in a heterologous reporter assay using human HEK293 cells. In vivo, Mediator is recruited to ncRNA-a target genes and regulates their expression. We show that ncRNA-a interact with Mediator to regulate its chromatin localization and kinase activity towards histone H3 serine 10. The Mediator complex harbouring disease- displays diminished ability to associate with activating ncRNAs. Chromosome conformation capture confirmed the presence of DNA looping between the ncRNA-a loci and its targets. Importantly, depletion of Mediator subunits or ncRNA-a reduced the chromatin looping between the two loci. Our results identify the human Mediator complex as the transducer of activating ncRNAs and highlight the importance of Mediator and activating ncRNA association in human disease.

Functional interrogation of non-coding DNA through CRISPR genome editing

PubMed Central

Canver, Matthew C.; Bauer, Daniel E.; Orkin, Stuart H.

2017-01-01

Methodologies to interrogate non-coding regions have lagged behind coding regions despite comprising the vast majority of the genome. However, the rapid evolution of clustered regularly interspaced short palindromic repeats (CRISPR)-based genome editing has provided a multitude of novel techniques for laboratory investigation including significant contributions to the toolbox for studying non-coding DNA. CRISPR-mediated loss-of-function strategies rely on direct disruption of the underlying sequence or repression of transcription without modifying the targeted DNA sequence. CRISPR-mediated gain-of-function approaches similarly benefit from methods to alter the targeted sequence through integration of customized sequence into the genome as well as methods to activate transcription. Here we review CRISPR-based loss- and gain-of-function techniques for the interrogation of non-coding DNA. PMID:28288828

A Multi-Label Classification Approach for Coding Cancer Information Service Chat Transcripts

PubMed Central

Rios, Anthony; Vanderpool, Robin; Shaw, Pam

2017-01-01

National Cancer Institute's (NCI) Cancer Information Service (CIS) offers online instant messaging based information service called LiveHelp to patients, family members, friends, and other cancer information consumers. A cancer information specialist (IS) ‘chats’ with a consumer and provides information on a variety of topics including clinical trials. After a LiveHelp chat session is finished, the IS codes about 20 different elements of metadata about the session in electronic contact record forms (ECRF), which are to be later used for quality control and reporting. Besides straightforward elements like age and gender, more specific elements to be coded include the purpose of contact, the subjects of interaction, and the different responses provided to the consumer, the latter two often taking on multiple values. As such, ECRF coding is a time consuming task and automating this process could help ISs to focus more on their primary goal of helping consumers with valuable cancer related information. As a first attempt in this task, we explored multi-label and multi-class text classification approaches to code the purpose, subjects of interaction, and the responses provided based on the chat transcripts. With a sample dataset of about 673 transcripts, we achieved example-based F-scores of 0.67 (for subjects) and 0.58 (responses). We also achieved label-based micro F-scores of 0.65 (for subjects), 0.62 (for responses), and 0.61 (for purpose). To our knowledge this is the first attempt in automatic coding of Live-Help transcripts and our initial results on the smaller corpus indicate promising future directions in this task. PMID:28736775

Long Non-coding RNAs in the X-inactivation Center

PubMed Central

Kalantry, Sundeep

2014-01-01

The X-inactivation center is a hotbed of functional long non-coding RNAs in eutherian mammals. These RNAs are thought to help orchestrate the epigenetic transcriptional states of the two X-chromosomes in females as well as of the single X-chromosome in males. To balance X-linked gene expression between the sexes, females undergo transcriptional silencing of most genes on one of the two X-chromosomes in a process termed X-chromosome inactivation. While one X-chromosome is inactivated, the other X-chromosome remains active. Moreover, with a few notable exceptions, the originally established epigenetic transcriptional profiles of the two is maintained as such through many rounds of cell division, essentially for the life of the organism. The stable divergent transcriptional fates of the two X-chromosomes, despite residing in a shared nucleoplasm, make X-inactivation a paradigm of epigenetic transcriptional regulation. Originally proposed in 1961 by Mary Lyon, the X-inactivation hypothesis has been validated through much experimentation over the last fifty years. In the last 25 years, the discovery and functional characterization has firmly established X-linked long non-coding RNAs as key players in choreographing X-chromosome inactivation. PMID:24297756

Evolution at protein ends: major contribution of alternative transcription initiation and termination to the transcriptome and proteome diversity in mammals

PubMed Central

Shabalina, Svetlana A.; Ogurtsov, Aleksey Y.; Spiridonov, Nikolay A.; Koonin, Eugene V.

2014-01-01

Alternative splicing (AS), alternative transcription initiation (ATI) and alternative transcription termination (ATT) create the extraordinary complexity of transcriptomes and make key contributions to the structural and functional diversity of mammalian proteomes. Analysis of mammalian genomic and transcriptomic data shows that contrary to the traditional view, the joint contribution of ATI and ATT to the transcriptome and proteome diversity is quantitatively greater than the contribution of AS. Although the mean numbers of protein-coding constitutive and alternative nucleotides in gene loci are nearly identical, their distribution along the transcripts is highly non-uniform. On average, coding exons in the variable 5′ and 3′ transcript ends that are created by ATI and ATT contain approximately four times more alternative nucleotides than core protein-coding regions that diversify exclusively via AS. Short upstream exons that encompass alternative 5′-untranslated regions and N-termini of proteins evolve under strong nucleotide-level selection whereas in 3′-terminal exons that encode protein C-termini, protein-level selection is significantly stronger. The groups of genes that are subject to ATI and ATT show major differences in biological roles, expression and selection patterns. PMID:24792168

Long Non-Coding RNAs Responsive to Salt and Boron Stress in the Hyper-Arid Lluteño Maize from Atacama Desert.

PubMed

Huanca-Mamani, Wilson; Arias-Carrasco, Raúl; Cárdenas-Ninasivincha, Steffany; Rojas-Herrera, Marcelo; Sepúlveda-Hermosilla, Gonzalo; Caris-Maldonado, José Carlos; Bastías, Elizabeth; Maracaja-Coutinho, Vinicius

2018-03-20

Long non-coding RNAs (lncRNAs) have been defined as transcripts longer than 200 nucleotides, which lack significant protein coding potential and possess critical roles in diverse cellular processes. Long non-coding RNAs have recently been functionally characterized in plant stress-response mechanisms. In the present study, we perform a comprehensive identification of lncRNAs in response to combined stress induced by salinity and excess of boron in the Lluteño maize, a tolerant maize landrace from Atacama Desert, Chile. We use deep RNA sequencing to identify a set of 48,345 different lncRNAs, of which 28,012 (58.1%) are conserved with other maize (B73, Mo17 or Palomero), with the remaining 41.9% belonging to potentially Lluteño exclusive lncRNA transcripts. According to B73 maize reference genome sequence, most Lluteño lncRNAs correspond to intergenic transcripts. Interestingly, Lluteño lncRNAs presents an unusual overall higher expression compared to protein coding genes under exposure to stressed conditions. In total, we identified 1710 putatively responsive to the combined stressed conditions of salt and boron exposure. We also identified a set of 848 stress responsive potential trans natural antisense transcripts ( trans -NAT) lncRNAs, which seems to be regulating genes associated with regulation of transcription, response to stress, response to abiotic stimulus and participating of the nicotianamine metabolic process. Reverse transcription-quantitative PCR (RT-qPCR) experiments were performed in a subset of lncRNAs, validating their existence and expression patterns. Our results suggest that a diverse set of maize lncRNAs from leaves and roots is responsive to combined salt and boron stress, being the first effort to identify lncRNAs from a maize landrace adapted to extreme conditions such as the Atacama Desert. The information generated is a starting point to understand the genomic adaptabilities suffered by this maize to surpass this extremely stressed environment.

Long Non-Coding RNAs Responsive to Salt and Boron Stress in the Hyper-Arid Lluteño Maize from Atacama Desert

PubMed Central

Huanca-Mamani, Wilson; Arias-Carrasco, Raúl; Cárdenas-Ninasivincha, Steffany; Rojas-Herrera, Marcelo; Sepúlveda-Hermosilla, Gonzalo; Caris-Maldonado, José Carlos; Bastías, Elizabeth; Maracaja-Coutinho, Vinicius

2018-01-01

Long non-coding RNAs (lncRNAs) have been defined as transcripts longer than 200 nucleotides, which lack significant protein coding potential and possess critical roles in diverse cellular processes. Long non-coding RNAs have recently been functionally characterized in plant stress–response mechanisms. In the present study, we perform a comprehensive identification of lncRNAs in response to combined stress induced by salinity and excess of boron in the Lluteño maize, a tolerant maize landrace from Atacama Desert, Chile. We use deep RNA sequencing to identify a set of 48,345 different lncRNAs, of which 28,012 (58.1%) are conserved with other maize (B73, Mo17 or Palomero), with the remaining 41.9% belonging to potentially Lluteño exclusive lncRNA transcripts. According to B73 maize reference genome sequence, most Lluteño lncRNAs correspond to intergenic transcripts. Interestingly, Lluteño lncRNAs presents an unusual overall higher expression compared to protein coding genes under exposure to stressed conditions. In total, we identified 1710 putatively responsive to the combined stressed conditions of salt and boron exposure. We also identified a set of 848 stress responsive potential trans natural antisense transcripts (trans-NAT) lncRNAs, which seems to be regulating genes associated with regulation of transcription, response to stress, response to abiotic stimulus and participating of the nicotianamine metabolic process. Reverse transcription-quantitative PCR (RT-qPCR) experiments were performed in a subset of lncRNAs, validating their existence and expression patterns. Our results suggest that a diverse set of maize lncRNAs from leaves and roots is responsive to combined salt and boron stress, being the first effort to identify lncRNAs from a maize landrace adapted to extreme conditions such as the Atacama Desert. The information generated is a starting point to understand the genomic adaptabilities suffered by this maize to surpass this extremely stressed environment. PMID:29558449

Quality control of mRNP biogenesis: networking at the transcription site.

PubMed

Eberle, Andrea B; Visa, Neus

2014-08-01

Eukaryotic cells carry out quality control (QC) over the processes of RNA biogenesis to inactivate or eliminate defective transcripts, and to avoid their production. In the case of protein-coding transcripts, the quality controls can sense defects in the assembly of mRNA-protein complexes, in the processing of the precursor mRNAs, and in the sequence of open reading frames. Different types of defect are monitored by different specialized mechanisms. Some of them involve dedicated factors whose function is to identify faulty molecules and target them for degradation. Others are the result of a more subtle balance in the kinetics of opposing activities in the mRNA biogenesis pathway. One way or another, all such mechanisms hinder the expression of the defective mRNAs through processes as diverse as rapid degradation, nuclear retention and transcriptional silencing. Three major degradation systems are responsible for the destruction of the defective transcripts: the exosome, the 5'-3' exoribonucleases, and the nonsense-mediated mRNA decay (NMD) machinery. This review summarizes recent findings on the cotranscriptional quality control of mRNA biogenesis, and speculates that a protein-protein interaction network integrates multiple mRNA degradation systems with the transcription machinery. Copyright © 2014 Elsevier Ltd. All rights reserved.

Surfactant Protein-C Promoter Variants Associated with Neonatal Respiratory Distress Syndrome Reduce Transcription

PubMed Central

Wambach, Jennifer A.; Yang, Ping; Wegner, Daniel J.; An, Ping; Hackett, Brian P.; Cole, F. S.; Hamvas, Aaron

2010-01-01

Dominant mutations in coding regions of the surfactant protein-C gene (SFTPC) cause respiratory distress syndrome (RDS) in infants. However, the contribution of variants in noncoding regions of SFTPC to pulmonary phenotypes is unknown. Using a case-control group of infants ≥34 weeks gestation (n=538), we used complete resequencing of SFTPC and its promoter, genotyping, and logistic regression to identify 80 single nucleotide polymorphisms (SNPs). Three promoter SNPs were statistically associated with neonatal RDS among European descent infants. To assess the transcriptional effects of these three promoter SNPs, we selectively mutated the SFTPC promoter and performed transient transfection using MLE-15 cells and a firefly luciferase reporter vector. Each promoter SNP decreased SFTPC transcription. The combination of two variants in high linkage dysequilibrium also decreased SFTPC transcription. In silico evaluation of transcription factor binding demonstrated that the rare allele at g.-1167 disrupts a SOX (SRY-related high mobility group box) consensus motif and introduces a GATA-1 site, at g.-2385 removes a MZF-1 (myeloid zinc finger) binding site, and at g.-1647 removes a potential methylation site. This combined statistical, in vitro, and in silico approach suggests that reduced SFTPC transcription contributes to the genetic risk for neonatal RDS in developmentally susceptible infants. PMID:20539253

Long Non-Coding RNAs: A Novel Paradigm for Toxicology.

PubMed

Dempsey, Joseph L; Cui, Julia Yue

2017-01-01

Long non-coding RNAs (lncRNAs) are over 200 nucleotides in length and are transcribed from the mammalian genome in a tissue-specific and developmentally regulated pattern. There is growing recognition that lncRNAs are novel biomarkers and/or key regulators of toxicological responses in humans and animal models. Lacking protein-coding capacity, the numerous types of lncRNAs possess a myriad of transcriptional regulatory functions that include cis and trans gene expression, transcription factor activity, chromatin remodeling, imprinting, and enhancer up-regulation. LncRNAs also influence mRNA processing, post-transcriptional regulation, and protein trafficking. Dysregulation of lncRNAs has been implicated in various human health outcomes such as various cancers, Alzheimer's disease, cardiovascular disease, autoimmune diseases, as well as intermediary metabolism such as glucose, lipid, and bile acid homeostasis. Interestingly, emerging evidence in the literature over the past five years has shown that lncRNA regulation is impacted by exposures to various chemicals such as polycyclic aromatic hydrocarbons, benzene, cadmium, chlorpyrifos-methyl, bisphenol A, phthalates, phenols, and bile acids. Recent technological advancements, including next-generation sequencing technologies and novel computational algorithms, have enabled the profiling and functional characterizations of lncRNAs on a genomic scale. In this review, we summarize the biogenesis and general biological functions of lncRNAs, highlight the important roles of lncRNAs in human diseases and especially during the toxicological responses to various xenobiotics, evaluate current methods for identifying aberrant lncRNA expression and molecular target interactions, and discuss the potential to implement these tools to address fundamental questions in toxicology. © The Author 2016. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Changes in the Coding and Non-coding Transcriptome and DNA Methylome that Define the Schwann Cell Repair Phenotype after Nerve Injury.

PubMed

Arthur-Farraj, Peter J; Morgan, Claire C; Adamowicz, Martyna; Gomez-Sanchez, Jose A; Fazal, Shaline V; Beucher, Anthony; Razzaghi, Bonnie; Mirsky, Rhona; Jessen, Kristjan R; Aitman, Timothy J

2017-09-12

Repair Schwann cells play a critical role in orchestrating nerve repair after injury, but the cellular and molecular processes that generate them are poorly understood. Here, we perform a combined whole-genome, coding and non-coding RNA and CpG methylation study following nerve injury. We show that genes involved in the epithelial-mesenchymal transition are enriched in repair cells, and we identify several long non-coding RNAs in Schwann cells. We demonstrate that the AP-1 transcription factor C-JUN regulates the expression of certain micro RNAs in repair Schwann cells, in particular miR-21 and miR-34. Surprisingly, unlike during development, changes in CpG methylation are limited in injury, restricted to specific locations, such as enhancer regions of Schwann cell-specific genes (e.g., Nedd4l), and close to local enrichment of AP-1 motifs. These genetic and epigenomic changes broaden our mechanistic understanding of the formation of repair Schwann cell during peripheral nervous system tissue repair. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

The gene and the genon concept: a functional and information-theoretic analysis

PubMed Central

Scherrer, Klaus; Jost, Jürgen

2007-01-01

‘Gene' has become a vague and ill-defined concept. To set the stage for mathematical analysis of gene storage and expression, we return to the original concept of the gene as a function encoded in the genome, basis of genetic analysis, that is a polypeptide or other functional product. The additional information needed to express a gene is contained within each mRNA as an ensemble of signals, added to or superimposed onto the coding sequence. To designate this programme, we introduce the term ‘genon'. Individual genons are contained in the pre-mRNA forming a pre-genon. A genomic domain contains a proto-genon, with the signals of transcription activation in addition to the pre-genon in the transcripts. Some contain several mRNAs and hence genons, to be singled out by RNA processing and differential splicing. The programme in the genon in cis is implemented by corresponding factors of protein or RNA nature contained in the transgenon of the cell or organism. The gene, the cis programme contained in the individual domain and transcript, and the trans programme of factors, can be analysed by information theory. PMID:17353929

The minisatellite of the GPI/AMF/NLK/MF gene: interspecies conservation and transcriptional activity.

PubMed

Williams, R R; Hassan-Walker, A F; Lavender, F L; Morgan, M; Faik, P; Ragoussis, J

2001-05-16

Minisatellites are tandemly repeated DNA sequences found throughout the genomes of all eukaryotes. They are regions often prone to instability and hence hypervariability; thus repeat unit sequence is generally not conserved beyond closely related species. We have studied the minisatellite located in intron 9 of the human glucose phosphate isomerase (GPI) gene (also known as neuroleukin, autocrine motility factor, maturation and differentiation factor) and have found, by Zoo blotting coupled with PCR amplification and DNA sequencing, that similar repeat units are present in seven other species of mammal. There is also evidence for the presence of the minisatellite in chicken. The repeat unit does not appear to be present at any other locus in these genomes. Minisatellite DNA has been reported to be involved in recombination activity, control of gene expression of nearby gene(s) (both transcriptional and translational), whilst others form protein coding regions. The high level of conservation exhibited by the GPI minisatellite, coupled with the unique location, strongly suggests a functional role. Our results from transient and stable transfections using luciferase reporter constructs have shown that the GPI minisatellite region can act to increase transcription from the SV40 promoter, CMV promoter and the human GPI promoter.

Profiling and bioinformatic analysis of circular RNA expression regulated by c-Myc.

PubMed

Gou, Qiheng; Wu, Ke; Zhou, Jian-Kang; Xie, Yuxin; Liu, Lunxu; Peng, Yong

2017-09-22

The c-Myc transcription factor is involved in cell proliferation, cell cycle and apoptosis by activating or repressing transcription of multiple genes. Circular RNAs (circRNAs) are widely expressed non-coding RNAs participating in the regulation of gene expression. Using a high-throughput microarray assay, we showed that Myc regulates the expression of certain circRNAs. A total of 309 up- and 252 down-regulated circRNAs were identified. Among them, randomly selected 8 circRNAs were confirmed by real-time PCR. Subsequently, Myc-binding sites were found to generally exist in the promoter regions of differentially expressed circRNAs. Based on miRNA sponge mechanism, we constructed circRNAs/miRNAs network regulated by Myc, suggesting that circRNAs may widely regulate protein expression through miRNA sponge mechanism. Lastly, we took advantage of Gene Ontology and KEGG analyses to point out that Myc-regulated circRNAs could impact cell proliferation through affecting Ras signaling pathway and pathways in cancer. Our study for the first time demonstrated that Myc transcription factor regulates the expression of circRNAs, adding a novel component of the Myc tumorigenic program and opening a window to investigate the function of certain circRNAs in tumorigenesis.

Open Source Subtitle Editor Software Study for Section 508 Close Caption Applications

NASA Technical Reports Server (NTRS)

Murphy, F. Brandon

2013-01-01

This paper will focus on a specific item within the NASA Electronic Information Accessibility Policy - Multimedia Presentation shall have synchronized caption; thus making information accessible to a person with hearing impairment. This synchronized caption will assist a person with hearing or cognitive disability to access the same information as everyone else. This paper focuses on the research and implementation for CC (subtitle option) support to video multimedia. The goal of this research is identify the best available open-source (free) software to achieve synchronized captions requirement and achieve savings, while meeting the security requirement for Government information integrity and assurance. CC and subtitling are processes that display text within a video to provide additional or interpretive information for those whom may need it or those whom chose it. Closed captions typically show the transcription of the audio portion of a program (video) as it occurs (either verbatim or in its edited form), sometimes including non-speech elements (such as sound effects). The transcript can be provided by a third party source or can be extracted word for word from the video. This feature can be made available for videos in two forms: either Soft-Coded or Hard-Coded. Soft-Coded is the more optional version of CC, where you can chose to turn them on if you want, or you can turn them off. Most of the time, when using the Soft-Coded option, the transcript is also provided to the view along-side the video. This option is subject to compromise, whereas the transcript is merely a text file that can be changed by anyone who has access to it. With this option the integrity of the CC is at the mercy of the user. Hard-Coded CC is a more permanent form of CC. A Hard-Coded CC transcript is embedded within a video, without the option of removal.

A chromatin activity based chemoproteomic approach reveals a transcriptional repressome for gene-specific silencing

PubMed Central

Liu, Cui; Yu, Yanbao; Liu, Feng; Wei, Xin; Wrobel, John A.; Gunawardena, Harsha P.; Zhou, Li; Jin, Jian; Chen, Xian

2015-01-01

Immune cells develop endotoxin tolerance (ET) after prolonged stimulation. ET increases the level of a repression mark H3K9me2 in the transcriptional-silent chromatin specifically associated with pro-inflammatory genes. However, it is not clear what proteins are functionally involved in this process. Here we show that a novel chromatin activity based chemoproteomic (ChaC) approach can dissect the functional chromatin protein complexes that regulate ET-associated inflammation. Using UNC0638 that binds the enzymatically active H3K9-specific methyltransferase G9a/GLP, ChaC reveals that G9a is constitutively active at a G9a-dependent mega-dalton repressome in primary endotoxin-tolerant macrophages. G9a/GLP broadly impacts the ET-specific reprogramming of the histone code landscape, chromatin remodeling, and the activities of select transcription factors. We discover that the G9a-dependent epigenetic environment promotes the transcriptional repression activity of c-Myc for gene-specific co-regulation of chronic inflammation. ChaC may be also applicable to dissect other functional protein complexes in the context of phenotypic chromatin architectures. PMID:25502336

Differentially-Expressed Pseudogenes in HIV-1 Infection

PubMed Central

Gupta, Aditi; Brown, C. Titus; Zheng, Yong-Hui; Adami, Christoph

2015-01-01

Not all pseudogenes are transcriptionally silent as previously thought. Pseudogene transcripts, although not translated, contribute to the non-coding RNA pool of the cell that regulates the expression of other genes. Pseudogene transcripts can also directly compete with the parent gene transcripts for mRNA stability and other cell factors, modulating their expression levels. Tissue-specific and cancer-specific differential expression of these “functional” pseudogenes has been reported. To ascertain potential pseudogene:gene interactions in HIV-1 infection, we analyzed transcriptomes from infected and uninfected T-cells and found that 21 pseudogenes are differentially expressed in HIV-1 infection. This is interesting because parent genes of one-third of these differentially-expressed pseudogenes are implicated in HIV-1 life cycle, and parent genes of half of these pseudogenes are involved in different viral infections. Our bioinformatics analysis identifies candidate pseudogene:gene interactions that may be of significance in HIV-1 infection. Experimental validation of these interactions would establish that retroviruses exploit this newly-discovered layer of host gene expression regulation for their own benefit. PMID:26426037

Long Non-coding RNAs and Their Biological Roles in Plants

PubMed Central

Liu, Xue; Hao, Lili; Li, Dayong; Zhu, Lihuang; Hu, Songnian

2015-01-01

With the development of genomics and bioinformatics, especially the extensive applications of high-throughput sequencing technology, more transcriptional units with little or no protein-coding potential have been discovered. Such RNA molecules are called non-protein-coding RNAs (npcRNAs or ncRNAs). Among them, long npcRNAs or ncRNAs (lnpcRNAs or lncRNAs) represent diverse classes of transcripts longer than 200 nucleotides. In recent years, the lncRNAs have been considered as important regulators in many essential biological processes. In plants, although a large number of lncRNA transcripts have been predicted and identified in few species, our current knowledge of their biological functions is still limited. Here, we have summarized recent studies on their identification, characteristics, classification, bioinformatics, resources, and current exploration of their biological functions in plants. PMID:25936895

Differential regulation of transcription through distinct Suppressor of Hairless DNA binding site architectures during Notch signaling in proneural clusters.

PubMed

Cave, John W; Xia, Li; Caudy, Michael

2011-01-01

In Drosophila melanogaster, achaete (ac) and m8 are model basic helix-loop-helix activator (bHLH A) and repressor genes, respectively, that have the opposite cell expression pattern in proneural clusters during Notch signaling. Previous studies have shown that activation of m8 transcription in specific cells within proneural clusters by Notch signaling is programmed by a "combinatorial" and "architectural" DNA transcription code containing binding sites for the Su(H) and proneural bHLH A proteins. Here we show the novel result that the ac promoter contains a similar combinatorial code of Su(H) and bHLH A binding sites but contains a different Su(H) site architectural code that does not mediate activation during Notch signaling, thus programming a cell expression pattern opposite that of m8 in proneural clusters.

The long non-coding RNA ROCR contributes to SOX9 expression and chondrogenic differentiation of human mesenchymal stem cells

PubMed Central

Hyatt, Sam; Cheung, Kat; Skelton, Andrew J.; Xu, Yaobo; Clark, Ian M.

2017-01-01

Long non-coding RNAs (lncRNAs) are expressed in a highly tissue-specific manner and function in various aspects of cell biology, often as key regulators of gene expression. In this study, we established a role for lncRNAs in chondrocyte differentiation. Using RNA sequencing we identified a human articular chondrocyte repertoire of lncRNAs from normal hip cartilage donated by neck of femur fracture patients. Of particular interest are lncRNAs upstream of the master chondrocyte transcription factor SOX9 locus. SOX9 is an HMG-box transcription factor that plays an essential role in chondrocyte development by directing the expression of chondrocyte-specific genes. Two of these lncRNAs are upregulated during chondrogenic differentiation of mesenchymal stem cells (MSCs). Depletion of one of these lncRNAs, LOC102723505, which we termed ROCR (regulator of chondrogenesis RNA), by RNA interference disrupted MSC chondrogenesis, concomitant with reduced cartilage-specific gene expression and incomplete matrix component production, indicating an important role in chondrocyte biology. Specifically, SOX9 induction was significantly ablated in the absence of ROCR, and overexpression of SOX9 rescued the differentiation of MSCs into chondrocytes. Our work sheds further light on chondrocyte-specific SOX9 expression and highlights a novel method of chondrocyte gene regulation involving a lncRNA. PMID:29084806

Genome-wide survey by ChIP-seq reveals YY1 regulation of lincRNAs in skeletal myogenesis

PubMed Central

Lu, Leina; Sun, Kun; Chen, Xiaona; Zhao, Yu; Wang, Lijun; Zhou, Liang; Sun, Hao; Wang, Huating

2013-01-01

Skeletal muscle differentiation is orchestrated by a network of transcription factors, epigenetic regulators, and non-coding RNAs. The transcription factor Yin Yang 1 (YY1) silences multiple target genes in myoblasts (MBs) by recruiting Ezh2 (Enhancer of Zeste Homologue2). To elucidate genome-wide YY1 binding in MBs, we performed chromatin immunoprecipitation (ChIP)-seq and found 1820 specific binding sites in MBs with a large portion residing in intergenic regions. Detailed analysis demonstrated that YY1 acts as an activator for many loci in addition to its known repressor function. No significant co-occupancy was found between YY1 and Ezh2, suggesting an additional Ezh2-independent function for YY1 in MBs. Further analysis of intergenic binding sites showed that YY1 potentially regulates dozens of large intergenic non-coding RNAs (lincRNAs), whose function in myogenesis is underexplored. We characterized a novel muscle-associated lincRNA (Yam-1) that is positively regulated by YY1. Yam-1 is downregulated upon differentiation and acts as an inhibitor of myogenesis. We demonstrated that Yam-1 functions through in cis regulation of miR-715, which in turn targets Wnt7b. Our findings not only provide the first genome-wide picture of YY1 association in muscle cells, but also uncover the functional role of lincRNA Yam-1. PMID:23942234

Computational systems biology analysis of biomarkers in lung cancer; unravelling genomic regions which frequently encode biomarkers, enriched pathways, and new candidates.

PubMed

Alanazi, Ibrahim O; AlYahya, Sami A; Ebrahimie, Esmaeil; Mohammadi-Dehcheshmeh, Manijeh

2018-06-15

Exponentially growing scientific knowledge in scientific publications has resulted in the emergence of a new interdisciplinary science of literature mining. In text mining, the machine reads the published literature and transfers the discovered knowledge to mathematical-like formulas. In an integrative approach in this study, we used text mining in combination with network discovery, pathway analysis, and enrichment analysis of genomic regions for better understanding of biomarkers in lung cancer. Particular attention was paid to non-coding biomarkers. In total, 60 MicroRNA biomarkers were reported for lung cancer, including some prognostic biomarkers. MIR21, MIR155, MALAT1, and MIR31 were the top non-coding RNA biomarkers of lung cancer. Text mining identified 447 proteins which have been studied as biomarkers in lung cancer. EGFR (receptor), TP53 (transcription factor), KRAS, CDKN2A, ENO2, KRT19, RASSF1, GRP (ligand), SHOX2 (transcription factor), and ERBB2 (receptor) were the most studied proteins. Within small molecules, thymosin-a1, oestrogen, and 8-OHdG have received more attention. We found some chromosomal bands, such as 7q32.2, 18q12.1, 6p12, 11p15.5, and 3p21.3 that are highly involved in deriving lung cancer biomarkers. Copyright © 2018 Elsevier B.V. All rights reserved.

GATA: A graphic alignment tool for comparative sequenceanalysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nix, David A.; Eisen, Michael B.

2005-01-01

Several problems exist with current methods used to align DNA sequences for comparative sequence analysis. Most dynamic programming algorithms assume that conserved sequence elements are collinear. This assumption appears valid when comparing orthologous protein coding sequences. Functional constraints on proteins provide strong selective pressure against sequence inversions, and minimize sequence duplications and feature shuffling. For non-coding sequences this collinearity assumption is often invalid. For example, enhancers contain clusters of transcription factor binding sites that change in number, orientation, and spacing during evolution yet the enhancer retains its activity. Dotplot analysis is often used to estimate non-coding sequence relatedness. Yet dotmore » plots do not actually align sequences and thus cannot account well for base insertions or deletions. Moreover, they lack an adequate statistical framework for comparing sequence relatedness and are limited to pairwise comparisons. Lastly, dot plots and dynamic programming text outputs fail to provide an intuitive means for visualizing DNA alignments.« less

[Neuromuscular system and aging: involutions and implications].

PubMed

Paillard, Thierry

2013-12-01

In aged human, the number of muscle fibers and motor units decreases. The remaining motor units lose their functionality (decrease of the discharge frequency, greater fluctuation of the discharge) particularly those which contain type II fibers. The renewal of intracellular proteins declines which creates a negative balance between the daily protein losses and the capacities to renew them. The activity of the protein kinase (Akt) that stimulates the synthesis of regulation proteins (mTOR, p70S6, IGFBP-5) declines whereas the factors of degradation of proteins (NF-kappa B) are activated. Besides, the process of activation and proliferation of satellite cells is affected and the production of anabolic hormones and local factors is decreased. After a strength training program, muscle hypertrophy is linked to the protein synthesis at the level of myosin heavy chain (MHC) isoforms in older subjects. However, the transcription of the genes that code the MHC-I (slow form) increases and the transcription of the genes that code the MHC-II (fast form) decreases. Thus, the transition of the phenotype towards a slower form cannot be inverted by strength training during the advanced in age. Moreover, strength training enables to decrease the proportion of fibers containing MHC of hybrid form in the process of evolution. Hence, strength training can engender a stabilization of the muscular phenotype i.e. different isoforms of MHC. In addition, strength training counteracts the noxious effects mentioned above by generating muscular hypertrophy thanks to a reactive increase in the production of anabolic hormones. A program of aerobic training can induce an increase in the synthesis of ARN messengers coding isoforms related to the oxidative metabolism (MHC-I and to a lesser extent MHC-IIa) while the transcribed for the type MHC-IIx decrease.

CHIR99021 promotes self-renewal of mouse embryonic stem cells by modulation of protein-encoding gene and long intergenic non-coding RNA expression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wu, Yongyan; Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A and F University, Yangling 712100, Shaanxi; Ai, Zhiying

2013-10-15

Embryonic stem cells (ESCs) can proliferate indefinitely in vitro and differentiate into cells of all three germ layers. These unique properties make them exceptionally valuable for drug discovery and regenerative medicine. However, the practical application of ESCs is limited because it is difficult to derive and culture ESCs. It has been demonstrated that CHIR99021 (CHIR) promotes self-renewal and enhances the derivation efficiency of mouse (m)ESCs. However, the downstream targets of CHIR are not fully understood. In this study, we identified CHIR-regulated genes in mESCs using microarray analysis. Our microarray data demonstrated that CHIR not only influenced the Wnt/β-catenin pathway bymore » stabilizing β-catenin, but also modulated several other pluripotency-related signaling pathways such as TGF-β, Notch and MAPK signaling pathways. More detailed analysis demonstrated that CHIR inhibited Nodal signaling, while activating bone morphogenetic protein signaling in mESCs. In addition, we found that pluripotency-maintaining transcription factors were up-regulated by CHIR, while several developmental-related genes were down-regulated. Furthermore, we found that CHIR altered the expression of epigenetic regulatory genes and long intergenic non-coding RNAs. Quantitative real-time PCR results were consistent with microarray data, suggesting that CHIR alters the expression pattern of protein-encoding genes (especially transcription factors), epigenetic regulatory genes and non-coding RNAs to establish a relatively stable pluripotency-maintaining network. - Highlights: • Combined use of CHIR with LIF promotes self-renewal of J1 mESCs. • CHIR-regulated genes are involved in multiple pathways. • CHIR inhibits Nodal signaling and promotes Bmp4 expression to activate BMP signaling. • Expression of epigenetic regulatory genes and lincRNAs is altered by CHIR.« less

Structural and functional studies of a family of Dictyostelium discoideum developmentally regulated, prestalk genes coding for small proteins.

PubMed

Vicente, Juan J; Galardi-Castilla, María; Escalante, Ricardo; Sastre, Leandro

2008-01-03

The social amoeba Dictyostelium discoideum executes a multicellular development program upon starvation. This morphogenetic process requires the differential regulation of a large number of genes and is coordinated by extracellular signals. The MADS-box transcription factor SrfA is required for several stages of development, including slug migration and spore terminal differentiation. Subtractive hybridization allowed the isolation of a gene, sigN (SrfA-induced gene N), that was dependent on the transcription factor SrfA for expression at the slug stage of development. Homology searches detected the existence of a large family of sigN-related genes in the Dictyostelium discoideum genome. The 13 most similar genes are grouped in two regions of chromosome 2 and have been named Group1 and Group2 sigN genes. The putative encoded proteins are 87-89 amino acids long. All these genes have a similar structure, composed of a first exon containing a 13 nucleotides long open reading frame and a second exon comprising the remaining of the putative coding region. The expression of these genes is induced at10 hours of development. Analyses of their promoter regions indicate that these genes are expressed in the prestalk region of developing structures. The addition of antibodies raised against SigN Group 2 proteins induced disintegration of multi-cellular structures at the mound stage of development. A large family of genes coding for small proteins has been identified in D. discoideum. Two groups of very similar genes from this family have been shown to be specifically expressed in prestalk cells during development. Functional studies using antibodies raised against Group 2 SigN proteins indicate that these genes could play a role during multicellular development.

Pregnancy Complicated by Obesity Induces Global Transcript Expression Alterations in Visceral and Subcutaneous Fat

PubMed Central

Bashiri, Asher; Heo, Hye J.; Ben-Avraham, Danny; Mazor, Moshe; Budagov, Temuri; Einstein, Francine H.; Atzmon, Gil

2014-01-01

Maternal obesity is a significant risk factor for development of both maternal and fetal metabolic complications. Increase in visceral fat and insulin resistance is a metabolic hallmark of pregnancy, yet little is known how obesity alters adipose cellular function and how this may contribute to pregnancy morbidities. We sought to identify alterations in genome-wide transcription expression in both visceral (omental) and abdominal subcutaneous fat deposits in pregnancy complicated by obesity. Visceral and abdominal subcutaneous fat deposits were collected from normal weight and obese pregnant women (n=4/group) at time of scheduled uncomplicated cesarean section. A genome-wide expression array (Affymetrix Human Exon 1.0 st platform), validated by quantitative real-time PCR, was utilized to establish the gene transcript expression profile in both visceral and abdominal subcutaneous fat in normal weight and obese pregnant women. Global alteration in gene expression was identified in pregnancy complicated by obesity. These regions of variations lead to identification of indolethylamine N-methyltransferase (INMT), tissue factor pathway inhibitor-2 (TFPI-2), and ephrin type-B receptor 6 (EPHB6), not previously associated with fat metabolism during pregnancy. In addition, subcutaneous fat of obese pregnant women demonstrated increased coding protein transcripts associated with apoptosis compared to lean counterparts. Global alteration of gene expression in adipose tissue may contribute to adverse pregnancy outcomes associated with obesity. PMID:24696292

The Mediator complex of Caenorhabditis elegans: insights into the developmental and physiological roles of a conserved transcriptional coregulator.

PubMed

Grants, Jennifer M; Goh, Grace Y S; Taubert, Stefan

2015-02-27

The Mediator multiprotein complex ('Mediator') is an important transcriptional coregulator that is evolutionarily conserved throughout eukaryotes. Although some Mediator subunits are essential for the transcription of all protein-coding genes, others influence the expression of only subsets of genes and participate selectively in cellular signaling pathways. Here, we review the current knowledge of Mediator subunit function in the nematode Caenorhabditis elegans, a metazoan in which established and emerging genetic technologies facilitate the study of developmental and physiological regulation in vivo. In this nematode, unbiased genetic screens have revealed critical roles for Mediator components in core developmental pathways such as epidermal growth factor (EGF) and Wnt/β-catenin signaling. More recently, important roles for C. elegans Mediator subunits have emerged in the regulation of lipid metabolism and of systemic stress responses, engaging conserved transcription factors such as nuclear hormone receptors (NHRs). We emphasize instances where similar functions for individual Mediator subunits exist in mammals, highlighting parallels between Mediator subunit action in nematode development and in human cancer biology. We also discuss a parallel between the association of the Mediator subunit MED12 with several human disorders and the role of its C. elegans ortholog mdt-12 as a regulatory hub that interacts with numerous signaling pathways. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

Caregivers' psychosocial factors underlying sugar-sweetened beverage intake among non-Hispanic black preschoolers: an elicitation study.

PubMed

Tipton, Julia A

2014-01-01

The purpose of this study was to explore caregivers' beliefs and perceptions regarding serving sugar-sweetened beverages (SSBs) to non-Hispanic black preschoolers. The Theory of Planned Behavior (TpB) was used as the framework for conducting elicitation interviews among a sample of (n = 19) caregivers. Thematic coding of interview transcripts revealed that the decision to serve SSBs to preschoolers is driven by numerous individual, familial, cultural, and environmental factors. Salient factors associated with serving SSBs included convenience, cost, taste, potential health consequences, availability, and pressure from other parents. Population-specific interventions aimed at reducing SSB intake among non-Hispanic preschoolers are discussed. © 2013.

A long natural-antisense RNA is accumulated in the conidia of Aspergillus oryzae.

PubMed

Tsujii, Masaru; Okuda, Satoshi; Ishi, Kazutomo; Madokoro, Kana; Takeuchi, Michio; Yamagata, Youhei

2016-01-01

Analysis of expressed sequence tag libraries from various culture conditions revealed the existence of conidia-specific transcripts assembled to putative conidiation-specific reductase gene (csrA) in Aspergillus oryzae. However, the all transcripts were transcribed with opposite direction to the gene csrA. The sequence analysis of the transcript revealed that the RNA overlapped mRNA of csrA with 3'-end, and did not code protein longer than 60 amino acid residues. We designated the transcript Conidia Specific Long Natural-antisense RNA (CSLNR). The real-time PCR analysis demonstrated that the CSLNR is conidia-specific transcript, which cannot be transcribed in the absence of brlA, and the amount of CSLNR was much more than that of the transcript from csrA in conidia. Furthermore, the csrA deletion, also lacking coding region of CSLNR in A. oryzae reduced the number of conidia. Overexpression of CsrA demonstrated the inhibition of growth and conidiation, while CSLNR did not affect conidiation.

Transcription and DNA Damage: Holding Hands or Crossing Swords?

PubMed

D'Alessandro, Giuseppina; d'Adda di Fagagna, Fabrizio

2017-10-27

Transcription has classically been considered a potential threat to genome integrity. Collision between transcription and DNA replication machinery, and retention of DNA:RNA hybrids, may result in genome instability. On the other hand, it has been proposed that active genes repair faster and preferentially via homologous recombination. Moreover, while canonical transcription is inhibited in the proximity of DNA double-strand breaks, a growing body of evidence supports active non-canonical transcription at DNA damage sites. Small non-coding RNAs accumulate at DNA double-strand break sites in mammals and other organisms, and are involved in DNA damage signaling and repair. Furthermore, RNA binding proteins are recruited to DNA damage sites and participate in the DNA damage response. Here, we discuss the impact of transcription on genome stability, the role of RNA binding proteins at DNA damage sites, and the function of small non-coding RNAs generated upon damage in the signaling and repair of DNA lesions. Copyright © 2016 Elsevier Ltd. All rights reserved.

The primary transcriptome of the marine diazotroph Trichodesmium erythraeum IMS101

NASA Astrophysics Data System (ADS)

Pfreundt, Ulrike; Kopf, Matthias; Belkin, Natalia; Berman-Frank, Ilana; Hess, Wolfgang R.

2014-08-01

Blooms of the dinitrogen-fixing marine cyanobacterium Trichodesmium considerably contribute to new nitrogen inputs into tropical oceans. Intriguingly, only 60% of the Trichodesmium erythraeum IMS101 genome sequence codes for protein, compared with ~85% in other sequenced cyanobacterial genomes. The extensive non-coding genome fraction suggests space for an unusually high number of unidentified, potentially regulatory non-protein-coding RNAs (ncRNAs). To identify the transcribed fraction of the genome, here we present a genome-wide map of transcriptional start sites (TSS) at single nucleotide resolution, revealing the activity of 6,080 promoters. We demonstrate that T. erythraeum has the highest number of actively splicing group II introns and the highest percentage of TSS yielding ncRNAs of any bacterium examined to date. We identified a highly transcribed retroelement that serves as template repeat for the targeted mutation of at least 12 different genes by mutagenic homing. Our findings explain the non-coding portion of the T. erythraeum genome by the transcription of an unusually high number of non-coding transcripts in addition to the known high incidence of transposable elements. We conclude that riboregulation and RNA maturation-dependent processes constitute a major part of the Trichodesmium regulatory apparatus.

The transcription factor NRSF contributes to epileptogenesis by selective repression of a subset of target genes

PubMed Central

McClelland, Shawn; Brennan, Gary P; Dubé, Celine; Rajpara, Seeta; Iyer, Shruti; Richichi, Cristina; Bernard, Christophe; Baram, Tallie Z

2014-01-01

The mechanisms generating epileptic neuronal networks following insults such as severe seizures are unknown. We have previously shown that interfering with the function of the neuron-restrictive silencer factor (NRSF/REST), an important transcription factor that influences neuronal phenotype, attenuated development of this disorder. In this study, we found that epilepsy-provoking seizures increased the low NRSF levels in mature hippocampus several fold yet surprisingly, provoked repression of only a subset (∼10%) of potential NRSF target genes. Accordingly, the repressed gene-set was rescued when NRSF binding to chromatin was blocked. Unexpectedly, genes selectively repressed by NRSF had mid-range binding frequencies to the repressor, a property that rendered them sensitive to moderate fluctuations of NRSF levels. Genes selectively regulated by NRSF during epileptogenesis coded for ion channels, receptors, and other crucial contributors to neuronal function. Thus, dynamic, selective regulation of NRSF target genes may play a role in influencing neuronal properties in pathological and physiological contexts. DOI: http://dx.doi.org/10.7554/eLife.01267.001 PMID:25117540

Promoter- and RNA polymerase II–dependent hsp-16 gene association with nuclear pores in Caenorhabditis elegans

PubMed Central

Rohner, Sabine; Kalck, Veronique; Wang, Xuefei; Ikegami, Kohta; Lieb, Jason D.; Meister, Peter

2013-01-01

Some inducible yeast genes relocate to nuclear pores upon activation, but the general relevance of this phenomenon has remained largely unexplored. Here we show that the bidirectional hsp-16.2/41 promoter interacts with the nuclear pore complex upon activation by heat shock in the nematode Caenorhabditis elegans. Direct pore association was confirmed by both super-resolution microscopy and chromatin immunoprecipitation. The hsp-16.2 promoter was sufficient to mediate perinuclear positioning under basal level conditions of expression, both in integrated transgenes carrying from 1 to 74 copies of the promoter and in a single-copy genomic insertion. Perinuclear localization of the uninduced gene depended on promoter elements essential for induction and required the heat-shock transcription factor HSF-1, RNA polymerase II, and ENY-2, a factor that binds both SAGA and the THO/TREX mRNA export complex. After induction, colocalization with nuclear pores increased significantly at the promoter and along the coding sequence, dependent on the same promoter-associated factors, including active RNA polymerase II, and correlated with nascent transcripts. PMID:23460676

P-class pentatricopeptide repeat proteins are required for efficient 5′ end formation of plant mitochondrial transcripts

PubMed Central

Binder, Stefan; Stoll, Katrin; Stoll, Birgit

2013-01-01

It is well recognized that flowering plants maintain a particularly broad spectrum of factors to support gene expression in mitochondria. Many of these factors are pentatricopeptide repeat (PPR) proteins that participate in virtually all processes dealing with RNA. One of these processes is the post-transcriptional generation of mature 5′ termini of RNA. Several PPR proteins are required for efficient 5′ maturation of mitochondrial mRNA and rRNA. These so-called RNA PROCESSING FACTORs (RPF) exclusively represent P-class PPR proteins, mainly composed of canonical PPR motifs without any extra domains. Applying the recent PPR-nucleotide recognition code, binding sites of RPF are predicted on the 5′ leader sequences. The sequence-specific interaction of an RPF with one or a few RNA substrates probably directly or indirectly recruits an as-yet-unidentified endonuclease to the processing site(s). The identification and characterization of RPF is a major step toward the understanding of the role of 5′ end maturation in flowering plant mitochondria. PMID:24184847

The evolution of transcriptional regulation in eukaryotes

NASA Technical Reports Server (NTRS)

Wray, Gregory A.; Hahn, Matthew W.; Abouheif, Ehab; Balhoff, James P.; Pizer, Margaret; Rockman, Matthew V.; Romano, Laura A.

2003-01-01

Gene expression is central to the genotype-phenotype relationship in all organisms, and it is an important component of the genetic basis for evolutionary change in diverse aspects of phenotype. However, the evolution of transcriptional regulation remains understudied and poorly understood. Here we review the evolutionary dynamics of promoter, or cis-regulatory, sequences and the evolutionary mechanisms that shape them. Existing evidence indicates that populations harbor extensive genetic variation in promoter sequences, that a substantial fraction of this variation has consequences for both biochemical and organismal phenotype, and that some of this functional variation is sorted by selection. As with protein-coding sequences, rates and patterns of promoter sequence evolution differ considerably among loci and among clades for reasons that are not well understood. Studying the evolution of transcriptional regulation poses empirical and conceptual challenges beyond those typically encountered in analyses of coding sequence evolution: promoter organization is much less regular than that of coding sequences, and sequences required for the transcription of each locus reside at multiple other loci in the genome. Because of the strong context-dependence of transcriptional regulation, sequence inspection alone provides limited information about promoter function. Understanding the functional consequences of sequence differences among promoters generally requires biochemical and in vivo functional assays. Despite these challenges, important insights have already been gained into the evolution of transcriptional regulation, and the pace of discovery is accelerating.

Molecular cloning and analysis of Schizosaccharomyces pombe Reb1p: sequence-specific recognition of two sites in the far upstream rDNA intergenic spacer.

PubMed Central

Zhao, A; Guo, A; Liu, Z; Pape, L

1997-01-01

The coding sequences for a Schizosaccharomyces pombe sequence-specific DNA binding protein, Reb1p, have been cloned. The predicted S. pombe Reb1p is 24-29% identical to mouse TTF-1 (transcription termination factor-1) and Saccharomyces cerevisiae REB1 protein, both of which direct termination of RNA polymerase I catalyzed transcripts. The S.pombe Reb1 cDNA encodes a predicted polypeptide of 504 amino acids with a predicted molecular weight of 58.4 kDa. The S. pombe Reb1p is unusual in that the bipartite DNA binding motif identified originally in S.cerevisiae and Klyveromyces lactis REB1 proteins is uninterrupted and thus S.pombe Reb1p may contain the smallest natural REB1 homologous DNA binding domain. Its genomic coding sequences were shown to be interrupted by two introns. A recombinant histidine-tagged Reb1 protein bearing the rDNA binding domain has two homologous, sequence-specific binding sites in the S. pomber DNA intergenic spacer, located between 289 and 480 nt downstream of the end of the approximately 25S rRNA coding sequences. Each binding site is 13-14 bp downstream of two of the three proposed in vivo termination sites. The core of this 17 bp site, AGGTAAGGGTAATGCAC, is specifically protected by Reb1p in footprinting analysis. PMID:9016645

Assessing Attachment in Psychotherapy: Validation of the Patient Attachment Coding System (PACS).

PubMed

Talia, Alessandro; Miller-Bottome, Madeleine; Daniel, Sarah I F

2017-01-01

The authors present and validate the Patient Attachment Coding System (PACS), a transcript-based instrument that assesses clients' in-session attachment based on any session of psychotherapy, in multiple treatment modalities. One-hundred and sixty clients in different types of psychotherapy (cognitive-behavioural, cognitive-behavioural-enhanced, psychodynamic, relational, supportive) and from three different countries were administered the Adult Attachment Interview (AAI) prior to treatment, and one session for each client was rated with the PACS by independent coders. Results indicate strong inter-rater reliability, and high convergent validity of the PACS scales and classifications with the AAI. These results present the PACS as a practical alternative to the AAI in psychotherapy research and suggest that clinicians using the PACS can assess clients' attachment status on an ongoing basis by monitoring clients' verbal activity. These results also provide information regarding the ways in which differences in attachment status play out in therapy sessions and further the study of attachment in psychotherapy from a pre-treatment client factor to a process variable. Copyright © 2015 John Wiley & Sons, Ltd. The Patient Attachment Coding System is a valid measure of attachment that can classify clients' attachment based on any single psychotherapy transcript, in many therapeutic modalities Client differences in attachment manifest in part independently of the therapist's contributions Client adult attachment patterns are likely to affect psychotherapeutic processes. Copyright © 2015 John Wiley & Sons, Ltd.

Gene end-like sequences within the 3' non-coding region of the Nipah virus genome attenuate viral gene transcription.

PubMed

Sugai, Akihiro; Sato, Hiroki; Yoneda, Misako; Kai, Chieko

2017-08-01

The regulation of transcription during Nipah virus (NiV) replication is poorly understood. Using a bicistronic minigenome system, we investigated the involvement of non-coding regions (NCRs) in the transcriptional re-initiation efficiency of NiV RNA polymerase. Reporter assays revealed that attenuation of NiV gene expression was not constant at each gene junction, and that the attenuating property was controlled by the 3' NCR. However, this regulation was independent of the gene-end, gene-start and intergenic regions. Northern blot analysis indicated that regulation of viral gene expression by the phosphoprotein (P) and large protein (L) 3' NCRs occurred at the transcription level. We identified uridine-rich tracts within the L 3' NCR that are similar to gene-end signals. These gene-end-like sequences were recognized as weak transcription termination signals by the viral RNA polymerase, thereby reducing downstream gene transcription. Thus, we suggest that NiV has a unique mechanism of transcriptional regulation. Copyright © 2017 Elsevier Inc. All rights reserved.

Transcription Factor KLF5 Binds a Cyclin E1 Polymorphic Intronic Enhancer to Confer Increased Bladder Cancer Risk

PubMed Central

Pattison, Jillian M.; Posternak, Valeriya; Cole, Michael D.

2016-01-01

It is well established that environmental toxins, such as exposure to arsenic, are risk factors in the development of urinary bladder cancer, yet recent genome-wide association studies (GWAS) provide compelling evidence that there is a strong genetic component associated with disease predisposition. A single nucleotide polymorphism (SNP), rs8102137, was identified on chromosome 19q12, residing 6 kb upstream of the important cell cycle regulator and proto-oncogene, Cyclin E1 (CCNE1). However, the functional role of this variant in bladder cancer predisposition has been unclear since it lies within a non-coding region of the genome. Here, it is demonstrated that bladder cancer cells heterozygous for this SNP exhibit biased allelic expression of CCNE1 with 1.5-fold more transcription occurring from the risk allele. Furthermore, using chromatin immunoprecipitation assays, a novel enhancer element was identified within the first intron of CCNE1 that binds Kruppel-like Factor 5 (KLF5), a known transcriptional activator in bladder cancer. Moreover, the data reveal that the presence of rs200996365, a SNP in high linkage disequilibrium with rs8102137 residing in the center of a KLF5 motif, alters KLF5 binding to this genomic region. Through luciferase assays and CRISPR-Cas9 genome editing, a novel polymorphic intronic regulatory element controlling CCNE1 transcription is characterized. These studies uncover how a cancer-associated polymorphism mechanistically contributes to an increased predisposition for bladder cancer development. Implications A polymorphic KLF5 binding site near the CCNE1 gene explains genetic risk identified through genome wide association studies. PMID:27514407

Functional interrogation of non-coding DNA through CRISPR genome editing.

PubMed

Canver, Matthew C; Bauer, Daniel E; Orkin, Stuart H

2017-05-15

Methodologies to interrogate non-coding regions have lagged behind coding regions despite comprising the vast majority of the genome. However, the rapid evolution of clustered regularly interspaced short palindromic repeats (CRISPR)-based genome editing has provided a multitude of novel techniques for laboratory investigation including significant contributions to the toolbox for studying non-coding DNA. CRISPR-mediated loss-of-function strategies rely on direct disruption of the underlying sequence or repression of transcription without modifying the targeted DNA sequence. CRISPR-mediated gain-of-function approaches similarly benefit from methods to alter the targeted sequence through integration of customized sequence into the genome as well as methods to activate transcription. Here we review CRISPR-based loss- and gain-of-function techniques for the interrogation of non-coding DNA. Copyright © 2017 Elsevier Inc. All rights reserved.

Regulatory activities of transposable elements: from conflicts to benefits

PubMed Central

Chuong, Edward B.; Elde, Nels C.; Feschotte, Cédric

2017-01-01

Transposable elements (TEs) are a prolific source of tightly regulated, biochemically active non-coding elements, such as transcription factor binding sites and non-coding RNAs. A wealth of recent studies reinvigorates the idea that these elements are pervasively co-opted for the regulation of host genes. We argue that the inherent genetic properties of TEs and conflicting relationships with their hosts facilitate their recruitment for regulatory functions in diverse genomes. We review recent findings supporting the long-standing hypothesis that the waves of TE invasions endured by organisms for eons have catalyzed the evolution of gene regulatory networks. We also discuss the challenges of dissecting and interpreting the phenotypic impact of regulatory activities encoded by TEs in health and disease. PMID:27867194

Bovine Herpes Virus 1 (BHV-1) and Herpes Simplex Virus Type 1 (HSV-1) Promote Survival of Latently Infected Sensory Neurons, in Part by Inhibiting Apoptosis

PubMed Central

Jones, Clinton

2013-01-01

α-Herpesvirinae subfamily members, including herpes simplex virus type 1 (HSV-1) and bovine herpes virus 1 (BHV-1), initiate infection in mucosal surfaces. BHV-1 and HSV-1 enter sensory neurons by cell-cell spread where a burst of viral gene expression occurs. When compared to non-neuronal cells, viral gene expression is quickly extinguished in sensory neurons resulting in neuronal survival and latency. The HSV-1 latency associated transcript (LAT), which is abundantly expressed in latently infected neurons, inhibits apoptosis, viral transcription, and productive infection, and directly or indirectly enhances reactivation from latency in small animal models. Three anti-apoptosis genes can be substituted for LAT, which will restore wild type levels of reactivation from latency to a LAT null mutant virus. Two small non-coding RNAs encoded by LAT possess anti-apoptosis functions in transfected cells. The BHV-1 latency related RNA (LR-RNA), like LAT, is abundantly expressed during latency. The LR-RNA encodes a protein (ORF2) and two microRNAs that are expressed in certain latently infected neurons. Wild-type expression of LR gene products is required for stress-induced reactivation from latency in cattle. ORF2 has anti-apoptosis functions and interacts with certain cellular transcription factors that stimulate viral transcription and productive infection. ORF2 is predicted to promote survival of infected neurons by inhibiting apoptosis and sequestering cellular transcription factors which stimulate productive infection. In addition, the LR encoded microRNAs inhibit viral transcription and apoptosis. In summary, the ability of BHV-1 and HSV-1 to interfere with apoptosis and productive infection in sensory neurons is crucial for the life-long latency-reactivation cycle in their respective hosts. PMID:25278776

Natural antisense transcripts associated with salinity response in alfalfa

USDA-ARS?s Scientific Manuscript database

Natural antisense transcripts (NATs) are long non-coding RNAs (lncRNAs) complimentary to the messenger (sense) RNA (Wang et al. 2014). Many of them are involved in regulation of their own sense transcripts thus playing pivotal biological roles in all processes of organismal development and responses...

Regulated Formation of lncRNA-DNA Hybrids Enables Faster Transcriptional Induction and Environmental Adaptation.

PubMed

Cloutier, Sara C; Wang, Siwen; Ma, Wai Kit; Al Husini, Nadra; Dhoondia, Zuzer; Ansari, Athar; Pascuzzi, Pete E; Tran, Elizabeth J

2016-02-04

Long non-coding (lnc)RNAs, once thought to merely represent noise from imprecise transcription initiation, have now emerged as major regulatory entities in all eukaryotes. In contrast to the rapidly expanding identification of individual lncRNAs, mechanistic characterization has lagged behind. Here we provide evidence that the GAL lncRNAs in the budding yeast S. cerevisiae promote transcriptional induction in trans by formation of lncRNA-DNA hybrids or R-loops. The evolutionarily conserved RNA helicase Dbp2 regulates formation of these R-loops as genomic deletion or nuclear depletion results in accumulation of these structures across the GAL cluster gene promoters and coding regions. Enhanced transcriptional induction is manifested by lncRNA-dependent displacement of the Cyc8 co-repressor and subsequent gene looping, suggesting that these lncRNAs promote induction by altering chromatin architecture. Moreover, the GAL lncRNAs confer a competitive fitness advantage to yeast cells because expression of these non-coding molecules correlates with faster adaptation in response to an environmental switch. Copyright © 2016 Elsevier Inc. All rights reserved.

Regulatory consequences of neuronal ELAV-like protein binding to coding and non-coding RNAs in human brain

PubMed Central

Scheckel, Claudia; Drapeau, Elodie; Frias, Maria A; Park, Christopher Y; Fak, John; Zucker-Scharff, Ilana; Kou, Yan; Haroutunian, Vahram; Ma'ayan, Avi

2016-01-01

Neuronal ELAV-like (nELAVL) RNA binding proteins have been linked to numerous neurological disorders. We performed crosslinking-immunoprecipitation and RNAseq on human brain, and identified nELAVL binding sites on 8681 transcripts. Using knockout mice and RNAi in human neuroblastoma cells, we showed that nELAVL intronic and 3' UTR binding regulates human RNA splicing and abundance. We validated hundreds of nELAVL targets among which were important neuronal and disease-associated transcripts, including Alzheimer's disease (AD) transcripts. We therefore investigated RNA regulation in AD brain, and observed differential splicing of 150 transcripts, which in some cases correlated with differential nELAVL binding. Unexpectedly, the most significant change of nELAVL binding was evident on non-coding Y RNAs. nELAVL/Y RNA complexes were specifically remodeled in AD and after acute UV stress in neuroblastoma cells. We propose that the increased nELAVL/Y RNA association during stress may lead to nELAVL sequestration, redistribution of nELAVL target binding, and altered neuronal RNA splicing. DOI: http://dx.doi.org/10.7554/eLife.10421.001 PMID:26894958

Topic models: A novel method for modeling couple and family text data

PubMed Central

Atkins, David C.; Rubin, Tim N.; Steyvers, Mark; Doeden, Michelle A.; Baucom, Brian R.; Christensen, Andrew

2012-01-01

Couple and family researchers often collect open-ended linguistic data – either through free response questionnaire items or transcripts of interviews or therapy sessions. Because participant's responses are not forced into a set number of categories, text-based data can be very rich and revealing of psychological processes. At the same time it is highly unstructured and challenging to analyze. Within family psychology analyzing text data typically means applying a coding system, which can quantify text data but also has several limitations, including the time needed for coding, difficulties with inter-rater reliability, and defining a priori what should be coded. The current article presents an alternative method for analyzing text data called topic models (Steyvers & Griffiths, 2006), which has not yet been applied within couple and family psychology. Topic models have similarities with factor analysis and cluster analysis in that topic models identify underlying clusters of words with semantic similarities (i.e., the “topics”). In the present article, a non-technical introduction to topic models is provided, highlighting how these models can be used for text exploration and indexing (e.g., quickly locating text passages that share semantic meaning) and how output from topic models can be used to predict behavioral codes or other types of outcomes. Throughout the article a collection of transcripts from a large couple therapy trial (Christensen et al., 2004) is used as example data to highlight potential applications. Practical resources for learning more about topic models and how to apply them are discussed. PMID:22888778

Topic models: a novel method for modeling couple and family text data.

PubMed

Atkins, David C; Rubin, Timothy N; Steyvers, Mark; Doeden, Michelle A; Baucom, Brian R; Christensen, Andrew

2012-10-01

Couple and family researchers often collect open-ended linguistic data-either through free-response questionnaire items, or transcripts of interviews or therapy sessions. Because participants' responses are not forced into a set number of categories, text-based data can be very rich and revealing of psychological processes. At the same time, it is highly unstructured and challenging to analyze. Within family psychology, analyzing text data typically means applying a coding system, which can quantify text data but also has several limitations, including the time needed for coding, difficulties with interrater reliability, and defining a priori what should be coded. The current article presents an alternative method for analyzing text data called topic models (Steyvers & Griffiths, 2006), which has not yet been applied within couple and family psychology. Topic models have similarities to factor analysis and cluster analysis in that they identify underlying clusters of words with semantic similarities (i.e., the "topics"). In the present article, a nontechnical introduction to topic models is provided, highlighting how these models can be used for text exploration and indexing (e.g., quickly locating text passages that share semantic meaning) and how output from topic models can be used to predict behavioral codes or other types of outcomes. Throughout the article, a collection of transcripts from a large couple-therapy trial (Christensen et al., 2004) is used as example data to highlight potential applications. Practical resources for learning more about topic models and how to apply them are discussed. (PsycINFO Database Record (c) 2012 APA, all rights reserved).

The Non-Coding RNA Ncr0700/PmgR1 is Required for Photomixotrophic Growth and the Regulation of Glycogen Accumulation in the Cyanobacterium Synechocystis sp. PCC 6803.

PubMed

de Porcellinis, Alice J; Klähn, Stephan; Rosgaard, Lisa; Kirsch, Rebekka; Gutekunst, Kirstin; Georg, Jens; Hess, Wolfgang R; Sakuragi, Yumiko

2016-10-01

Carbohydrate metabolism is a tightly regulated process in photosynthetic organisms. In the cyanobacterium Synechocystis sp. PCC 6803, the photomixotrophic growth protein A (PmgA) is involved in the regulation of glucose and storage carbohydrate (i.e. glycogen) metabolism, while its biochemical activity and possible factors acting downstream of PmgA are unknown. Here, a genome-wide microarray analysis of a ΔpmgA strain identified the expression of 36 protein-coding genes and 42 non-coding transcripts as significantly altered. From these, the non-coding RNA Ncr0700 was identified as the transcript most strongly reduced in abundance. Ncr0700 is widely conserved among cyanobacteria. In Synechocystis its expression is inversely correlated with light intensity. Similarly to a ΔpmgA mutant, a Δncr0700 deletion strain showed an approximately 2-fold increase in glycogen content under photoautotrophic conditions and wild-type-like growth. Moreover, its growth was arrested by 38 h after a shift to photomixotrophic conditions. Ectopic expression of Ncr0700 in Δncr0700 and ΔpmgA restored the glycogen content and photomixotrophic growth to wild-type levels. These results indicate that Ncr0700 is required for photomixotrophic growth and the regulation of glycogen accumulation, and acts downstream of PmgA. Hence Ncr0700 is renamed here as PmgR1 for photomixotrophic growth RNA 1. © The Author 2016. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Mitochondrial transcription factor A (Tfam) gene sequencing and mitochondrial evaluation in inherited retinal dysplasia in miniature schnauzer dogs

PubMed Central

Bauer, Bianca S.; Forsyth, George W.; Sandmeyer, Lynne S.; Grahn, Bruce H.

2011-01-01

Mitochondrial transcription factor A (Tfam) has been implicated in the pathogenesis of retinal dysplasia in miniature schnauzer dogs and it has been proposed that affected dogs have altered mitochondrial numbers, size, and morphology. To test these hypotheses the Tfam gene of affected and normal miniature schnauzer dogs with retinal dysplasia was sequenced and lymphocyte mitochondria were quantified, measured, and the morphology was compared in normal and affected dogs using transmission electron microscopy. For Tfam sequencing, retina, retinal pigment epithelium (RPE), and whole blood samples were collected. Total RNA was isolated from the retina and RPE and reverse transcribed to make cDNA. Genomic DNA was extracted from white blood cell pellets obtained from the whole blood samples. The Tfam coding sequence, 5′ promoter region, intron1 and the 3′ non-coding sequence of normal and affected dogs were amplified using polymerase chain reaction (PCR), cloned and sequenced. For electron microscopy, lymphocytes from affected and normal dogs were photographed and the mitochondria within each cross-section were identified, quantified, and the mitochondrial area (μm2) per lymphocyte cross-section was calculated. Lastly, using a masked technique, mitochondrial morphology was compared between the 2 groups. Sequencing of the miniature schnauzer Tfam gene revealed no functional sequence variation between affected and normal dogs. Lymphocyte and mitochondrial area, mitochondrial quantification, and morphology assessment also revealed no significant difference between the 2 groups. Further investigation into other candidate genes or factors causing retinal dysplasia in the miniature schnauzer is warranted. PMID:21731185

Mitochondrial transcription factor A (Tfam) gene sequencing and mitochondrial evaluation in inherited retinal dysplasia in miniature schnauzer dogs.

PubMed

Bauer, Bianca S; Forsyth, George W; Sandmeyer, Lynne S; Grahn, Bruce H

2011-04-01

Mitochondrial transcription factor A (Tfam) has been implicated in the pathogenesis of retinal dysplasia in miniature schnauzer dogs and it has been proposed that affected dogs have altered mitochondrial numbers, size, and morphology. To test these hypotheses the Tfam gene of affected and normal miniature schnauzer dogs with retinal dysplasia was sequenced and lymphocyte mitochondria were quantified, measured, and the morphology was compared in normal and affected dogs using transmission electron microscopy. For Tfam sequencing, retina, retinal pigment epithelium (RPE), and whole blood samples were collected. Total RNA was isolated from the retina and RPE and reverse transcribed to make cDNA. Genomic DNA was extracted from white blood cell pellets obtained from the whole blood samples. The Tfam coding sequence, 5' promoter region, intron1 and the 3' non-coding sequence of normal and affected dogs were amplified using polymerase chain reaction (PCR), cloned and sequenced. For electron microscopy, lymphocytes from affected and normal dogs were photographed and the mitochondria within each cross-section were identified, quantified, and the mitochondrial area (μm²) per lymphocyte cross-section was calculated. Lastly, using a masked technique, mitochondrial morphology was compared between the 2 groups. Sequencing of the miniature schnauzer Tfam gene revealed no functional sequence variation between affected and normal dogs. Lymphocyte and mitochondrial area, mitochondrial quantification, and morphology assessment also revealed no significant difference between the 2 groups. Further investigation into other candidate genes or factors causing retinal dysplasia in the miniature schnauzer is warranted.

Functional Characterization of the Poplar R2R3-MYB Transcription Factor PtoMYB216 Involved in the Regulation of Lignin Biosynthesis during Wood Formation

PubMed Central

Lu, Wanxiang; Yang, Li; Jiang, Yuanzhong; Luo, Keming

2013-01-01

Because of the importance of wood in many industrial applications, tremendous studies have been performed on wood formation, especially in lignin biosynthesis. MYB transcription factors (TFs), which consist of a large family of plant TFs, have been reported to directly regulate lignin biosynthetic genes in a number of plants. In this study, we describe the cloning and functional characterization of PtoMYB216, a cDNA isolated from Chinese white poplar (Populus tomentosa Carr.). PtoMYB216 encodes a protein belonging to the R2R3-MYB family and displays significant similarity with other MYB factors shown to regulate lignin synthesis in Arabidopsis. Gene expression profiling studies showed that PtoMYB216 mRNA is specifically expressed during secondary wall formation in wood. The 1.8-kb promoter sequence of PtoMYB216 was fused to the GUS coding sequence and introduced into wild-type A. thaliana. GUS expression was shown to be restricted to tissues undergoing secondary cell wall formation. Overexpression of PtoMYB216 specifically activated the expression of the upstream genes in the lignin biosynthetic pathway and resulted in ectopic deposition of lignin in cells that are normally unligninified. These results suggest that PtoMYB216 is specific transcriptional activators of lignin biosynthesis and involved in the regulation of wood formation in poplar. PMID:24204619

Identification and characterization of microRNAs and their targets in high-altitude stress-adaptive plant maca (Lepidium meyenii Walp).

PubMed

Paul, Sujay

2017-06-01

MicroRNAs (miRNAs) are endogenous, short (~21-nucleotide), non-coding RNA molecules that play pivotal roles in plant growth, development, and stress response signaling. In this study using recently published draft genome sequence of a high-altitude plant maca (Lepidium meyenii Walp) and applying genome-wide computational-based approaches, a total of 62 potentially conserved miRNAs belonging to 28 families were identified and four (lme-miR160a, lme-miR164c, lme-miR 166a, and lme-miR 319a) of them further validated by RT-PCR. Deploying psRNATarget tool a total of 99 potential miRNA target transcripts were also identified in maca. Targets include a number of transcription factors like Squamosa promoter-binding, NAC, MYB, auxin response factor, APETALA, WRKY, and F-box protein. To the best of my knowledge, this is the first genome-based miRNA profiling of a high-altitude plant.

OnTheFly: a database of Drosophila melanogaster transcription factors and their binding sites.

PubMed

Shazman, Shula; Lee, Hunjoong; Socol, Yakov; Mann, Richard S; Honig, Barry

2014-01-01

We present OnTheFly (http://bhapp.c2b2.columbia.edu/OnTheFly/index.php), a database comprising a systematic collection of transcription factors (TFs) of Drosophila melanogaster and their DNA-binding sites. TFs predicted in the Drosophila melanogaster genome are annotated and classified and their structures, obtained via experiment or homology models, are provided. All known preferred TF DNA-binding sites obtained from the B1H, DNase I and SELEX methodologies are presented. DNA shape parameters predicted for these sites are obtained from a high throughput server or from crystal structures of protein-DNA complexes where available. An important feature of the database is that all DNA-binding domains and their binding sites are fully annotated in a eukaryote using structural criteria and evolutionary homology. OnTheFly thus provides a comprehensive view of TFs and their binding sites that will be a valuable resource for deciphering non-coding regulatory DNA.

Genome-wide analysis of the DNA-binding with one zinc finger (Dof) transcription factor family in bananas.

PubMed

Dong, Chen; Hu, Huigang; Xie, Jianghui

2016-12-01

DNA-binding with one finger (Dof) domain proteins are a multigene family of plant-specific transcription factors involved in numerous aspects of plant growth and development. In this study, we report a genome-wide search for Musa acuminata Dof (MaDof) genes and their expression profiles at different developmental stages and in response to various abiotic stresses. In addition, a complete overview of the Dof gene family in bananas is presented, including the gene structures, chromosomal locations, cis-regulatory elements, conserved protein domains, and phylogenetic inferences. Based on the genome-wide analysis, we identified 74 full-length protein-coding MaDof genes unevenly distributed on 11 chromosomes. Phylogenetic analysis with Dof members from diverse plant species showed that MaDof genes can be classified into four subgroups (StDof I, II, III, and IV). The detailed genomic information of the MaDof gene homologs in the present study provides opportunities for functional analyses to unravel the exact role of the genes in plant growth and development.

CCCTC-Binding Factor Acts as a Heterochromatin Barrier on Herpes Simplex Viral Latent Chromatin and Contributes to Poised Latent Infection

PubMed Central

2018-01-01

ABSTRACT Herpes simplex virus 1 (HSV-1) establishes latent infection in neurons via a variety of epigenetic mechanisms that silence its genome. The cellular CCCTC-binding factor (CTCF) functions as a mediator of transcriptional control and chromatin organization and has binding sites in the HSV-1 genome. We constructed an HSV-1 deletion mutant that lacked a pair of CTCF-binding sites (CTRL2) within the latency-associated transcript (LAT) coding sequences and found that loss of these CTCF-binding sites did not alter lytic replication or levels of establishment of latent infection, but their deletion reduced the ability of the virus to reactivate from latent infection. We also observed increased heterochromatin modifications on viral chromatin over the LAT promoter and intron. We therefore propose that CTCF binding at the CTRL2 sites acts as a chromatin insulator to keep viral chromatin in a form that is poised for reactivation, a state which we call poised latency. PMID:29437926

Human papillomavirus 16 E6 modulates the expression of miR-496 in oropharyngeal cancer.

PubMed

Mason, Dayna; Zhang, Xiaoying; Marques, Tânia Monteiro; Rose, Barbara; Khoury, Samantha; Hill, Meredith; Deutsch, Fiona; Lyons, J Guy; Gama-Carvalho, Margarida; Tran, Nham

2018-06-20

Human papillomavirus (HPV), notably type 16, is a risk factor for up to 75% of oropharyngeal squamous cell carcinomas (SCC). It has been demonstrated that small non-coding RNAs known as microRNAs play a vital role in the cellular transformation process. In this study, we used an LNA array to further investigate the impact of HPV16 on the expression of microRNAs in oropharyngeal (tonsillar) cancer. A number of miRNAs were found to be deregulated, with miR-496 showing a four-fold decrease. Over-expression of the high risk E6 oncoprotein down-regulated miR-496, impacting upon the post-transcriptional control of the transcription factor E2F2. These HPV specific miRNAs were integrated with the HPV16 interactome to identify possible mechanistic pathways. These analyses provide insights into novel molecular interactions between HPV16 and miRNAs in oropharyngeal cancers. Copyright © 2018. Published by Elsevier Inc.

miRNA-dependent gene silencing involving Ago2-mediated cleavage of a circular antisense RNA

PubMed Central

Hansen, Thomas B; Wiklund, Erik D; Bramsen, Jesper B; Villadsen, Sune B; Statham, Aaron L; Clark, Susan J; Kjems, Jørgen

2011-01-01

MicroRNAs (miRNAs) are ∼22 nt non-coding RNAs that typically bind to the 3′ UTR of target mRNAs in the cytoplasm, resulting in mRNA destabilization and translational repression. Here, we report that miRNAs can also regulate gene expression by targeting non-coding antisense transcripts in human cells. Specifically, we show that miR-671 directs cleavage of a circular antisense transcript of the Cerebellar Degeneration-Related protein 1 (CDR1) locus in an Ago2-slicer-dependent manner. The resulting downregulation of circular antisense has a concomitant decrease in CDR1 mRNA levels, independently of heterochromatin formation. This study provides the first evidence for non-coding antisense transcripts as functional miRNA targets, and a novel regulatory mechanism involving a positive correlation between mRNA and antisense circular RNA levels. PMID:21964070

Long non-coding RNAs and their biological roles in plants.

PubMed

Liu, Xue; Hao, Lili; Li, Dayong; Zhu, Lihuang; Hu, Songnian

2015-06-01

With the development of genomics and bioinformatics, especially the extensive applications of high-throughput sequencing technology, more transcriptional units with little or no protein-coding potential have been discovered. Such RNA molecules are called non-protein-coding RNAs (npcRNAs or ncRNAs). Among them, long npcRNAs or ncRNAs (lnpcRNAs or lncRNAs) represent diverse classes of transcripts longer than 200 nucleotides. In recent years, the lncRNAs have been considered as important regulators in many essential biological processes. In plants, although a large number of lncRNA transcripts have been predicted and identified in few species, our current knowledge of their biological functions is still limited. Here, we have summarized recent studies on their identification, characteristics, classification, bioinformatics, resources, and current exploration of their biological functions in plants. Copyright © 2015 The Authors. Production and hosting by Elsevier Ltd.. All rights reserved.

Technological Developments in lncRNA Biology.

PubMed

Jathar, Sonali; Kumar, Vikram; Srivastava, Juhi; Tripathi, Vidisha

2017-01-01

It is estimated that more than 90% of the mammalian genome is transcribed as non-coding RNAs. Recent evidences have established that these non-coding transcripts are not junk or just transcriptional noise, but they do serve important biological purpose. One of the rapidly expanding fields of this class of transcripts is the regulatory lncRNAs, which had been a major challenge in terms of their molecular functions and mechanisms of action. The emergence of high-throughput technologies and the development in various conventional approaches have led to the expansion of the lncRNA world. The combination of multidisciplinary approaches has proven to be essential to unravel the complexity of their regulatory networks and helped establish the importance of their existence. Here, we review the current methodologies available for discovering and investigating functions of long non-coding RNAs (lncRNAs) and focus on the powerful technological advancement available to specifically address their functional importance.

No involvement of the nerve growth factor gene locus in hypertension in spontaneously hypertensive rats.

PubMed

Nemoto, Kiyomitsu; Sekimoto, Masashi; Fukamachi, Katsumi; Kageyama, Haruaki; Degawa, Masakuni; Hamadai, Masanori; Hendley, Edith D; Macrae, I Mhairi; Clark, James S; Dominiczak, Anna F; Ueyama, Takashi

2005-02-01

Sympathetic hyper-innervation and increased levels of nerve growth factor (NGF), an essential neurotrophic factor for sympathetic neurons, have been observed in the vascular tissues of spontaneously hypertensive rats (SHRs). Such observations have suggested that the pathogenesis of hypertension might involve a qualitative or quantitative abnormality in the NGF protein, resulting from a significant mutation in the gene's promoter or coding region. In the present study, we analyzed the nucleotide sequences of the cis-element of the NGF gene in SHRs, stroke-prone SHRs (SHRSPs), and normotensive Wistar-Kyoto (WKY) rats. The present analyses revealed some differences in the 3-kb promoter region, coding exon, and 3' untranslated region (3'UTR) for the NGF gene among those strains. However, the observed differences did not lead to changes in promoter activity or to amino acid substitution; nor did they represent a link between the 3'UTR mutation of SHRSPs and elevated blood pressure in an F2 generation produced by crossbreeding SHRSPs with WKY rats. These results suggest that the NGF gene locus is not involved in hypertension in SHR/ SHRSP strains. The present study also revealed two differences between SHRs and WKY rats, as found in cultured vascular smooth muscle cells and in mRNA prepared from each strain. First, SHRs had higher expression levels of c-fos and c-jun genes, which encode the component of the AP-1 transcription factor that activates NGF gene transcription. Second, NGF mRNAs prepared from SHRs had a longer 3'UTR than those prepared from WKY rats. Although it remains to be determined whether these events play a role in the hypertension of SHR/SHRSP strains, the present results emphasize the importance of actively searching for aberrant trans-acting factor(s) leading to the enhanced expression of the NGF gene and NGF protein in SHR/SHRSP strains.

Epigenetic regulation of TTF-I-mediated promoter–terminator interactions of rRNA genes

PubMed Central

Németh, Attila; Guibert, Sylvain; Tiwari, Vijay Kumar; Ohlsson, Rolf; Längst, Gernot

2008-01-01

Ribosomal RNA synthesis is the eukaryotic cell's main transcriptional activity, but little is known about the chromatin domain organization and epigenetics of actively transcribed rRNA genes. Here, we show epigenetic and spatial organization of mouse rRNA genes at the molecular level. TTF-I-binding sites subdivide the rRNA transcription unit into functional chromatin domains and sharply delimit transcription factor occupancy. H2A.Z-containing nucleosomes occupy the spacer promoter next to a newly characterized TTF-I-binding site. The spacer and the promoter proximal TTF-I-binding sites demarcate the enhancer. DNA from both the enhancer and the coding region is hypomethylated in actively transcribed repeats. 3C analysis revealed an interaction between promoter and terminator regions, which brings the beginning and end of active rRNA genes into close contact. Reporter assays show that TTF-I mediates this interaction, thereby linking topology and epigenetic regulation of the rRNA genes. PMID:18354495

Proteomic Analysis of Mitotic RNA Polymerase II Reveals Novel Interactors and Association With Proteins Dysfunctional in Disease*

PubMed Central

Möller, André; Xie, Sheila Q.; Hosp, Fabian; Lang, Benjamin; Phatnani, Hemali P.; James, Sonya; Ramirez, Francisco; Collin, Gayle B.; Naggert, Jürgen K.; Babu, M. Madan; Greenleaf, Arno L.; Selbach, Matthias; Pombo, Ana

2012-01-01

RNA polymerase II (RNAPII) transcribes protein-coding genes in eukaryotes and interacts with factors involved in chromatin remodeling, transcriptional activation, elongation, and RNA processing. Here, we present the isolation of native RNAPII complexes using mild extraction conditions and immunoaffinity purification. RNAPII complexes were extracted from mitotic cells, where they exist dissociated from chromatin. The proteomic content of native complexes in total and size-fractionated extracts was determined using highly sensitive LC-MS/MS. Protein associations with RNAPII were validated by high-resolution immunolocalization experiments in both mitotic cells and in interphase nuclei. Functional assays of transcriptional activity were performed after siRNA-mediated knockdown. We identify >400 RNAPII associated proteins in mitosis, among these previously uncharacterized proteins for which we show roles in transcriptional elongation. We also identify, as novel functional RNAPII interactors, two proteins involved in human disease, ALMS1 and TFG, emphasizing the importance of gene regulation for normal development and physiology. PMID:22199231

Genome-wide transcriptional profiling of human glioblastoma cells in response to ITE treatment

PubMed Central

Kang, Bo; Zhou, Yanwen; Zheng, Min; Wang, Ying-Jie

2015-01-01

A ligand-activated transcription factor aryl hydrocarbon receptor (AhR) is recently revealed to play a key role in embryogenesis and tumorigenesis (Feng et al. [1], Safe et al. [2]) and 2-(1′H-indole-3′-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) (Song et al. [3]) is an endogenous AhR ligand that possesses anti-tumor activity. In order to gain insights into how ITE acts via the AhR in embryogenesis and tumorigenesis, we analyzed the genome-wide transcriptional profiles of the following three groups of cells: the human glioblastoma U87 parental cells, U87 tumor sphere cells treated with vehicle (DMSO) and U87 tumor sphere cells treated with ITE. Here, we provide the details of the sample gathering strategy and show the quality controls and the analyses associated with our gene array data deposited into the Gene Expression Omnibus (GEO) under the accession code of GSE67986. PMID:26484269

Retrotransposon Tf1 is targeted to pol II promoters by transcription activators

PubMed Central

Leem, Young-Eun; Ripmaster, Tracy; Kelly, Felice; Ebina, Hirotaka; Heincelman, Marc; Zhang, Ke; Grewal, Shiv I. S.; Hoffman, Charles S.; Levin, Henry L.

2008-01-01

SUMMARY The LTR-retrotransposon Tf1 preserves the coding capacity of its host Schizosaccharomyces pombe by integrating upstream of open reading frames (ORFs). To determine which features of the target sites were recognized by the transposon, we introduced plasmids containing candidate insertion sites into S. pombe and mapped the positions of integration. We found that Tf1 was targeted specifically to the promoters of pol II transcribed genes. A detailed analysis of integration in plasmids that contained either ade6 or fbp1 revealed insertions occurred in the promoters at positions where transcription factors bound. Further experiments revealed that the activator Atf1p and its binding site were required for directing integration to the promoter of fbp1. An interaction between Tf1 integrase and Atf1p was observed indicating that integration at fbp1 was mediated by the activator bound to its promoter. Surprisingly we found Tf1 contained sequences that activated transcription and these substituted for elements of the ade6 promoter disrupted by integration. PMID:18406330

Retrotransposon Tf1 is targeted to Pol II promoters by transcription activators.

PubMed

Leem, Young-Eun; Ripmaster, Tracy L; Kelly, Felice D; Ebina, Hirotaka; Heincelman, Marc E; Zhang, Ke; Grewal, Shiv I S; Hoffman, Charles S; Levin, Henry L

2008-04-11

The LTR-retrotransposon Tf1 preserves the coding capacity of its host Schizosaccharomyces pombe by integrating upstream of open reading frames (ORFs). To determine which features of the target sites were recognized by the transposon, we introduced plasmids containing candidate insertion sites into S. pombe and mapped the positions of integration. We found that Tf1 was targeted specifically to the promoters of Pol II-transcribed genes. A detailed analysis of integration in plasmids that contained either ade6 or fbp1 revealed insertions occurred in the promoters at positions where transcription factors bound. Further experiments revealed that the activator Atf1p and its binding site were required for directing integration to the promoter of fbp1. An interaction between Tf1 integrase and Atf1p was observed, indicating that integration at fbp1 was mediated by the activator bound to its promoter. Surprisingly, we found Tf1 contained sequences that activated transcription, and these substituted for elements of the ade6 promoter disrupted by integration.

Single-cell topological RNA-Seq analysis reveals insights into cellular differentiation and development

PubMed Central

Rizvi, Abbas H.; Camara, Pablo G.; Kandror, Elena K.; Roberts, Thomas J.; Schieren, Ira; Maniatis, Tom; Rabadan, Raul

2017-01-01

Transcriptional programs control cellular lineage commitment and differentiation during development. Understanding cell fate has been advanced by studying single-cell RNA-seq, but is limited by the assumptions of current analytic methods regarding the structure of data. We present single-cell topological data analysis (scTDA), an algorithm for topology-based computational analyses to study temporal, unbiased transcriptional regulation. Compared to other methods, scTDA is a non-linear, model-independent, unsupervised statistical framework that can characterize transient cellular states. We applied scTDA to the analysis of murine embryonic stem cell (mESC) differentiation in vitro in response to inducers of motor neuron differentiation. scTDA resolved asynchrony and continuity in cellular identity over time, and identified four transient states (pluripotent, precursor, progenitor, and fully differentiated cells) based on changes in stage-dependent combinations of transcription factors, RNA-binding proteins and long non-coding RNAs. scTDA can be applied to study asynchronous cellular responses to either developmental cues or environmental perturbations. PMID:28459448

RNA-directed DNA methylation involves co-transcriptional small-RNA-guided slicing of polymerase V transcripts in Arabidopsis.

PubMed

Liu, Wanlu; Duttke, Sascha H; Hetzel, Jonathan; Groth, Martin; Feng, Suhua; Gallego-Bartolome, Javier; Zhong, Zhenhui; Kuo, Hsuan Yu; Wang, Zonghua; Zhai, Jixian; Chory, Joanne; Jacobsen, Steven E

2018-03-01

Small RNAs regulate chromatin modifications such as DNA methylation and gene silencing across eukaryotic genomes. In plants, RNA-directed DNA methylation (RdDM) requires 24-nucleotide small interfering RNAs (siRNAs) that bind to ARGONAUTE 4 (AGO4) and target genomic regions for silencing. RdDM also requires non-coding RNAs transcribed by RNA polymerase V (Pol V) that probably serve as scaffolds for binding of AGO4-siRNA complexes. Here, we used a modified global nuclear run-on protocol followed by deep sequencing to capture Pol V nascent transcripts genome-wide. We uncovered unique characteristics of Pol V RNAs, including a uracil (U) common at position 10. This uracil was complementary to the 5' adenine found in many AGO4-bound 24-nucleotide siRNAs and was eliminated in a siRNA-deficient mutant as well as in the ago4/6/9 triple mutant, suggesting that the +10 U signature is due to siRNA-mediated co-transcriptional slicing of Pol V transcripts. Expression of wild-type AGO4 in ago4/6/9 mutants was able to restore slicing of Pol V transcripts, but a catalytically inactive AGO4 mutant did not correct the slicing defect. We also found that Pol V transcript slicing required SUPPRESSOR OF TY INSERTION 5-LIKE (SPT5L), an elongation factor whose function is not well understood. These results highlight the importance of Pol V transcript slicing in RNA-mediated transcriptional gene silencing, which is a conserved process in many eukaryotes.

Identification of a novel herpes simplex virus type 1 transcript and protein (AL3) expressed during latency.

PubMed

Jaber, Tareq; Henderson, Gail; Li, Sumin; Perng, Guey-Chuen; Carpenter, Dale; Wechsler, Steven L; Jones, Clinton

2009-10-01

The herpes simplex virus type 1 (HSV-1) latency-associated transcript (LAT) is abundantly expressed in latently infected sensory neurons. In small animal models of infection, expression of the first 1.5 kb of LAT coding sequences is necessary and sufficient for wild-type reactivation from latency. The ability of LAT to inhibit apoptosis is important for reactivation from latency. Within the first 1.5 kb of LAT coding sequences and LAT promoter sequences, additional transcripts have been identified. For example, the anti-sense to LAT transcript (AL) is expressed in the opposite direction to LAT from the 5' end of LAT and LAT promoter sequences. In addition, the upstream of LAT (UOL) transcript is expressed in the LAT direction from sequences in the LAT promoter. Further examination of the first 1.5 kb of LAT coding sequences revealed two small ORFs that are anti-sense with respect to LAT (AL2 and AL3). A transcript spanning AL3 was detected in productively infected cells, mouse neuroblastoma cells stably expressing LAT and trigeminal ganglia (TG) of latently infected mice. Peptide-specific IgG directed against AL3 specifically recognized a protein migrating near 15 kDa in cells stably transfected with LAT, mouse neuroblastoma cells transfected with a plasmid containing the AL3 ORF and TG of latently infected mice. The inability to detect the AL3 protein during productive infection may have been because the 5' terminus of the AL3 transcript was downstream of the first in-frame methionine of the AL3 ORF during productive infection.

Expression Profiling Smackdown: Human Transcriptome Array HTA 2.0 vs. RNA-Seq

PubMed Central

Palermo, Meghann; Driscoll, Heather; Tighe, Scott; Dragon, Julie; Bond, Jeff; Shukla, Arti; Vangala, Mahesh; Vincent, James; Hunter, Tim

2014-01-01

The advent of both microarray and massively parallel sequencing have revolutionized high-throughput analysis of the human transcriptome. Due to limitations in microarray technology, detecting and quantifying coding transcript isoforms, in addition to non-coding transcripts, has been challenging. As a result, RNA-Seq has been the preferred method for characterizing the full human transcriptome, until now. A new high-resolution array from Affymetrix, GeneChip Human Transcriptome Array 2.0 (HTA 2.0), has been designed to interrogate all transcript isoforms in the human transcriptome with >6 million probes targeting coding transcripts, exon-exon splice junctions, and non-coding transcripts. Here we compare expression results from GeneChip HTA 2.0 and RNA-Seq data using identical RNA extractions from three samples each of healthy human mesothelial cells in culture, LP9-C1, and healthy mesothelial cells treated with asbestos, LP9-A1. For GeneChip HTA 2.0 sample preparation, we chose to compare two target preparation methods, NuGEN Ovation Pico WTA V2 with the Encore Biotin Module versus Affymetrix's GeneChip WT PLUS with the WT Terminal Labeling Kit, on identical RNA extractions from both untreated and treated samples. These same RNA extractions were used for the RNA-Seq library preparation. All analyses were performed in Partek Genomics Suite 6.6. Expression profiles for control and asbestos-treated mesothelial cells prepared with NuGEN versus Affymetrix target preparation methods (GeneChip HTA 2.0) are compared to each other as well as to RNA-Seq results.

RNAi screening of subtracted transcriptomes reveals tumor suppression by taurine-activated GABAA receptors involved in volume regulation

PubMed Central

van Nierop, Pim; Vormer, Tinke L.; Foijer, Floris; Verheij, Joanne; Lodder, Johannes C.; Andersen, Jesper B.; Mansvelder, Huibert D.; te Riele, Hein

2018-01-01

To identify coding and non-coding suppressor genes of anchorage-independent proliferation by efficient loss-of-function screening, we have developed a method for enzymatic production of low complexity shRNA libraries from subtracted transcriptomes. We produced and screened two LEGO (Low-complexity by Enrichment for Genes shut Off) shRNA libraries that were enriched for shRNA vectors targeting coding and non-coding polyadenylated transcripts that were reduced in transformed Mouse Embryonic Fibroblasts (MEFs). The LEGO shRNA libraries included ~25 shRNA vectors per transcript which limited off-target artifacts. Our method identified 79 coding and non-coding suppressor transcripts. We found that taurine-responsive GABAA receptor subunits, including GABRA5 and GABRB3, were induced during the arrest of non-transformed anchor-deprived MEFs and prevented anchorless proliferation. We show that taurine activates chloride currents through GABAA receptors on MEFs, causing seclusion of cell volume in large membrane protrusions. Volume seclusion from cells by taurine correlated with reduced proliferation and, conversely, suppression of this pathway allowed anchorage-independent proliferation. In human cholangiocarcinomas, we found that several proteins involved in taurine signaling via GABAA receptors were repressed. Low GABRA5 expression typified hyperproliferative tumors, and loss of taurine signaling correlated with reduced patient survival, suggesting this tumor suppressive mechanism operates in vivo. PMID:29787571

Pervasive transcription: detecting functional RNAs in bacteria.

PubMed

Lybecker, Meghan; Bilusic, Ivana; Raghavan, Rahul

2014-01-01

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

3' rapid amplification of cDNA ends (RACE) walking for rapid structural analysis of large transcripts.

PubMed

Ozawa, Tatsuhiko; Kondo, Masato; Isobe, Masaharu

2004-01-01

The 3' rapid amplification of cDNA ends (3' RACE) is widely used to isolate the cDNA of unknown 3' flanking sequences. However, the conventional 3' RACE often fails to amplify cDNA from a large transcript if there is a long distance between the 5' gene-specific primer and poly(A) stretch, since the conventional 3' RACE utilizes 3' oligo-dT-containing primer complementary to the poly(A) tail of mRNA at the first strand cDNA synthesis. To overcome this problem, we have developed an improved 3' RACE method suitable for the isolation of cDNA derived from very large transcripts. By using the oligonucleotide-containing random 9mer together with the GC-rich sequence for the suppression PCR technology at the first strand of cDNA synthesis, we have been able to amplify the cDNA from a very large transcript, such as the microtubule-actin crosslinking factor 1 (MACF1) gene, which codes a transcript of 20 kb in size. When there is no splicing variant, our highly specific amplification allows us to perform the direct sequencing of 3' RACE products without requiring cloning in bacterial hosts. Thus, this stepwise 3' RACE walking will help rapid characterization of the 3' structure of a gene, even when it encodes a very large transcript.

The Global Regulators GacA and ςS Form Part of a Cascade That Controls Alginate Production in Azotobacter vinelandii

PubMed Central

Castañeda, Miguel; Sánchez, Judith; Moreno, Soledad; Núñez, Cinthia; Espín, Guadalupe

2001-01-01

Transcription of the Azotobacter vinelandii algD gene, which encodes GDP-mannose dehydrogenase (the rate-limiting enzyme of alginate synthesis), starts from three sites: p1, p2, and p3. The sensor kinase GacS, a member of the two-component regulatory system, is required for transcription of algD from its three sites during the stationary phase. Here we show that algD is expressed constitutively throughout the growth cycle from the p2 and p3 sites and that transcription from p1 started at the transition between the exponential growth phase and stationary phase. We constructed A. vinelandii strains that carried mutations in gacA encoding the cognate response regulator of GacS and in rpoS coding for the stationary-phase ςS factor. The gacA mutation impaired alginate production and transcription of algD from its three promoters. Transcription of rpoS was also abolished by the gacA mutation. The rpoS mutation impaired transcription of algD from the p1 promoter and increased it from the p2 ςE promoter. The results of this study provide evidence for the predominant role of GacA in a regulatory cascade controlling alginate production and gene expression during the stationary phase in A. vinelandii. PMID:11698366

RNA editing site recognition in heterologous plant mitochondria.

PubMed

Choury, David; Araya, Alejandro

2006-12-01

RNA editing is a process that modifies the information content of mitochondrial messenger RNAs in flowering plants changing specific cytosine residues into uridine. To gain insight into editing site recognition, we used electroporation to introduce engineered wheat (Triticum aestivum) or potato (Solanum tuberosum) mitochondrial cox2 genes, and an atp9-containing chimeric gene, into non-cognate mitochondria, and observed the efficiency of editing in these contexts. Both wheat and potato mitochondria were able to express "foreign" constructs, and their products were properly spliced. Seventeen and twelve editing sites are present in the coding regions of wheat and potato cox2 transcripts, respectively. Eight are common to both plants, whereas nine are specific to wheat, and four to potato. An analogous situation is found for the atp9 mRNA coding regions from these species. We found that both mitochondria were able to recognize sites that are already present as T at the genomic level, making RNA editing unnecessary for that specific residue in the cognate organelle. Our results demonstrate that non-cognate mitochondria are able to edit residues that are not edited in their own transcripts, and support the hypothesis that the same trans-acting factor may recognize several editing sites.

Knockdown of long non-coding RNA XIST increases blood–tumor barrier permeability and inhibits glioma angiogenesis by targeting miR-137

PubMed Central

Yu, H; Xue, Y; Wang, P; Liu, X; Ma, J; Zheng, J; Li, Z; Li, Z; Cai, H; Liu, Y

2017-01-01

Antiangiogenic therapy plays a significant role in combined glioma treatment. However, poor permeability of the blood–tumor barrier (BTB) limits the transport of chemotherapeutic agents, including antiangiogenic drugs, into tumor tissues. Long non-coding RNAs (lncRNAs) have been implicated in various diseases, especially malignant tumors. The present study found that lncRNA X-inactive-specific transcript (XIST) was upregulated in endothelial cells that were obtained in a BTB model in vitro. XIST knockdown increased BTB permeability and inhibited glioma angiogenesis. The analysis of the mechanism of action revealed that the reduction of XIST inhibited the expression of the transcription factor forkhead box C1 (FOXC1) and zonula occludens 2 (ZO-2) by upregulating miR-137. FOXC1 decreased BTB permeability by increasing the promoter activity and expression of ZO-1 and occludin, and promoted glioma angiogenesis by increasing the promoter activity and expression of chemokine (C–X–C motif) receptor 7b (CXCR7). Overall, the present study demonstrates that XIST plays a pivotal role in BTB permeability and glioma angiogenesis, and the inhibition of XIST may be a potential target for the clinical management of glioma. PMID:28287613

Allelic expression mapping across cellular lineages to establish impact of non-coding SNPs

PubMed Central

Adoue, Veronique; Schiavi, Alicia; Light, Nicholas; Almlöf, Jonas Carlsson; Lundmark, Per; Ge, Bing; Kwan, Tony; Caron, Maxime; Rönnblom, Lars; Wang, Chuan; Chen, Shu-Huang; Goodall, Alison H; Cambien, Francois; Deloukas, Panos; Ouwehand, Willem H; Syvänen, Ann-Christine; Pastinen, Tomi

2014-01-01

Most complex disease-associated genetic variants are located in non-coding regions and are therefore thought to be regulatory in nature. Association mapping of differential allelic expression (AE) is a powerful method to identify SNPs with direct cis-regulatory impact (cis-rSNPs). We used AE mapping to identify cis-rSNPs regulating gene expression in 55 and 63 HapMap lymphoblastoid cell lines from a Caucasian and an African population, respectively, 70 fibroblast cell lines, and 188 purified monocyte samples and found 40–60% of these cis-rSNPs to be shared across cell types. We uncover a new class of cis-rSNPs, which disrupt footprint-derived de novo motifs that are predominantly bound by repressive factors and are implicated in disease susceptibility through overlaps with GWAS SNPs. Finally, we provide the proof-of-principle for a new approach for genome-wide functional validation of transcription factor–SNP interactions. By perturbing NFκB action in lymphoblasts, we identified 489 cis-regulated transcripts with altered AE after NFκB perturbation. Altogether, we perform a comprehensive analysis of cis-variation in four cell populations and provide new tools for the identification of functional variants associated to complex diseases. PMID:25326100

Programmable DNA-binding proteins from Burkholderia provide a fresh perspective on the TALE-like repeat domain

PubMed Central

de Lange, Orlando; Wolf, Christina; Dietze, Jörn; Elsaesser, Janett; Morbitzer, Robert; Lahaye, Thomas

2014-01-01

The tandem repeats of transcription activator like effectors (TALEs) mediate sequence-specific DNA binding using a simple code. Naturally, TALEs are injected by Xanthomonas bacteria into plant cells to manipulate the host transcriptome. In the laboratory TALE DNA binding domains are reprogrammed and used to target a fused functional domain to a genomic locus of choice. Research into the natural diversity of TALE-like proteins may provide resources for the further improvement of current TALE technology. Here we describe TALE-like proteins from the endosymbiotic bacterium Burkholderia rhizoxinica, termed Bat proteins. Bat repeat domains mediate sequence-specific DNA binding with the same code as TALEs, despite less than 40% sequence identity. We show that Bat proteins can be adapted for use as transcription factors and nucleases and that sequence preferences can be reprogrammed. Unlike TALEs, the core repeats of each Bat protein are highly polymorphic. This feature allowed us to explore alternative strategies for the design of custom Bat repeat arrays, providing novel insights into the functional relevance of non-RVD residues. The Bat proteins offer fertile grounds for research into the creation of improved programmable DNA-binding proteins and comparative insights into TALE-like evolution. PMID:24792163

SEASTAR: systematic evaluation of alternative transcription start sites in RNA.

PubMed

Qin, Zhiyi; Stoilov, Peter; Zhang, Xuegong; Xing, Yi

2018-05-04

Alternative first exons diversify the transcriptomes of eukaryotes by producing variants of the 5' Untranslated Regions (5'UTRs) and N-terminal coding sequences. Accurate transcriptome-wide detection of alternative first exons typically requires specialized experimental approaches that are designed to identify the 5' ends of transcripts. We developed a computational pipeline SEASTAR that identifies first exons from RNA-seq data alone then quantifies and compares alternative first exon usage across multiple biological conditions. The exons inferred by SEASTAR coincide with transcription start sites identified directly by CAGE experiments and bear epigenetic hallmarks of active promoters. To determine if differential usage of alternative first exons can yield insights into the mechanism controlling gene expression, we applied SEASTAR to an RNA-seq dataset that tracked the reprogramming of mouse fibroblasts into induced pluripotent stem cells. We observed dynamic temporal changes in the usage of alternative first exons, along with correlated changes in transcription factor expression. Using a combined sequence motif and gene set enrichment analysis we identified N-Myc as a regulator of alternative first exon usage in the pluripotent state. Our results demonstrate that SEASTAR can leverage the available RNA-seq data to gain insights into the control of gene expression and alternative transcript variation in eukaryotic transcriptomes.

Mediator MED23 regulates basal transcription in vivo via an interaction with P-TEFb.

PubMed

Wang, Wei; Yao, Xiao; Huang, Yan; Hu, Xiangming; Liu, Runzhong; Hou, Dongming; Chen, Ruichuan; Wang, Gang

2013-01-01

The Mediator is a multi-subunit complex that transduces regulatory information from transcription regulators to the RNA polymerase II apparatus. Growing evidence suggests that Mediator plays roles in multiple stages of eukaryotic transcription, including elongation. However, the detailed mechanism by which Mediator regulates elongation remains elusive. In this study, we demonstrate that Mediator MED23 subunit controls a basal level of transcription by recruiting elongation factor P-TEFb, via an interaction with its CDK9 subunit. The mRNA level of Egr1, a MED23-controlled model gene, is reduced 4-5 fold in Med23 (-/-) ES cells under an unstimulated condition, but Med23-deficiency does not alter the occupancies of RNAP II, GTFs, Mediator complex, or activator ELK1 at the Egr1 promoter. Instead, Med23 depletion results in a significant decrease in P-TEFb and RNAP II (Ser2P) binding at the coding region, but no changes for several other elongation regulators, such as DSIF and NELF. ChIP-seq revealed that Med23-deficiency partially reduced the P-TEFb occupancy at a set of MED23-regulated gene promoters. Further, we demonstrate that MED23 interacts with CDK9 in vivo and in vitro. Collectively, these results provide the mechanistic insight into how Mediator promotes RNAP II into transcription elongation.

Transcript profiling of genes expressed during fibre development in diploid cotton (Gossypium arboreum L.).

PubMed

Hande, Atul S; Katageri, Ishwarappa S; Jadhav, Mangesh P; Adiger, Sateesh; Gamanagatti, Savita; Padmalatha, Kethireddy Venkata; Dhandapani, Gurusamy; Kanakachari, Mogilicherla; Kumar, Polumetla Ananda; Reddy, Vanga Siva

2017-08-31

Cotton fibre is a single cell and it is one of the best platforms for unraveling the genes express during various stages of fibre development. There are reports devoted to comparative transcriptome study on fiber cell initiation and elongation in tetraploid cultivated cotton. However, in the present investigation, comparative transcriptome study was made in diploid cultivated cotton using isogenic fuzzy-lintless (Fl) and normal fuzzy linted (FL) lines belong to Gossypium arboreum, diploid species at two stages, 0 and 10 dpa (days post anthesis), using Affymetrix cotton GeneChip genome array. Scanning electron microscopy (SEM) analysis uncovered the occurrence of few fibre cell initials in the Fl line as compared to many in Normal FL at -2 and 0 dpa. However, at 10 dpa there were no fibre cells found elongated in Fl but many elongated cells were found in FL line. Up-regulation of transcription factors, AP2-EREBP, C2H2, C3H, HB and WRKY was observed at 0 dpa whereas in 10 dpa transcription factors, AP2-EREBP, AUX/IAA, bHLH, C2H2, C3H, HB, MYB, NAC, Orphans, PLATZ and WRKY were found down regulated in Fl line. These transcription factors were mainly involved in metabolic pathways such as phytohormone signaling, energy metabolism of cell, fatty acid metabolism, secondary metabolism and other signaling pathways and are related directly or indirectly in fiber development. Quantitative real-time PCR was performed to check fold up or down-regulation of these genes and transcription factors (TFs) down regulated in mutants as compared to normal at 0 and 10 dpa. This study elucidates that the up-regulation of transcription factors like AP2-EREBP, C2H2, C3H, HB, WRKY and phytohormone signaling genes at 0 dpa and their down-regulation at the 10 dpa might have constrain the fibre elongation in fuzzy-lintless line. Along with this the down-regulation of genes involved in synthesis of VLCFA chain, transcripts necessary for energy and cell wall metabolism, EXPANSINs, arabinogalactan proteins (AGPs), tubulin might also be the probable reason for reduced growth of fibres in the Fl. Plant receptor-like kinases (RLKs), Leucine Rich Repeats) LRR- family protein and signal transduction coding for mitogen-activated protein kinase (MAPK) cascade, have been engaged in coordination of cell elongation and SCW biosynthesis, down-regulation of these might loss the function leads to reduced fibre growth.

A Partial Least Squares Based Procedure for Upstream Sequence Classification in Prokaryotes.

PubMed

Mehmood, Tahir; Bohlin, Jon; Snipen, Lars

2015-01-01

The upstream region of coding genes is important for several reasons, for instance locating transcription factor, binding sites, and start site initiation in genomic DNA. Motivated by a recently conducted study, where multivariate approach was successfully applied to coding sequence modeling, we have introduced a partial least squares (PLS) based procedure for the classification of true upstream prokaryotic sequence from background upstream sequence. The upstream sequences of conserved coding genes over genomes were considered in analysis, where conserved coding genes were found by using pan-genomics concept for each considered prokaryotic species. PLS uses position specific scoring matrix (PSSM) to study the characteristics of upstream region. Results obtained by PLS based method were compared with Gini importance of random forest (RF) and support vector machine (SVM), which is much used method for sequence classification. The upstream sequence classification performance was evaluated by using cross validation, and suggested approach identifies prokaryotic upstream region significantly better to RF (p-value < 0.01) and SVM (p-value < 0.01). Further, the proposed method also produced results that concurred with known biological characteristics of the upstream region.

Diversity, expansion, and evolutionary novelty of plant DNA-binding transcription factor families.

PubMed

Lehti-Shiu, Melissa D; Panchy, Nicholas; Wang, Peipei; Uygun, Sahra; Shiu, Shin-Han

2017-01-01

Plant transcription factors (TFs) that interact with specific sequences via DNA-binding domains are crucial for regulating transcriptional initiation and are fundamental to plant development and environmental response. In addition, expansion of TF families has allowed functional divergence of duplicate copies, which has contributed to novel, and in some cases adaptive, traits in plants. Thus, TFs are central to the generation of the diverse plant species that we see today. Major plant agronomic traits, including those relevant to domestication, have also frequently arisen through changes in TF coding sequence or expression patterns. Here our goal is to provide an overview of plant TF evolution by first comparing the diversity of DNA-binding domains and the sizes of these domain families in plants and other eukaryotes. Because TFs are among the most highly expanded gene families in plants, the birth and death process of TFs as well as the mechanisms contributing to their retention are discussed. We also provide recent examples of how TFs have contributed to novel traits that are important in plant evolution and in agriculture.This article is part of a Special Issue entitled: Plant Gene Regulatory Mechanisms and Networks, edited by Dr. Erich Grotewold and Dr. Nathan Springer. Copyright © 2016 Elsevier B.V. All rights reserved.

Sequences analyses and expression profiles in tissues and embryos of Japanese flounder (Paralichthys olivaceus) PRDM1.

PubMed

Liu, Conghui; Liu, Wei; Fan, Lin; Liu, Jinxiang; Li, Peizhen; Zhang, Wei; Gao, Jinning; Li, Zan; Zhang, Quanqi; Wang, Xubo

2016-04-01

PRDM1 (PRDI-BF1-RIZ1 homologous domain containing 1) appears to be a pleiotropic regulatory factor in various processes. It contains a PR (PRDI-BF1-RIZ1 homologous) domain protein and five zinc fingers. In the present study, a gene coding the homolog of prdm1 and the 5' regulatory region of prdm1 was identified from the Paralichthys olivaceus (denoted Po-prdm1). Results of real-time quantitative polymerase chain reaction amplification (RT-qPCR) and in situ hybridization (ISH) in embryos revealed that Po-prdm1 was highly expressed between the early gastrula and tail bud stages, with its expression peaking in the mid-gastrula stage, whereas the results of RT-qPCR and ISH in tissues demonstrated that Po-prdm1 transcripts were ubiquitously detected in all tissues, which indicates its pleiotropic function in multiple processes. ISH of gonadal tissues revealed that the transcripts were located in the nucleus and cytoplasm of the oocytes in the ovaries but only in the spermatogonia and not in the spermatocytes in the testes. The Po-prdm1 transcription factor binding sites and their conserved binding region among vertebrates were analyzed in this study. The combined results suggest that Po-PRDM1 has a conserved function in teleosts and mammals.

Expression of transcription factors after short-term exposure of Arabidopsis thaliana cell cultures to hyper-g, and to simulated and sounding rocket micro-g

NASA Astrophysics Data System (ADS)

Hampp, R.; Babbick, M.

Previous microarray studies with cell cultures of Arabidopsis thaliana cv Columbia have shown responses in gene expression which were partly specific to exposure to microgravity sounding rocket experiment TEXUS In order to get access to early responses upon changes in gravitational fields we used exposure times as short as 2 min For this purpose we selected a range of genes which code for different groups of transcription factors WRKY ERF MYB MADS Samples were taken in 5-min clinorotation 2- and 3-dimensional hypergravity 8g and 2-min intervals sounding rocket experiment Amounts of transcripts were determined by quantitative RT PCR Most transcripts showed a significant transient change in content within a time frame of up to 30 min after changing the external gravitational field strength They could be grouped into 1 basic stress responses which occurred under all conditions 2 clinorotation-related effects which were either identical or opposite between 2D 60 rpm 4x10 -2 g and 3D clinorotation random positioning machine and 3 alterations specific to the microgravity exposure under sounding rocket conditions MAXUS The data are discussed in relation to gravitation-dependent signalling chains and with regard to the simulation of microgravity by means of clinorotation Supported by a grant from the Deutsches Zentrum f u r Luft- und Raumfahrt e V grant no 50 WB 0143

Gene expression analysis of WRKY transcription factors in Arabidopsis thaliana cell cultures during a parabolic flight

NASA Astrophysics Data System (ADS)

Babbick, Maren; Barjaktarović, Žarko; Hampp, Ruediger

Plants sense gravity by specialized cells (statocytes) and adjust growth and development accordingly. It has, however, also been shown that plant cells which are not part of specialized tissues are also able to sense gravitational forces. Therefore we used undifferentiated, homogeneous cell cultures of Arabidopsis thaliana (cv. Columbia) in order to identify early alterations in gene expression as a response to altered gravitational field strengths. In this contribution we report on cell cultures exposed to parabolic flights (approximately 20 sec of microgravity). For this short-term exposure study, we specifically checked for genes at the beginning of signal transduction chains, such as those coding for transcription factors (TFs). TFs are small proteins that regulate expression of their target genes by binding to specific promoter sequences. Our main focus were members of the so-called WRKY TF family. WRKY TFs are known to be involved in various physiological processes like senescence and pathogen defense. By quantifying transcriptional changes of these genes by real-time RT-PCR, we wanted to find out, how gene expression is affected by both hyperand microgravity conditions during a parabolic flight. For this purpose Arabidopsis thaliana callus cultures were metabolically quenched by the injection of RNAlater at the end of the microgravity-phase of each parabola. The data we present will show how fast changes in amounts of transcripts will occur, and to what degree the expression profiles are comparable with data obtained from exposures to hypergravity and simulated microgravity.

Regulation of host-pathogen interactions via the post-transcriptional Csr/Rsm system.

PubMed

Kusmierek, Maria; Dersch, Petra

2018-02-01

A successful colonization of specific hosts requires a rapid and efficient adaptation of the virulence-relevant gene expression program by bacterial pathogens. An important element in this endeavor is the Csr/Rsm system. This multi-component, post-transcriptional control system forms a central hub within complex regulatory networks and coordinately adjusts virulence properties with metabolic and physiological attributes of the pathogen. A key function is elicited by the RNA-binding protein CsrA/RsmA. CsrA/RsmA interacts with numerous target mRNAs, many of which encode crucial virulence factors, and alters their translation, stability or elongation of transcription. Recent studies highlighted that important colonization factors, toxins, and bacterial secretion systems are under CsrA/RsmA control. CsrA/RsmA deficiency impairs host colonization and attenuates virulence, making this post-transcriptional regulator a suitable drug target. The CsrA/RsmA protein can be inactivated through sequestration by non-coding RNAs, or via binding to specific highly abundant mRNAs and interacting proteins. The wide range of interaction partners and RNA targets, as well as the overarching, interlinked genetic control circuits illustrate the complexity of this regulatory system in the different pathogens. Future work addressing spatio-temporal changes of Csr/Rsm-mediated control during the course of an infection will help us to understand how bacteria reprogram their expression profile to cope with continuous changes experienced in colonized niches. Copyright © 2017 Elsevier Ltd. All rights reserved.

Inferring genome-wide functional modulatory network: a case study on NF-κB/RelA transcription factor.

PubMed

Li, Xueling; Zhu, Min; Brasier, Allan R; Kudlicki, Andrzej S

2015-04-01

How different pathways lead to the activation of a specific transcription factor (TF) with specific effects is not fully understood. We model context-specific transcriptional regulation as a modulatory network: triplets composed of a TF, target gene, and modulator. Modulators usually affect the activity of a specific TF at the posttranscriptional level in a target gene-specific action mode. This action may be classified as enhancement, attenuation, or inversion of either activation or inhibition. As a case study, we inferred, from a large collection of expression profiles, all potential modulations of NF-κB/RelA. The predicted modulators include many proteins previously not reported as physically binding to RelA but with relevant functions, such as RNA processing, cell cycle, mitochondrion, ubiquitin-dependent proteolysis, and chromatin modification. Modulators from different processes exert specific prevalent action modes on distinct pathways. Modulators from noncoding RNA, RNA-binding proteins, TFs, and kinases modulate the NF-κB/RelA activity with specific action modes consistent with their molecular functions and modulation level. The modulatory networks of NF-κB/RelA in the context epithelial-mesenchymal transition (EMT) and burn injury have different modulators, including those involved in extracellular matrix (FBN1), cytoskeletal regulation (ACTN1), and metastasis-associated lung adenocarcinoma transcript 1 (MALAT1), a long intergenic nonprotein coding RNA, and tumor suppression (FOXP1) for EMT, and TXNIP, GAPDH, PKM2, IFIT5, LDHA, NID1, and TPP1 for burn injury.

BMP15 c.-9C>G promoter sequence variant may contribute to the cause of non-syndromic premature ovarian failure.

PubMed

Fonseca, Dora Janeth; Ortega-Recalde, Oscar; Esteban-Perez, Clara; Moreno-Ortiz, Harold; Patiño, Liliana Catherine; Bermúdez, Olga María; Ortiz, Angela María; Restrepo, Carlos M; Lucena, Elkin; Laissue, Paul

2014-11-01

BMP15 has drawn particular attention in the pathophysiology of reproduction, as its mutations in mammalian species have been related to different reproductive phenotypes. In humans, BMP15 coding regions have been sequenced in large panels of women with premature ovarian failure (POF), but only some mutations have been definitely validated as causing the phenotype. A functional association between the BMP15 c.-9C>G promoter polymorphism and cause of POF have been reported. The aim of this study was to determine the potential functional effect of this sequence variant on specific BMP15 promoter transactivation disturbances. Bioinformatics was used to identify transcription factor binding sites located on the promoter region of BMP15. Reverse transcription polymerase chain reaction was used to study specific gene expression in ovarian tissue. Luciferase reporter assays were used to establish transactivation disturbances caused by the BMP15 c.-9C>G variant. The c.-9C>G variant was found to modify the PITX1 transcription factor binding site. PITX1 and BMP15 co-expressed in human and mouse ovarian tissue, and PITX1 transactivated both BMP15 promoter versions (-9C and -9G). It was found that the BMP15 c.-9G allele was related to BMP15 increased transcription, supporting c.-9C>G as a causal agent of POF. Copyright © 2014 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

Intergenic disease-associated regions are abundant in novel transcripts.

PubMed

Bartonicek, N; Clark, M B; Quek, X C; Torpy, J R; Pritchard, A L; Maag, J L V; Gloss, B S; Crawford, J; Taft, R J; Hayward, N K; Montgomery, G W; Mattick, J S; Mercer, T R; Dinger, M E

2017-12-28

Genotyping of large populations through genome-wide association studies (GWAS) has successfully identified many genomic variants associated with traits or disease risk. Unexpectedly, a large proportion of GWAS single nucleotide polymorphisms (SNPs) and associated haplotype blocks are in intronic and intergenic regions, hindering their functional evaluation. While some of these risk-susceptibility regions encompass cis-regulatory sites, their transcriptional potential has never been systematically explored. To detect rare tissue-specific expression, we employed the transcript-enrichment method CaptureSeq on 21 human tissues to identify 1775 multi-exonic transcripts from 561 intronic and intergenic haploblocks associated with 392 traits and diseases, covering 73.9 Mb (2.2%) of the human genome. We show that a large proportion (85%) of disease-associated haploblocks express novel multi-exonic non-coding transcripts that are tissue-specific and enriched for GWAS SNPs as well as epigenetic markers of active transcription and enhancer activity. Similarly, we captured transcriptomes from 13 melanomas, targeting nine melanoma-associated haploblocks, and characterized 31 novel melanoma-specific transcripts that include fusion proteins, novel exons and non-coding RNAs, one-third of which showed allelically imbalanced expression. This resource of previously unreported transcripts in disease-associated regions ( http://gwas-captureseq.dingerlab.org ) should provide an important starting point for the translational community in search of novel biomarkers, disease mechanisms, and drug targets.

Evolution of transcriptional enhancers and animal diversity

PubMed Central

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

SAGE analysis of early oogenesis in the silkworm, Bombyx mori.

PubMed

Funaguma, Shunsuke; Hashimoto, Shin-ichi; Suzuki, Yutaka; Omuro, Naoko; Sugano, Sumio; Mita, Kazuei; Katsuma, Susumu; Shimada, Toru

2007-02-01

To identify genes involved in the differentiation of Bombyx cystoblast, we constructed two 3' long serial analysis of gene expression (Long SAGE) libraries from stage 1-3 or stage 2-3 egg chambers and compared their gene expression profiles. In both libraries, the most frequent tags were derived from the same novel transcript. The transcript does not have any open reading frame capable of encoding a protein with over 100 amino acids in length. RNA blot analysis revealed that this transcript is specifically and abundantly expressed in the Bombyx ovary, mainly the germ line cells in the ovarioles. These results suggest that Bombyx oogenesis may be regulated by a previously unidentified non-coding RNA. Comparison of the gene expression profiles between the stage 1-3 and stage 2-3 egg chamber libraries revealed that 272 tags were significantly more abundant in stage 1-3 egg chambers (p<0.05 and at least two-fold change) than in library 2. Among the differentially expressed transcripts were the sequences that correspond to ATP synthase subunit d (3.1-fold enriched) and ATP synthase coupling factor 6 (9.1-fold enriched), suggesting that they are involved in regulation of cell cycle of cystocytes.

Improvement of acetic acid tolerance of Saccharomyces cerevisiae using a zinc-finger-based artificial transcription factor and identification of novel genes involved in acetic acid tolerance.

PubMed

Ma, Cui; Wei, Xiaowen; Sun, Cuihuan; Zhang, Fei; Xu, Jianren; Zhao, Xinqing; Bai, Fengwu

2015-03-01

Acetic acid is present in cellulosic hydrolysate as a potent inhibitor, and the superior acetic acid tolerance of Saccharomyces cerevisiae ensures good cell viability and efficient ethanol production when cellulosic raw materials are used as substrates. In this study, a mutant strain of S. cerevisiae ATCC4126 (Sc4126-M01) with improved acetic acid tolerance was obtained through screening strains transformed with an artificial zinc finger protein transcription factor (ZFP-TF) library. Further analysis indicated that improved acetic acid tolerance was associated with improved catalase (CAT) activity. The ZFP coding sequence associated with the improved phenotype was identified, and real-time RT-PCR analysis revealed that three of the possible genes involved in the enhanced acetic acid tolerance regulated by this ZFP-TF, namely YFL040W, QDR3, and IKS1, showed decreased transcription levels in Sc4126-M01 in the presence of acetic acid, compared to those in the control strain. Sc4126-M01 mutants having QDR3 and IKS1 deletion (ΔQDR3 and ΔIKS1) exhibited higher acetic acid tolerance than the wild-type strain under acetic acid treatment. Glucose consumption rate and ethanol productivity in the presence of 5 g/L acetic acid were improved in the ΔQDR3 mutant compared to the wild-type strain. Our studies demonstrated that the synthetic ZFP-TF library can be used to improve acetic acid tolerance of S. cerevisiae and that the employment of an artificial transcription factor can facilitate the exploration of novel functional genes involved in stress tolerance of S. cerevisiae.

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

NASA Astrophysics Data System (ADS)

Tarpine, Ryan; Istrail, Sorin

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

Epigenetic priors for identifying active transcription factor binding sites.

PubMed

Cuellar-Partida, Gabriel; Buske, Fabian A; McLeay, Robert C; Whitington, Tom; Noble, William Stafford; Bailey, Timothy L

2012-01-01

Accurate knowledge of the genome-wide binding of transcription factors in a particular cell type or under a particular condition is necessary for understanding transcriptional regulation. Using epigenetic data such as histone modification and DNase I, accessibility data has been shown to improve motif-based in silico methods for predicting such binding, but this approach has not yet been fully explored. We describe a probabilistic method for combining one or more tracks of epigenetic data with a standard DNA sequence motif model to improve our ability to identify active transcription factor binding sites (TFBSs). We convert each data type into a position-specific probabilistic prior and combine these priors with a traditional probabilistic motif model to compute a log-posterior odds score. Our experiments, using histone modifications H3K4me1, H3K4me3, H3K9ac and H3K27ac, as well as DNase I sensitivity, show conclusively that the log-posterior odds score consistently outperforms a simple binary filter based on the same data. We also show that our approach performs competitively with a more complex method, CENTIPEDE, and suggest that the relative simplicity of the log-posterior odds scoring method makes it an appealing and very general method for identifying functional TFBSs on the basis of DNA and epigenetic evidence. FIMO, part of the MEME Suite software toolkit, now supports log-posterior odds scoring using position-specific priors for motif search. A web server and source code are available at http://meme.nbcr.net. Utilities for creating priors are at http://research.imb.uq.edu.au/t.bailey/SD/Cuellar2011. t.bailey@uq.edu.au Supplementary data are available at Bioinformatics online.

Genome-wide characterization and expression analysis of citrus NUCLEAR FACTOR-Y (NF-Y) transcription factors identified a novel NF-YA gene involved in drought-stress response and tolerance.

PubMed

Pereira, Suzam L S; Martins, Cristina P S; Sousa, Aurizangela O; Camillo, Luciana R; Araújo, Caroline P; Alcantara, Grazielle M; Camargo, Danielle S; Cidade, Luciana C; de Almeida, Alex-Alan F; Costa, Marcio G C

2018-01-01

Nuclear factor Y (NF-Y) is a ubiquitous transcription factor found in eukaryotes. It is composed of three distinct subunits called NF-YA, NF-YB and NF-YC. NF-Ys have been identified as key regulators of multiple pathways in the control of development and tolerance to biotic and abiotic factors. The present study aimed to identify and characterize the complete repertoire of genes coding for NF-Y in citrus, as well as to perform the functional characterization of one of its members, namely CsNFYA5, in transgenic tobacco plants. A total of 22 genes coding for NF-Y were identified in the genomes of sweet orange (Citrus sinensis) and Clementine mandarin (C. clementina), including six CsNF-YAs, 11 CsNF-YBs and five CsNF-YCs. Phylogenetic analyses showed that there is a NF-Y orthologous in the Clementine genome for each sweet orange NF-Y gene; this was not observed when compared to Arabidopsis thaliana. CsNF-Y proteins shared the same conserved domains with their orthologous proteins in other organisms, including mouse. Analysis of gene expression by RNA-seq and EST data demonstrated that CsNF-Ys have a tissue-specific and stress inducible expression profile. qRT-PCR analysis revealed that CsNF-YA5 exhibits differential expression in response to water deficit in leaves and roots of citrus plants. Overexpression of CsNF-YA5 in transgenic tobacco plants contributed to the reduction of H2O2 production under dehydration conditions and increased plant growth and photosynthetic rate under normal conditions and drought stress. These biochemical and physiological responses to drought stress promoted by CsNF-YA5 may confer a productivity advantage in environments with frequent short-term soil water deficit.

Coral-zooxanthellae meta-transcriptomics reveals integrated response to pollutant stress.

PubMed

Gust, Kurt A; Najar, Fares Z; Habib, Tanwir; Lotufo, Guilherme R; Piggot, Alan M; Fouke, Bruce W; Laird, Jennifer G; Wilbanks, Mitchell S; Rawat, Arun; Indest, Karl J; Roe, Bruce A; Perkins, Edward J

2014-07-12

Corals represent symbiotic meta-organisms that require harmonization among the coral animal, photosynthetic zooxanthellae and associated microbes to survive environmental stresses. We investigated integrated-responses among coral and zooxanthellae in the scleractinian coral Acropora formosa in response to an emerging marine pollutant, the munitions constituent, 1,3,5-trinitro-1,3,5 triazine (RDX; 5 day exposures to 0 (control), 0.5, 0.9, 1.8, 3.7, and 7.2 mg/L, measured in seawater). RDX accumulated readily in coral soft tissues with bioconcentration factors ranging from 1.1 to 1.5. Next-generation sequencing of a normalized meta-transcriptomic library developed for the eukaryotic components of the A. formosa coral holobiont was leveraged to conduct microarray-based global transcript expression analysis of integrated coral/zooxanthellae responses to the RDX exposure. Total differentially expressed transcripts (DET) increased with increasing RDX exposure concentrations as did the proportion of zooxanthellae DET relative to the coral animal. Transcriptional responses in the coral demonstrated higher sensitivity to RDX compared to zooxanthellae where increased expression of gene transcripts coding xenobiotic detoxification mechanisms (i.e. cytochrome P450 and UDP glucuronosyltransferase 2 family) were initiated at the lowest exposure concentration. Increased expression of these detoxification mechanisms was sustained at higher RDX concentrations as well as production of a physical barrier to exposure through a 40% increase in mucocyte density at the maximum RDX exposure. At and above the 1.8 mg/L exposure concentration, DET coding for genes involved in central energy metabolism, including photosynthesis, glycolysis and electron-transport functions, were decreased in zooxanthellae although preliminary data indicated that zooxanthellae densities were not affected. In contrast, significantly increased transcript expression for genes involved in cellular energy production including glycolysis and electron-transport pathways was observed in the coral animal. Transcriptional network analysis for central energy metabolism demonstrated highly correlated responses to RDX among the coral animal and zooxanthellae indicative of potential compensatory responses to lost photosynthetic potential within the holobiont. These observations underscore the potential for complex integrated responses to RDX exposure among species comprising the coral holobiont and highlight the need to understand holobiont-species interactions to accurately assess pollutant impacts.

Long non-coding RNA HOTAIR, a c-Myc activated driver of malignancy, negatively regulates miRNA-130a in gallbladder cancer

PubMed Central

2014-01-01

Background Protein coding genes account for only about 2% of the human genome, whereas the vast majority of transcripts are non-coding RNAs including long non-coding RNAs. A growing volume of literature has proposed that lncRNAs are important players in cancer. HOTAIR was previously shown to be an oncogene and negative prognostic factor in a variety of cancers. However, the factors that contribute to its upregulation and the interaction between HOTAIR and miRNAs are largely unknown. Methods A computational screen of HOTAIR promoter was conducted to search for transcription-factor-binding sites. HOTAIR promoter activities were examined by luciferase reporter assay. The function of the c-Myc binding site in the HOTAIR promoter region was tested by a promoter assay with nucleotide substitutions in the putative E-box. The association of c-Myc with the HOTAIR promoter in vivo was confirmed by chromatin immunoprecipitation assay and Electrophoretic mobility shift assay. A search for miRNAs with complementary base paring with HOTAIR was performed utilizing online software program. Gain and loss of function approaches were employed to investigate the expression changes of HOTAIR or miRNA-130a. The expression levels of HOTAIR, c-Myc and miRNA-130a were examined in 65 matched pairs of gallbladder cancer tissues. The effects of HOTAIR and miRNA-130a on gallbladder cancer cell invasion and proliferation was tested using in vitro cell invasion and flow cytometric assays. Results We demonstrate that HOTAIR is a direct target of c-Myc through interaction with putative c-Myc target response element (RE) in the upstream region of HOTAIR in gallbladder cancer cells. A positive correlation between c-Myc and HOTAIR mRNA levels was observed in gallbladder cancer tissues. We predicted that HOTAIR harbors a miRNA-130a binding site. Our data showed that this binding site is vital for the regulation of miRNA-130a by HOTAIR. Moreover, a negative correlation between HOTAIR and miRNA-130a was observed in gallbladder cancer tissues. Finally, we demonstrate that the oncogenic activity of HOTAIR is in part through its negative regulation of miRNA-130a. Conclusion Together, these results suggest that HOTAIR is a c-Myc-activated driver of malignancy, which acts in part through repression of miRNA-130a. PMID:24953832

Barriers and facilitators to the implementation of a school-based physical activity policy in Canada: application of the theoretical domains framework.

PubMed

Weatherson, Katie A; McKay, Rhyann; Gainforth, Heather L; Jung, Mary E

2017-10-23

In British Columbia Canada, a Daily Physical Activity (DPA) policy was mandated that requires elementary school teachers to provide students with opportunities to achieve 30 min of physical activity during the school day. However, the implementation of school-based physical activity policies is influenced by many factors. A theoretical examination of the factors that impede and enhance teachers' implementation of physical activity policies is necessary in order to develop strategies to improve policy practice and achieve desired outcomes. This study used the Theoretical Domains Framework (TDF) to understand teachers' barriers and facilitators to the implementation of the DPA policy in one school district. Additionally, barriers and facilitators were examined and compared according to how the teacher implemented the DPA policy during the instructional school day. Interviews were conducted with thirteen teachers and transcribed verbatim. One researcher performed barrier and facilitator extraction, with double extraction occurring across a third of the interview transcripts by a second researcher. A deductive and inductive analytical approach in a two-stage process was employed whereby barriers and facilitators were deductively coded using TDF domains (content analysis) and analyzed for sub-themes within each domain. Two researchers performed coding. A total of 832 items were extracted from the interview transcripts. Some items were coded into multiple TDF domains, resulting in a total of 1422 observations. The most commonly coded TDF domains accounting for 75% of the total were Environmental context and resources (ECR; n = 250), Beliefs about consequences (n = 225), Social influences (n = 193), Knowledge (n = 100), and Intentions (n = 88). Teachers who implemented DPA during instructional time differed from those who relied on non-instructional time in relation to Goals, Behavioural regulation, Social/professional role and identity, Beliefs about Consequences. Forty-one qualitative sub-themes were identified across the fourteen domains and exemplary quotes were highlighted. Teachers identified barriers and facilitators relating to all TDF domains, with ECR, Beliefs about consequences, Social influences, Knowledge and Intentions being the most often discussed influencers of DPA policy implementation. Use of the TDF to understand the implementation factors can assist with the systematic development of future interventions to improve implementation.

Identification of Circular RNAs from the Parental Genes Involved in Multiple Aspects of Cellular Metabolism in Barley

PubMed Central

Darbani, Behrooz; Noeparvar, Shahin; Borg, Søren

2016-01-01

RNA circularization made by head-to-tail back-splicing events is involved in the regulation of gene expression from transcriptional to post-translational levels. By exploiting RNA-Seq data and down-stream analysis, we shed light on the importance of circular RNAs in plants. The results introduce circular RNAs as novel interactors in the regulation of gene expression in plants and imply the comprehensiveness of this regulatory pathway by identifying circular RNAs for a diverse set of genes. These genes are involved in several aspects of cellular metabolism as hormonal signaling, intracellular protein sorting, carbohydrate metabolism and cell-wall biogenesis, respiration, amino acid biosynthesis, transcription and translation, and protein ubiquitination. Additionally, these parental loci of circular RNAs, from both nuclear and mitochondrial genomes, encode for different transcript classes including protein coding transcripts, microRNA, rRNA, and long non-coding/microprotein coding RNAs. The results shed light on the mitochondrial exonic circular RNAs and imply the importance of circular RNAs for regulation of mitochondrial genes. Importantly, we introduce circular RNAs in barley and elucidate their cellular-level alterations across tissues and in response to micronutrients iron and zinc. In further support of circular RNAs' functional roles in plants, we report several cases where fluctuations of circRNAs do not correlate with the levels of their parental-loci encoded linear transcripts. PMID:27375638

Keeping abreast with long non-coding RNAs in mammary gland development and breast cancer

PubMed Central

Hansji, Herah; Leung, Euphemia Y.; Baguley, Bruce C.; Finlay, Graeme J.; Askarian-Amiri, Marjan E.

2014-01-01

The majority of the human genome is transcribed, even though only 2% of transcripts encode proteins. Non-coding transcripts were originally dismissed as evolutionary junk or transcriptional noise, but with the development of whole genome technologies, these non-coding RNAs (ncRNAs) are emerging as molecules with vital roles in regulating gene expression. While shorter ncRNAs have been extensively studied, the functional roles of long ncRNAs (lncRNAs) are still being elucidated. Studies over the last decade show that lncRNAs are emerging as new players in a number of diseases including cancer. Potential roles in both oncogenic and tumor suppressive pathways in cancer have been elucidated, but the biological functions of the majority of lncRNAs remain to be identified. Accumulated data are identifying the molecular mechanisms by which lncRNA mediates both structural and functional roles. LncRNA can regulate gene expression at both transcriptional and post-transcriptional levels, including splicing and regulating mRNA processing, transport, and translation. Much current research is aimed at elucidating the function of lncRNAs in breast cancer and mammary gland development, and at identifying the cellular processes influenced by lncRNAs. In this paper we review current knowledge of lncRNAs contributing to these processes and present lncRNA as a new paradigm in breast cancer development. PMID:25400658

Transcripts with in silico predicted RNA structure are enriched everywhere in the mouse brain

PubMed Central

2012-01-01

Background Post-transcriptional control of gene expression is mostly conducted by specific elements in untranslated regions (UTRs) of mRNAs, in collaboration with specific binding proteins and RNAs. In several well characterized cases, these RNA elements are known to form stable secondary structures. RNA secondary structures also may have major functional implications for long noncoding RNAs (lncRNAs). Recent transcriptional data has indicated the importance of lncRNAs in brain development and function. However, no methodical efforts to investigate this have been undertaken. Here, we aim to systematically analyze the potential for RNA structure in brain-expressed transcripts. Results By comprehensive spatial expression analysis of the adult mouse in situ hybridization data of the Allen Mouse Brain Atlas, we show that transcripts (coding as well as non-coding) associated with in silico predicted structured probes are highly and significantly enriched in almost all analyzed brain regions. Functional implications of these RNA structures and their role in the brain are discussed in detail along with specific examples. We observe that mRNAs with a structure prediction in their UTRs are enriched for binding, transport and localization gene ontology categories. In addition, after manual examination we observe agreement between RNA binding protein interaction sites near the 3’ UTR structures and correlated expression patterns. Conclusions Our results show a potential use for RNA structures in expressed coding as well as noncoding transcripts in the adult mouse brain, and describe the role of structured RNAs in the context of intracellular signaling pathways and regulatory networks. Based on this data we hypothesize that RNA structure is widely involved in transcriptional and translational regulatory mechanisms in the brain and ultimately plays a role in brain function. PMID:22651826

Deep Sequencing Reveals Uncharted Isoform Heterogeneity of the Protein-Coding Transcriptome in Cerebral Ischemia.

PubMed

Bhattarai, Sunil; Aly, Ahmed; Garcia, Kristy; Ruiz, Diandra; Pontarelli, Fabrizio; Dharap, Ashutosh

2018-06-03

Gene expression in cerebral ischemia has been a subject of intense investigations for several years. Studies utilizing probe-based high-throughput methodologies such as microarrays have contributed significantly to our existing knowledge but lacked the capacity to dissect the transcriptome in detail. Genome-wide RNA-sequencing (RNA-seq) enables comprehensive examinations of transcriptomes for attributes such as strandedness, alternative splicing, alternative transcription start/stop sites, and sequence composition, thus providing a very detailed account of gene expression. Leveraging this capability, we conducted an in-depth, genome-wide evaluation of the protein-coding transcriptome of the adult mouse cortex after transient focal ischemia at 6, 12, or 24 h of reperfusion using RNA-seq. We identified a total of 1007 transcripts at 6 h, 1878 transcripts at 12 h, and 1618 transcripts at 24 h of reperfusion that were significantly altered as compared to sham controls. With isoform-level resolution, we identified 23 splice variants arising from 23 genes that were novel mRNA isoforms. For a subset of genes, we detected reperfusion time-point-dependent splice isoform switching, indicating an expression and/or functional switch for these genes. Finally, for 286 genes across all three reperfusion time-points, we discovered multiple, distinct, simultaneously expressed and differentially altered isoforms per gene that were generated via alternative transcription start/stop sites. Of these, 165 isoforms derived from 109 genes were novel mRNAs. Together, our data unravel the protein-coding transcriptome of the cerebral cortex at an unprecedented depth to provide several new insights into the flexibility and complexity of stroke-related gene transcription and transcript organization.

A lncRNA Perspective into (Re)Building the Heart.

PubMed

Frank, Stefan; Aguirre, Aitor; Hescheler, Juergen; Kurian, Leo

2016-01-01

Our conception of the human genome, long focused on the 2% that codes for proteins, has profoundly changed since its first draft assembly in 2001. Since then, an unanticipatedly expansive functionality and convolution has been attributed to the majority of the genome that is transcribed in a cell-type/context-specific manner into transcripts with no apparent protein coding ability. While the majority of these transcripts, currently annotated as long non-coding RNAs (lncRNAs), are functionally uncharacterized, their prominent role in embryonic development and tissue homeostasis, especially in the context of the heart, is emerging. In this review, we summarize and discuss the latest advances in understanding the relevance of lncRNAs in (re)building the heart.

A systemic identification approach for primary transcription start site of Arabidopsis miRNAs from multidimensional omics data.

PubMed

You, Qi; Yan, Hengyu; Liu, Yue; Yi, Xin; Zhang, Kang; Xu, Wenying; Su, Zhen

2017-05-01

The 22-nucleotide non-coding microRNAs (miRNAs) are mostly transcribed by RNA polymerase II and are similar to protein-coding genes. Unlike the clear process from stem-loop precursors to mature miRNAs, the primary transcriptional regulation of miRNA, especially in plants, still needs to be further clarified, including the original transcription start site, functional cis-elements and primary transcript structures. Due to several well-characterized transcription signals in the promoter region, we proposed a systemic approach integrating multidimensional "omics" (including genomics, transcriptomics, and epigenomics) data to improve the genome-wide identification of primary miRNA transcripts. Here, we used the model plant Arabidopsis thaliana to improve the ability to identify candidate promoter locations in intergenic miRNAs and to determine rules for identifying primary transcription start sites of miRNAs by integrating high-throughput omics data, such as the DNase I hypersensitive sites, chromatin immunoprecipitation-sequencing of polymerase II and H3K4me3, as well as high throughput transcriptomic data. As a result, 93% of refined primary transcripts could be confirmed by the primer pairs from a previous study. Cis-element and secondary structure analyses also supported the feasibility of our results. This work will contribute to the primary transcriptional regulatory analysis of miRNAs, and the conserved regulatory pattern may be a suitable miRNA characteristic in other plant species.

The Long Noncoding RNA Transcriptome of Dictyostelium discoideum Development.

PubMed

Rosengarten, Rafael D; Santhanam, Balaji; Kokosar, Janez; Shaulsky, Gad

2017-02-09

Dictyostelium discoideum live in the soil as single cells, engulfing bacteria and growing vegetatively. Upon starvation, tens of thousands of amoebae enter a developmental program that includes aggregation, multicellular differentiation, and sporulation. Major shifts across the protein-coding transcriptome accompany these developmental changes. However, no study has presented a global survey of long noncoding RNAs (ncRNAs) in D. discoideum To characterize the antisense and long intergenic noncoding RNA (lncRNA) transcriptome, we analyzed previously published developmental time course samples using an RNA-sequencing (RNA-seq) library preparation method that selectively depletes ribosomal RNAs (rRNAs). We detected the accumulation of transcripts for 9833 protein-coding messenger RNAs (mRNAs), 621 lncRNAs, and 162 putative antisense RNAs (asRNAs). The noncoding RNAs were interspersed throughout the genome, and were distinct in expression level, length, and nucleotide composition. The noncoding transcriptome displayed a temporal profile similar to the coding transcriptome, with stages of gradual change interspersed with larger leaps. The transcription profiles of some noncoding RNAs were strongly correlated with known differentially expressed coding RNAs, hinting at a functional role for these molecules during development. Examining the mitochondrial transcriptome, we modeled two novel antisense transcripts. We applied yet another ribosomal depletion method to a subset of the samples to better retain transfer RNA (tRNA) transcripts. We observed polymorphisms in tRNA anticodons that suggested a post-transcriptional means by which D. discoideum compensates for codons missing in the genomic complement of tRNAs. We concluded that the prevalence and characteristics of long ncRNAs indicate that these molecules are relevant to the progression of molecular and cellular phenotypes during development. Copyright © 2017 Rosengarten et al.

Banana NAC transcription factor MusaNAC042 is positively associated with drought and salinity tolerance.

PubMed

Tak, Himanshu; Negi, Sanjana; Ganapathi, T R

2017-03-01

Banana is an important fruit crop and its yield is hampered by multiple abiotic stress conditions encountered during its growth. The NAC (NAM, ATAF, and CUC) transcription factors are involved in plant response to biotic and abiotic stresses. In the present study, we studied the induction of banana NAC042 transcription factor in drought and high salinity conditions and its overexpression in transgenic banana to improve drought and salinity tolerance. MusaNAC042 expression was positively associated with stress conditions like salinity and drought and it encoded a nuclear localized protein. Transgenic lines of banana cultivar Rasthali overexpressing MusaNAC042 were generated by Agrobacterium-mediated transformation of banana embryogenic cells and T-DNA insertion was confirmed by PCR and Southern blot analysis. Our results using leaf disc assay indicated that transgenic banana lines were able to tolerate drought and high salinity stress better than the control plants and retained higher level of total chlorophyll and lower level of MDA content (malondialdehyde). Transgenic lines analyzed for salinity (250 mM NaCl) and drought (Soil gravimetric water content 0.15) tolerance showed higher proline content, better Fv/Fm ratio, and lower levels of MDA content than control suggesting that MusaNAC042 may be involved in responses to higher salinity and drought stresses in banana. Expression of several abiotic stress-related genes like those coding for CBF/DREB, LEA, and WRKY factors was altered in transgenic lines indicating that MusaNAC042 is an efficient modulator of abiotic stress response in banana.

Mucin acts as a nutrient source and a signal for the differential expression of genes coding for cellular processes and virulence factors in Acinetobacter baumannii

PubMed Central

Ohneck, Emily J.; Arivett, Brock A.; Fiester, Steven E.; Wood, Cecily R.; Metz, Maeva L.; Simeone, Gabriella M.

2018-01-01

The capacity of Acinetobacter baumannii to persist and cause infections depends on its interaction with abiotic and biotic surfaces, including those found on medical devices and host mucosal surfaces. However, the extracellular stimuli affecting these interactions are poorly understood. Based on our previous observations, we hypothesized that mucin, a glycoprotein secreted by lung epithelial cells, particularly during respiratory infections, significantly alters A. baumannii’s physiology and its interaction with the surrounding environment. Biofilm, virulence and growth assays showed that mucin enhances the interaction of A. baumannii ATCC 19606T with abiotic and biotic surfaces and its cytolytic activity against epithelial cells while serving as a nutrient source. The global effect of mucin on the physiology and virulence of this pathogen is supported by RNA-Seq data showing that its presence in a low nutrient medium results in the differential transcription of 427 predicted protein-coding genes. The reduced expression of ion acquisition genes and the increased transcription of genes coding for energy production together with the detection of mucin degradation indicate that this host glycoprotein is a nutrient source. The increased expression of genes coding for adherence and biofilm biogenesis on abiotic and biotic surfaces, the degradation of phenylacetic acid and the production of an active type VI secretion system further supports the role mucin plays in virulence. Taken together, our observations indicate that A. baumannii recognizes mucin as an environmental signal, which triggers a response cascade that allows this pathogen to acquire critical nutrients and promotes host-pathogen interactions that play a role in the pathogenesis of bacterial infections. PMID:29309434

Identification of functional elements and regulatory circuits by Drosophila modENCODE

DOE Office of Scientific and Technical Information (OSTI.GOV)

Roy, Sushmita; Ernst, Jason; Kharchenko, Peter V.

2010-12-22

To gain insight into how genomic information is translated into cellular and developmental programs, the Drosophila model organism Encyclopedia of DNA Elements (modENCODE) project is comprehensively mapping transcripts, histone modifications, chromosomal proteins, transcription factors, replication proteins and intermediates, and nucleosome properties across a developmental time course and in multiple cell lines. We have generated more than 700 data sets and discovered protein-coding, noncoding, RNA regulatory, replication, and chromatin elements, more than tripling the annotated portion of the Drosophila genome. Correlated activity patterns of these elements reveal a functional regulatory network, which predicts putative new functions for genes, reveals stage- andmore » tissue-specific regulators, and enables gene-expression prediction. Our results provide a foundation for directed experimental and computational studies in Drosophila and related species and also a model for systematic data integration toward comprehensive genomic and functional annotation. Several years after the complete genetic sequencing of many species, it is still unclear how to translate genomic information into a functional map of cellular and developmental programs. The Encyclopedia of DNA Elements (ENCODE) (1) and model organism ENCODE (modENCODE) (2) projects use diverse genomic assays to comprehensively annotate the Homo sapiens (human), Drosophila melanogaster (fruit fly), and Caenorhabditis elegans (worm) genomes, through systematic generation and computational integration of functional genomic data sets. Previous genomic studies in flies have made seminal contributions to our understanding of basic biological mechanisms and genome functions, facilitated by genetic, experimental, computational, and manual annotation of the euchromatic and heterochromatic genome (3), small genome size, short life cycle, and a deep knowledge of development, gene function, and chromosome biology. The functions of {approx}40% of the protein and nonprotein-coding genes [FlyBase 5.12 (4)] have been determined from cDNA collections (5, 6), manual curation of gene models (7), gene mutations and comprehensive genome-wide RNA interference screens (8-10), and comparative genomic analyses (11, 12). The Drosophila modENCODE project has generated more than 700 data sets that profile transcripts, histone modifications and physical nucleosome properties, general and specific transcription factors (TFs), and replication programs in cell lines, isolated tissues, and whole organisms across several developmental stages (Fig. 1). Here, we computationally integrate these data sets and report (i) improved and additional genome annotations, including full-length proteincoding genes and peptides as short as 21 amino acids; (ii) noncoding transcripts, including 132 candidate structural RNAs and 1608 nonstructural transcripts; (iii) additional Argonaute (Ago)-associated small RNA genes and pathways, including new microRNAs (miRNAs) encoded within protein-coding exons and endogenous small interfering RNAs (siRNAs) from 3-inch untranslated regions; (iv) chromatin 'states' defined by combinatorial patterns of 18 chromatin marks that are associated with distinct functions and properties; (v) regions of high TF occupancy and replication activity with likely epigenetic regulation; (vi)mixed TF and miRNA regulatory networks with hierarchical structure and enriched feed-forward loops; (vii) coexpression- and co-regulation-based functional annotations for nearly 3000 genes; (viii) stage- and tissue-specific regulators; and (ix) predictive models of gene expression levels and regulator function.« less

Deep sequencing approaches for the analysis of prokaryotic transcriptional boundaries and dynamics.

PubMed

James, Katherine; Cockell, Simon J; Zenkin, Nikolay

2017-05-01

The identification of the protein-coding regions of a genome is straightforward due to the universality of start and stop codons. However, the boundaries of the transcribed regions, conditional operon structures, non-coding RNAs and the dynamics of transcription, such as pausing of elongation, are non-trivial to identify, even in the comparatively simple genomes of prokaryotes. Traditional methods for the study of these areas, such as tiling arrays, are noisy, labour-intensive and lack the resolution required for densely-packed bacterial genomes. Recently, deep sequencing has become increasingly popular for the study of the transcriptome due to its lower costs, higher accuracy and single nucleotide resolution. These methods have revolutionised our understanding of prokaryotic transcriptional dynamics. Here, we review the deep sequencing and data analysis techniques that are available for the study of transcription in prokaryotes, and discuss the bioinformatic considerations of these analyses. Copyright © 2017 Elsevier Inc. All rights reserved.

A Rapid, Extensive, and Transient Transcriptional Response to Estrogen Signaling in Breast Cancer Cells

PubMed Central

Hah, Nasun; Danko, Charles G.; Core, Leighton; Waterfall, Joshua J.; Siepel, Adam; Lis, John T.; Kraus, W. Lee

2011-01-01

Summary We report the immediate effects of estrogen signaling on the transcriptome of breast cancer cells using Global Run-On and sequencing (GRO-seq). The data were analyzed using a new bioinformatic approach that allowed us to identify transcripts directly from the GRO-seq data. We found that estrogen signaling directly regulates a strikingly large fraction of the transcriptome in a rapid, robust, and unexpectedly transient manner. In addition to protein coding genes, estrogen regulates the distribution and activity of all three RNA polymerases, and virtually every class of non-coding RNA that has been described to date. We also identified a large number of previously undetected estrogen-regulated intergenic transcripts, many of which are found proximal to estrogen receptor binding sites. Collectively, our results provide the most comprehensive measurement of the primary and immediate estrogen effects to date and a resource for understanding rapid signal-dependent transcription in other systems. PMID:21549415

Methods for Modeling Brassinosteroid-Mediated Signaling in Plant Development.

PubMed

Frigola, David; Caño-Delgado, Ana I; Ibañes, Marta

2017-01-01

Mathematical modeling of biological processes is a useful tool to draw conclusions that are contained in the data, but not directly reachable, as well as to make predictions and select the most efficient follow-up experiments. Here we outline a method to model systems of a few proteins that interact transcriptionally and/or posttranscriptionally, by representing the system as Ordinary Differential Equations and to study the model dynamics and stationary states. We exemplify this method by focusing on the regulation by the brassinosteroid (BR) signaling component BRASSINOSTEROID INSENSITIVE1 ETHYL METHYL SULFONATE SUPPRESSOR1 (BES1) of BRAVO, a quiescence-regulating transcription factor expressed in the quiescent cells of Arabidopsis thaliana roots. The method to extract the stationary states and the dynamics is provided as a Mathematica code and requires basic knowledge of the Mathematica software to be executed.

Overexpression of Grain Amaranth (Amaranthus hypochondriacus) AhERF or AhDOF Transcription Factors in Arabidopsis thaliana Increases Water Deficit- and Salt-Stress Tolerance, Respectively, via Contrasting Stress-Amelioration Mechanisms

PubMed Central

Massange-Sánchez, Julio A.; Palmeros-Suárez, Paola A.; Espitia-Rangel, Eduardo; Rodríguez-Arévalo, Isaac; Sánchez-Segura, Lino; Martínez-Gallardo, Norma A.; Alatorre-Cobos, Fulgencio; Tiessen, Axel; Délano-Frier, John P.

2016-01-01

Two grain amaranth transcription factor (TF) genes were overexpressed in Arabidopsis plants. The first, coding for a group VII ethylene response factor TF (i.e., AhERF-VII) conferred tolerance to water-deficit stress (WS) in transgenic Arabidopsis without affecting vegetative or reproductive growth. A significantly lower water-loss rate in detached leaves coupled to a reduced stomatal opening in leaves of plants subjected to WS was associated with this trait. WS tolerance was also associated with an increased antioxidant enzyme activity and the accumulation of putative stress-related secondary metabolites. However, microarray and GO data did not indicate an obvious correlation between WS tolerance, stomatal closure, and abscisic acid (ABA)-related signaling. This scenario suggested that stomatal closure during WS in these plants involved ABA-independent mechanisms, possibly involving reactive oxygen species (ROS). WS tolerance may have also involved other protective processes, such as those employed for methyl glyoxal detoxification. The second, coding for a class A and cluster I DNA binding with one finger TF (i.e., AhDof-AI) provided salt-stress (SS) tolerance with no evident fitness penalties. The lack of an obvious development-related phenotype contrasted with microarray and GO data showing an enrichment of categories and genes related to developmental processes, particularly flowering. SS tolerance also correlated with increased superoxide dismutase activity but not with augmented stomatal closure. Additionally, microarray and GO data indicated that, contrary to AhERF-VII, SS tolerance conferred by AhDof-AI in Arabidopsis involved ABA-dependent and ABA-independent stress amelioration mechanisms. PMID:27749893

[Long non-coding RNAs in the pathophysiology of atherosclerosis].

PubMed

Novak, Jan; Vašků, Julie Bienertová; Souček, Miroslav

2018-01-01

The human genome contains about 22 000 protein-coding genes that are transcribed to an even larger amount of messenger RNAs (mRNA). Interestingly, the results of the project ENCODE from 2012 show, that despite up to 90 % of our genome being actively transcribed, protein-coding mRNAs make up only 2-3 % of the total amount of the transcribed RNA. The rest of RNA transcripts is not translated to proteins and that is why they are referred to as "non-coding RNAs". Earlier the non-coding RNA was considered "the dark matter of genome", or "the junk", whose genes has accumulated in our DNA during the course of evolution. Today we already know that non-coding RNAs fulfil a variety of regulatory functions in our body - they intervene into epigenetic processes from chromatin remodelling to histone methylation, or into the transcription process itself, or even post-transcription processes. Long non-coding RNAs (lncRNA) are one of the classes of non-coding RNAs that have more than 200 nucleotides in length (non-coding RNAs with less than 200 nucleotides in length are called small non-coding RNAs). lncRNAs represent a widely varied and large group of molecules with diverse regulatory functions. We can identify them in all thinkable cell types or tissues, or even in an extracellular space, which includes blood, specifically plasma. Their levels change during the course of organogenesis, they are specific to different tissues and their changes also occur along with the development of different illnesses, including atherosclerosis. This review article aims to present lncRNAs problematics in general and then focuses on some of their specific representatives in relation to the process of atherosclerosis (i.e. we describe lncRNA involvement in the biology of endothelial cells, vascular smooth muscle cells or immune cells), and we further describe possible clinical potential of lncRNA, whether in diagnostics or therapy of atherosclerosis and its clinical manifestations.Key words: atherosclerosis - lincRNA - lncRNA - MALAT - MIAT.

Quantifying circular RNA expression from RNA-seq data using model-based framework.

PubMed

Li, Musheng; Xie, Xueying; Zhou, Jing; Sheng, Mengying; Yin, Xiaofeng; Ko, Eun-A; Zhou, Tong; Gu, Wanjun

2017-07-15

Circular RNAs (circRNAs) are a class of non-coding RNAs that are widely expressed in various cell lines and tissues of many organisms. Although the exact function of many circRNAs is largely unknown, the cell type-and tissue-specific circRNA expression has implicated their crucial functions in many biological processes. Hence, the quantification of circRNA expression from high-throughput RNA-seq data is becoming important to ascertain. Although many model-based methods have been developed to quantify linear RNA expression from RNA-seq data, these methods are not applicable to circRNA quantification. Here, we proposed a novel strategy that transforms circular transcripts to pseudo-linear transcripts and estimates the expression values of both circular and linear transcripts using an existing model-based algorithm, Sailfish. The new strategy can accurately estimate transcript expression of both linear and circular transcripts from RNA-seq data. Several factors, such as gene length, amount of expression and the ratio of circular to linear transcripts, had impacts on quantification performance of circular transcripts. In comparison to count-based tools, the new computational framework had superior performance in estimating the amount of circRNA expression from both simulated and real ribosomal RNA-depleted (rRNA-depleted) RNA-seq datasets. On the other hand, the consideration of circular transcripts in expression quantification from rRNA-depleted RNA-seq data showed substantial increased accuracy of linear transcript expression. Our proposed strategy was implemented in a program named Sailfish-cir. Sailfish-cir is freely available at https://github.com/zerodel/Sailfish-cir . tongz@medicine.nevada.edu or wanjun.gu@gmail.com. Supplementary data are available at Bioinformatics online. © The Author (2017). Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com

Multiple PAR and E4BP4 bZIP transcription factors in zebrafish: diverse spatial and temporal expression patterns.

PubMed

Ben-Moshe, Zohar; Vatine, Gad; Alon, Shahar; Tovin, Adi; Mracek, Philipp; Foulkes, Nicholas S; Gothilf, Yoav

2010-09-01

Circadian rhythms of physiology and behavior are generated by an autonomous circadian oscillator that is synchronized daily with the environment, mainly by light input. The PAR subfamily of transcriptional activators and the related E4BP4 repressor belonging to the basic leucine zipper (bZIP) family are clock-controlled genes that are suggested to mediate downstream circadian clock processes and to feedback onto the core oscillator. Here, the authors report the characterization of these genes in the zebrafish, an increasingly important model in the field of chronobiology. Five novel PAR and six novel e4bp4 zebrafish homolog genes were identified using bioinformatic tools and their coding sequences were cloned. Based on their evolutionary relationships, these genes were annotated as ztef2, zhlf1 and zhlf2, zdbp1 and zdbp2, and ze4bp4-1 to -6. The spatial and temporal mRNA expression pattern of each of these factors was characterized in zebrafish embryos in the context of a functional circadian clock and regulation by light. Nine of the factors exhibited augmented and rhythmic expression in the pineal gland, a central clock organ in zebrafish. Moreover, these genes were found to be regulated, to variable extents, by the circadian clock and/or by light. Differential expression patterns of multiple paralogs in zebrafish suggest multiple roles for these factors within the vertebrate circadian clock. This study, in the genetically accessible zebrafish model, lays the foundation for further research regarding the involvement and specific roles of PAR and E4BP4 transcription factors in the vertebrate circadian clock mechanism.

Systematical analysis of cutaneous squamous cell carcinoma network of microRNAs, transcription factors, and target and host genes.

PubMed

Wang, Ning; Xu, Zhi-Wen; Wang, Kun-Hao

2014-01-01

MicroRNAs (miRNAs) are small non-coding RNA molecules found in multicellular eukaryotes which are implicated in development of cancer, including cutaneous squamous cell carcinoma (cSCC). Expression is controlled by transcription factors (TFs) that bind to specific DNA sequences, thereby controlling the flow (or transcription) of genetic information from DNA to messenger RNA. Interactions result in biological signal control networks. Molecular components involved in cSCC were here assembled at abnormally expressed, related and global levels. Networks at these three levels were constructed with corresponding biological factors in term of interactions between miRNAs and target genes, TFs and miRNAs, and host genes and miRNAs. Up/down regulation or mutation of the factors were considered in the context of the regulation and significant patterns were extracted. Participants of the networks were evaluated based on their expression and regulation of other factors. Sub-networks with two core TFs, TP53 and EIF2C2, as the centers are identified. These share self-adapt feedback regulation in which a mutual restraint exists. Up or down regulation of certain genes and miRNAs are discussed. Some, for example the expression of MMP13, were in line with expectation while others, including FGFR3, need further investigation of their unexpected behavior. The present research suggests that dozens of components, miRNAs, TFs, target genes and host genes included, unite as networks through their regulation to function systematically in human cSCC. Networks built under the currently available sources provide critical signal controlling pathways and frequent patterns. Inappropriate controlling signal flow from abnormal expression of key TFs may push the system into an incontrollable situation and therefore contributes to cSCC development.

Dehydration-induced WRKY genes from tobacco and soybean respond to jasmonic acid treatments in BY-2 cell culture.

PubMed

Rabara, Roel C; Tripathi, Prateek; Lin, Jun; Rushton, Paul J

2013-02-15

Drought is one of the important environmental factors affecting crop production worldwide and therefore understanding the molecular response of plant to stress is an important step in crop improvement. WRKY transcription factors are one of the 10 largest transcription factor families across the green lineage. In this study, highly upregulated dehydration-induced WRKY and enzyme-coding genes from tobacco and soybean were selected from microarray data for promoter analyses. Putative stress-related cis-regulatory elements such as TGACG motif, ABRE-like elements; W and G-like sequences were identified by an in silico analyses of promoter region of the selected genes. GFP quantification of transgenic BY-2 cell culture showed these promoters direct higher expression in-response to 100 μM JA treatment compared to 100 μM ABA, 10% PEG and 85 mM NaCl treatments. Thus promoter activity upon JA treatment and enrichment of MeJA-responsive elements in the promoter of the selected genes provides insights for these genes to be jasmonic acid responsive with potential of mediating cross-talk during dehydration responses. Copyright © 2013 Elsevier Inc. All rights reserved.

Identification of the SRC pyrimidine-binding protein (SPy) as hnRNP K: implications in the regulation of SRC1A transcription

PubMed Central

Ritchie, Shawn A.; Pasha, Mohammed K.; Batten, Danielle J. P.; Sharma, Rajendra K.; Olson, Douglas J. H.; Ross, Andrew R. S.; Bonham, Keith

2003-01-01

The human SRC gene encodes pp60c–src, a non-receptor tyrosine kinase involved in numerous signaling pathways. Activation or overexpression of c-Src has also been linked to a number of important human cancers. Transcription of the SRC gene is complex and regulated by two closely linked but highly dissimilar promoters, each associated with its own distinct non-coding exon. In many tissues SRC expression is regulated by the housekeeping-like SRC1A promoter. In addition to other regulatory elements, three substantial polypurine:polypyrimidine (TC) tracts within this promoter are required for full transcriptional activity. Previously, we described an unusual factor called SRC pyrimidine-binding protein (SPy) that could bind to two of these TC tracts in their double-stranded form, but was also capable of interacting with higher affinity to all three pyrimidine tracts in their single-stranded form. Mutations in the TC tracts, which abolished the ability of SPy to interact with its double-stranded DNA target, significantly reduced SRC1A promoter activity, especially in concert with mutations in critical Sp1 binding sites. Here we expand upon our characterization of this interesting factor and describe the purification of SPy from human SW620 colon cancer cells using a DNA affinity-based approach. Subsequent in-gel tryptic digestion of purified SPy followed by MALDI-TOF mass spectrometric analysis identified SPy as heterogeneous nuclear ribonucleoprotein K (hnRNP K), a known nucleic-acid binding protein implicated in various aspects of gene expression including transcription. These data provide new insights into the double- and single-stranded DNA-binding specificity, as well as functional properties of hnRNP K, and suggest that hnRNP K is a critical component of SRC1A transcriptional processes. PMID:12595559

5′UTR of the Neurogenic bHLH Nex1/MATH-2/NeuroD6 Gene Is Regulated by Two Distinct Promoters Through CRE and C/EBP Binding Sites

PubMed Central

Uittenbogaard, Martine; Martinka, Debra L.; Johnson, Peter F.; Vinson, Charles; Chiaramello, Anne

2009-01-01

Expression of the bHLH transcription factor Nex1/MATH-2/NeuroD6, a member of the NeuroD subfamily, parallels overt neuronal differentiation and synaptogenesis during brain development. Our previous studies have shown that Nex1 is a critical effector of the NGF pathway and promotes neuronal differentiation and survival of PC12 cells in the absence of growth factors. In this study, we investigated the transcriptional regulation of the Nex1 gene during NGF-induced neuronal differentiation. We found that Nex1 expression is under the control of two conserved promoters, Nex1-P1 and Nex1-P2, located in two distinct non-coding exons. Both promoters are TATA-less with multiple transcription start sites, and are activated on NGF or cAMP exposure. Luciferase-reporter assays showed that the Nex1-P2 promoter activity is stronger than the Nex1-P1 promoter activity, which supports the previously reported differential expression levels of Nex1 transcripts throughout brain development. Using a combination of DNaseI footprinting, EMSA assays, and site-directed mutagenesis, we identified the essential regulatory elements within the first 2 kb of the Nex1 5′UTR. The Nex1-P1 promoter is mainly regulated by a conserved CRE element, whereas the Nex1-P2 promoter is under the control of a conserved C/EBP binding site. Overexpression of wild-type C/EBPβ resulted in increased Nex1-P2 promoter activity in NGF-differentiated PC12 cells. The fact that Nex1 is a target gene of C/EBPβ provides new insight into the C/EBP transcriptional cascade known to promote neurogenesis, while repressing gliogenesis. PMID:17075921

The long non-coding RNA LSINCT5 promotes malignancy in non-small cell lung cancer by stabilizing HMGA2.

PubMed

Tian, Yuheng; Zhang, Lina; Chen, Shuwen; Ma, Yuan; Liu, Yanyan

2018-06-08

Long non-coding RNAs (lncRNAs) can actively participate in tumorigenesis in various cancers. However, the involvement of lncRNA long stress induced non-coding transcripts 5 (LSINCT5) in non-small cell lung cancer (NSCLC) remains largely unknown. Here we showed a novel lncRNA signature in NSCLC through lncRNA profiling. Increased LSINCT5 expression positively correlates with malignant clinicopathological features and poor survival. LSINCT5 can promote migration and viability of various NSCLC cells in vitro and also enhance lung cancer progression in vivo. RNA immunoprecipitation followed by mass spectrometry has identified that LSINCT5 interacts with HMGA2. This physical interaction can increase the stability of HMGA2 by inhibiting proteasome-mediated degradation. Therefore, LSINCT5 may possibly contribute to NSCLC tumorigenesis by stabilizing the oncogenic factor of HMGA2. This novel LSINCT5/HMGA2 axis can modulate lung cancer progression and might be a promising target for pharmacological intervention.

Identification of significantly mutated regions across cancer types highlights a rich landscape of functional molecular alterations

PubMed Central

Araya, Carlos L.; Cenik, Can; Reuter, Jason A.; Kiss, Gert; Pande, Vijay S.; Snyder, Michael P.; Greenleaf, William J.

2015-01-01

Cancer sequencing studies have primarily identified cancer-driver genes by the accumulation of protein-altering mutations. An improved method would be annotation-independent, sensitive to unknown distributions of functions within proteins, and inclusive of non-coding drivers. We employed density-based clustering methods in 21 tumor types to detect variably-sized significantly mutated regions (SMRs). SMRs reveal recurrent alterations across a spectrum of coding and non-coding elements, including transcription factor binding sites and untranslated regions mutated in up to ∼15% of specific tumor types. SMRs reveal spatial clustering of mutations at molecular domains and interfaces, often with associated changes in signaling. Mutation frequencies in SMRs demonstrate that distinct protein regions are differentially mutated among tumor types, as exemplified by a linker region of PIK3CA in which biophysical simulations suggest mutations affect regulatory interactions. The functional diversity of SMRs underscores both the varied mechanisms of oncogenic misregulation and the advantage of functionally-agnostic driver identification. PMID:26691984

Single-Nucleosome Mapping of Histone Modifications in S. cerevisiae

PubMed Central

Kim, Minkyu; Buratowski, Stephen; Schreiber, Stuart L; Friedman, Nir

2005-01-01

Covalent modification of histone proteins plays a role in virtually every process on eukaryotic DNA, from transcription to DNA repair. Many different residues can be covalently modified, and it has been suggested that these modifications occur in a great number of independent, meaningful combinations. Published low-resolution microarray studies on the combinatorial complexity of histone modification patterns suffer from confounding effects caused by the averaging of modification levels over multiple nucleosomes. To overcome this problem, we used a high-resolution tiled microarray with single-nucleosome resolution to investigate the occurrence of combinations of 12 histone modifications on thousands of nucleosomes in actively growing S. cerevisiae. We found that histone modifications do not occur independently; there are roughly two groups of co-occurring modifications. One group of lysine acetylations shows a sharply defined domain of two hypo-acetylated nucleosomes, adjacent to the transcriptional start site, whose occurrence does not correlate with transcription levels. The other group consists of modifications occurring in gradients through the coding regions of genes in a pattern associated with transcription. We found no evidence for a deterministic code of many discrete states, but instead we saw blended, continuous patterns that distinguish nucleosomes at one location (e.g., promoter nucleosomes) from those at another location (e.g., over the 3′ ends of coding regions). These results are consistent with the idea of a simple, redundant histone code, in which multiple modifications share the same role. PMID:16122352

Non-coding RNA networks in cancer.

PubMed

Anastasiadou, Eleni; Jacob, Leni S; Slack, Frank J

2018-01-01

Thousands of unique non-coding RNA (ncRNA) sequences exist within cells. Work from the past decade has altered our perception of ncRNAs from 'junk' transcriptional products to functional regulatory molecules that mediate cellular processes including chromatin remodelling, transcription, post-transcriptional modifications and signal transduction. The networks in which ncRNAs engage can influence numerous molecular targets to drive specific cell biological responses and fates. Consequently, ncRNAs act as key regulators of physiological programmes in developmental and disease contexts. Particularly relevant in cancer, ncRNAs have been identified as oncogenic drivers and tumour suppressors in every major cancer type. Thus, a deeper understanding of the complex networks of interactions that ncRNAs coordinate would provide a unique opportunity to design better therapeutic interventions.

Md-miR156ab and Md-miR395 Target WRKY Transcription Factors to Influence Apple Resistance to Leaf Spot Disease.

PubMed

Zhang, Qiulei; Li, Yang; Zhang, Yi; Wu, Chuanbao; Wang, Shengnan; Hao, Li; Wang, Shengyuan; Li, Tianzhong

2017-01-01

MicroRNAs (miRNAs) are key regulators of gene expression that post-transcriptionally regulate transcription factors involved in plant physiological activities. Little is known about the effects of miRNAs in disease resistance in apple ( Malus × domestica ). We globally profiled miRNAs in the apple cultivar Golden Delicious (GD) infected or not with the apple leaf spot fungus Alternaria alternaria f. sp. mali (ALT1), and identified 58 miRNAs that exhibited more than a 2-fold upregulation upon ALT1 infection. We identified a pair of miRNAs that target protein-coding genes involved in the defense response against fungal pathogens; Md-miR156ab targets a novel WRKY transcription factor, MdWRKYN1, which harbors a TIR and a WRKY domain. Md-miR395 targets another transcription factor, MdWRKY26, which contains two WRKY domains. Real-time PCR analysis showed that Md-miR156ab and Md-miR395 levels increased, while MdWRKYN1 and MdWRKY26 expression decreased in ALT1-inoculated GD leaves; furthermore, the overexpression of Md-miR156ab and Md-miR395 resulted in a significant reduction in MdWRKYN1 and MdWRKY26 expression. To investigate whether these miRNAs and their targets play a crucial role in plant defense, we overexpressed MdWRKYN1 or knocked down Md-miR156ab activity, which in both cases enhanced the disease resistance of the plants by upregulating the expression of the WRKY-regulated pathogenesis-related (PR) protein-encoding genes MdPR3-1, MdPR3-2, MdPR4, MdPR5, MdPR10-1 , and MdPR10-2 . In a similar analysis, we overexpressed MdWRKY26 or suppressed Md-miR395 activity, and found that many PR protein-encoding genes were also regulated by MdWRKY26 . In GD, ALT-induced Md-miR156ab and Md-miR395 suppress MdWRKYN1 and MdWRKY26 expression, thereby decreasing the expression of some PR genes, and resulting in susceptibility to ALT1.

Regulation of expression of two LY-6 family genes by intron retention and transcription induced chimerism

PubMed Central

Calvanese, Vincenzo; Mallya, Meera; Campbell, R Duncan; Aguado, Begoña

2008-01-01

Background Regulation of the expression of particular genes can rely on mechanisms that are different from classical transcriptional and translational control. The LY6G5B and LY6G6D genes encode LY-6 domain proteins, whose expression seems to be regulated in an original fashion, consisting of an intron retention event which generates, through an early premature stop codon, a non-coding transcript, preventing expression in most cell lines and tissues. Results The MHC LY-6 non-coding transcripts have shown to be stable and very abundant in the cell, and not subject to Nonsense Mediated Decay (NMD). This retention event appears not to be solely dependent on intron features, because in the case of LY6G5B, when the intron is inserted in the artificial context of a luciferase expression plasmid, it is fully spliced but strongly stabilises the resulting luciferase transcript. In addition, by quantitative PCR we found that the retained and spliced forms are differentially expressed in tissues indicating an active regulation of the non-coding transcript. EST database analysis revealed that these genes have an alternative expression pathway with the formation of Transcription Induced Chimeras (TIC). This data was confirmed by RT-PCR, revealing the presence of different transcripts that would encode the chimeric proteins CSNKβ-LY6G5B and G6F-LY6G6D, in which the LY-6 domain would join to a kinase domain and an Ig-like domain, respectively. Conclusion In conclusion, the LY6G5B and LY6G6D intron-retained transcripts are not subjected to NMD and are more abundant than the properly spliced forms. In addition, these genes form chimeric transcripts with their neighbouring same orientation 5' genes. Of interest is the fact that the 5' genes (CSNKβ or G6F) undergo differential splicing only in the context of the chimera (CSNKβ-LY6G5B or G6F-LY6G6C) and not on their own. PMID:18817541

MEF2C loss-of-function mutation contributes to congenital heart defects.

PubMed

Qiao, Xiao-Hui; Wang, Fei; Zhang, Xian-Ling; Huang, Ri-Tai; Xue, Song; Wang, Juan; Qiu, Xing-Biao; Liu, Xing-Yuan; Yang, Yi-Qing

2017-01-01

Congenital heart disease (CHD) is the most common type of developmental abnormality in humans, and is a leading cause for substantially increased morbidity and mortality in affected individuals. Increasing studies demonstrates a pivotal role of genetic defects in the pathogenesis of CHD, and presently mutations in more than 60 genes have been associated with CHD. Nevertheless, CHD is of pronounced genetic heterogeneity, and the genetic basis underpinning CHD in a large proportion of patients remains unclear. In the present study, the whole coding exons and splicing donors/acceptors of the MEF2C gene, which codes for a transcription factor essential for normal cardiovascular development, were sequenced in 200 unrelated patients affected with CHD, and a novel heterozygous missense mutation, p.L38P, was identified in an index patient with patent ductus arteriosus (PDA) and ventricular septal defect (VSD). Genetic scan of the mutation carrier's family members available showed that the mutation was present in all affected family members but absent in unaffected family members. Analysis of the proband's pedigree revealed that the mutation co-segregated with PDA, which was transmitted as an autosomal dominant trait with complete penetrance. The mutation changed the amino acid that was completely conserved evolutionarily, and did not exist in 300 unrelated, ethnically-matched healthy individuals used as controls. Functional deciphers by using a dual-luciferase reporter assay system unveiled that the mutant MEF2C protein had a significantly reduced transcriptional activity. Furthermore, the mutation significantly diminished the synergistic activation between MEF2C and GATA4, another cardiac core transcription factor that has been causally linked to CHD. In conclusion, this is the first report on the association of a MEF2C loss-of-function mutation with an increased vulnerability to CHD in humans, which provides novel insight into the molecular mechanisms underlying CHD, implying potential implications for early diagnosis and timely prophylaxis of CHD.

MEF2C loss-of-function mutation contributes to congenital heart defects

PubMed Central

Qiao, Xiao-Hui; Wang, Fei; Zhang, Xian-Ling; Huang, Ri-Tai; Xue, Song; Wang, Juan; Qiu, Xing-Biao; Liu, Xing-Yuan; Yang, Yi-Qing

2017-01-01

Congenital heart disease (CHD) is the most common type of developmental abnormality in humans, and is a leading cause for substantially increased morbidity and mortality in affected individuals. Increasing studies demonstrates a pivotal role of genetic defects in the pathogenesis of CHD, and presently mutations in more than 60 genes have been associated with CHD. Nevertheless, CHD is of pronounced genetic heterogeneity, and the genetic basis underpinning CHD in a large proportion of patients remains unclear. In the present study, the whole coding exons and splicing donors/acceptors of the MEF2C gene, which codes for a transcription factor essential for normal cardiovascular development, were sequenced in 200 unrelated patients affected with CHD, and a novel heterozygous missense mutation, p.L38P, was identified in an index patient with patent ductus arteriosus (PDA) and ventricular septal defect (VSD). Genetic scan of the mutation carrier's family members available showed that the mutation was present in all affected family members but absent in unaffected family members. Analysis of the proband's pedigree revealed that the mutation co-segregated with PDA, which was transmitted as an autosomal dominant trait with complete penetrance. The mutation changed the amino acid that was completely conserved evolutionarily, and did not exist in 300 unrelated, ethnically-matched healthy individuals used as controls. Functional deciphers by using a dual-luciferase reporter assay system unveiled that the mutant MEF2C protein had a significantly reduced transcriptional activity. Furthermore, the mutation significantly diminished the synergistic activation between MEF2C and GATA4, another cardiac core transcription factor that has been causally linked to CHD. In conclusion, this is the first report on the association of a MEF2C loss-of-function mutation with an increased vulnerability to CHD in humans, which provides novel insight into the molecular mechanisms underlying CHD, implying potential implications for early diagnosis and timely prophylaxis of CHD. PMID:29104469

Transcriptator: An Automated Computational Pipeline to Annotate Assembled Reads and Identify Non Coding RNA.

PubMed

Tripathi, Kumar Parijat; Evangelista, Daniela; Zuccaro, Antonio; Guarracino, Mario Rosario

2015-01-01

RNA-seq is a new tool to measure RNA transcript counts, using high-throughput sequencing at an extraordinary accuracy. It provides quantitative means to explore the transcriptome of an organism of interest. However, interpreting this extremely large data into biological knowledge is a problem, and biologist-friendly tools are lacking. In our lab, we developed Transcriptator, a web application based on a computational Python pipeline with a user-friendly Java interface. This pipeline uses the web services available for BLAST (Basis Local Search Alignment Tool), QuickGO and DAVID (Database for Annotation, Visualization and Integrated Discovery) tools. It offers a report on statistical analysis of functional and Gene Ontology (GO) annotation's enrichment. It helps users to identify enriched biological themes, particularly GO terms, pathways, domains, gene/proteins features and protein-protein interactions related informations. It clusters the transcripts based on functional annotations and generates a tabular report for functional and gene ontology annotations for each submitted transcript to the web server. The implementation of QuickGo web-services in our pipeline enable the users to carry out GO-Slim analysis, whereas the integration of PORTRAIT (Prediction of transcriptomic non coding RNA (ncRNA) by ab initio methods) helps to identify the non coding RNAs and their regulatory role in transcriptome. In summary, Transcriptator is a useful software for both NGS and array data. It helps the users to characterize the de-novo assembled reads, obtained from NGS experiments for non-referenced organisms, while it also performs the functional enrichment analysis of differentially expressed transcripts/genes for both RNA-seq and micro-array experiments. It generates easy to read tables and interactive charts for better understanding of the data. The pipeline is modular in nature, and provides an opportunity to add new plugins in the future. Web application is freely available at: http://www-labgtp.na.icar.cnr.it/Transcriptator.

Microbial metatranscriptomics in a permanent marine oxygen minimum zone.

PubMed

Stewart, Frank J; Ulloa, Osvaldo; DeLong, Edward F

2012-01-01

Simultaneous characterization of taxonomic composition, metabolic gene content and gene expression in marine oxygen minimum zones (OMZs) has potential to broaden perspectives on the microbial and biogeochemical dynamics in these environments. Here, we present a metatranscriptomic survey of microbial community metabolism in the Eastern Tropical South Pacific OMZ off northern Chile. Community RNA was sampled in late austral autumn from four depths (50, 85, 110, 200 m) extending across the oxycline and into the upper OMZ. Shotgun pyrosequencing of cDNA yielded 180,000 to 550,000 transcript sequences per depth. Based on functional gene representation, transcriptome samples clustered apart from corresponding metagenome samples from the same depth, highlighting the discrepancies between metabolic potential and actual transcription. BLAST-based characterizations of non-ribosomal RNA sequences revealed a dominance of genes involved with both oxidative (nitrification) and reductive (anammox, denitrification) components of the marine nitrogen cycle. Using annotations of protein-coding genes as proxies for taxonomic affiliation, we observed depth-specific changes in gene expression by key functional taxonomic groups. Notably, transcripts most closely matching the genome of the ammonia-oxidizing archaeon Nitrosopumilus maritimus dominated the transcriptome in the upper three depths, representing one in five protein-coding transcripts at 85 m. In contrast, transcripts matching the anammox bacterium Kuenenia stuttgartiensis dominated at the core of the OMZ (200 m; 1 in 12 protein-coding transcripts). The distribution of N. maritimus-like transcripts paralleled that of transcripts matching ammonia monooxygenase genes, which, despite being represented by both bacterial and archaeal sequences in the community DNA, were dominated (> 99%) by archaeal sequences in the RNA, suggesting a substantial role for archaeal nitrification in the upper OMZ. These data, as well as those describing other key OMZ metabolic processes (e.g. sulfur oxidation), highlight gene-specific expression patterns in the context of the entire community transcriptome, as well as identify key functional groups for taxon-specific genomic profiling. © 2011 Society for Applied Microbiology and Blackwell Publishing Ltd.

Transcription Factor Amr1 Induces Melanin Biosynthesis and Suppresses Virulence in Alternaria brassicicola

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cho, Yangrae; Srivastava, Akhil; Ohm, Robin A.

2012-05-01

Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen. Several A. brassicicola genes have been characterized as affecting pathogenesis of Brassica species. To study regulatory mechanisms of pathogenesis, we mined 421 genes in silico encoding putative transcription factors in a machine-annotated, draft genome sequence of A. brassicicola. In this study, targeted gene disruption mutants for 117 of the transcription factor genes were produced and screened. Three of these genes were associated with pathogenesis. Disruption mutants of one gene (AbPacC) were nonpathogenic and another gene (AbVf8) caused lesions less than half the diameter of wild-type lesions. Unexpectedly, mutants of themore » third gene, Amr1, caused lesions with a two-fold larger diameter than the wild type and complementation mutants. Amr1 is a homolog of Cmr1, a transcription factor that regulates melanin biosynthesis in several fungi. We created gene deletion mutants of ?amr1 and characterized their phenotypes. The ?amr1 mutants used pectin as a carbon source more efficiently than the wild type, were melanin-deficient, and more sensitive to UV light and glucanase digestion. The AMR1 protein was localized in the nuclei of hyphae and in highly melanized conidia during the late stage of plant pathogenesis. RNA-seq analysis revealed that three genes in the melanin biosynthesis pathway, along with the deleted Amr1 gene, were expressed at low levels in the mutants. In contrast, many hydrolytic enzyme-coding genes were expressed at higher levels in the mutants than in the wild type during pathogenesis. The results of this study suggested that a gene important for survival in nature negatively affected virulence, probably by a less efficient use of plant cell-wall materials. We speculate that the functions of the Amr1 gene are important to the success of A. brassicicola as a competitive saprophyte and plant parasite.« less

Identification of heavy-ion radiation-induced microRNAs in rice

NASA Astrophysics Data System (ADS)

Zhang, Meng; Liang, Shujian; Hang, Xiaoming; Xiang, Yingxia; Cheng, Zhenlong; Li, Wenjian; Shi, Jinming; Huang, Lei; Sun, Yeqing

2011-03-01

MicroRNAs (miRNAs) are a family of small non-coding RNAs, which play significant roles in regulating development and stress responses in plant. As an excellent model organism for studying the effects of environmental stress, rice has been used to assess the damage of the space radiation environment for decades. Heavy-ions radiation show higher relative biological effectiveness compared to other cosmic-rays radiation. To identify the specific miRNAs that underlie biological effects of heavy-ion radiation, the germinated seeds of rice were exposed to 1 Gy, 10 Gy and 20 Gy dose of 12C heavy-ion radiation, respectively. Analysis of phenotype indicated that 20 Gy dose of heavy-ion radiation was the semi-lethal dose of rice seedling. The microarray of μparaflo™ chip was employed to monitor the expression profiles of miRNAs in rice (Oryza sativa) under 20 Gy dose of radiation stress. miR164a, miR164c, miR164d and miR156a-j were identified as heavy-ion radiation-induced miRNAs. miR164 and miR156 family were increased in all three exposed samples by using quantitative real-time PCR (qRT-RCP). As targets of miR156 and miR164, SQUAMOSA PROMOTER BINDING-LIKE (SPL) transcription factors and NAM/ATAF/CUC (NAC) transcription factors expression were down-regulated correlating with an up-regulated level of the regulated miRNAs. Since SPL transcription factors and NAC transcription factors regulated growth and development of plant, we used 2-dimension electrophoresis (2-DE) gel to analyze changes of functional proteins in 20 Gy exposed samples. It was evident that both the height and survival rates of seedlings were markedly decreased. The abundance of some developmentally regulated proteins was also changed. To our knowledge, this study is the first to report heavy-ion radiation stress responsive miRNAs in plant. Moreover, our findings are important to understand the molecular mechanism of space biology.

CisMiner: Genome-Wide In-Silico Cis-Regulatory Module Prediction by Fuzzy Itemset Mining

PubMed Central

Navarro, Carmen; Lopez, Francisco J.; Cano, Carlos; Garcia-Alcalde, Fernando; Blanco, Armando

2014-01-01

Eukaryotic gene control regions are known to be spread throughout non-coding DNA sequences which may appear distant from the gene promoter. Transcription factors are proteins that coordinately bind to these regions at transcription factor binding sites to regulate gene expression. Several tools allow to detect significant co-occurrences of closely located binding sites (cis-regulatory modules, CRMs). However, these tools present at least one of the following limitations: 1) scope limited to promoter or conserved regions of the genome; 2) do not allow to identify combinations involving more than two motifs; 3) require prior information about target motifs. In this work we present CisMiner, a novel methodology to detect putative CRMs by means of a fuzzy itemset mining approach able to operate at genome-wide scale. CisMiner allows to perform a blind search of CRMs without any prior information about target CRMs nor limitation in the number of motifs. CisMiner tackles the combinatorial complexity of genome-wide cis-regulatory module extraction using a natural representation of motif combinations as itemsets and applying the Top-Down Fuzzy Frequent- Pattern Tree algorithm to identify significant itemsets. Fuzzy technology allows CisMiner to better handle the imprecision and noise inherent to regulatory processes. Results obtained for a set of well-known binding sites in the S. cerevisiae genome show that our method yields highly reliable predictions. Furthermore, CisMiner was also applied to putative in-silico predicted transcription factor binding sites to identify significant combinations in S. cerevisiae and D. melanogaster, proving that our approach can be further applied genome-wide to more complex genomes. CisMiner is freely accesible at: http://genome2.ugr.es/cisminer. CisMiner can be queried for the results presented in this work and can also perform a customized cis-regulatory module prediction on a query set of transcription factor binding sites provided by the user. PMID:25268582

The role of Transposable Elements in shaping the combinatorial interaction of Transcription Factors

PubMed Central

2012-01-01

Background In the last few years several studies have shown that Transposable Elements (TEs) in the human genome are significantly associated with Transcription Factor Binding Sites (TFBSs) and that in several cases their expansion within the genome led to a substantial rewiring of the regulatory network. Another important feature of the regulatory network which has been thoroughly studied is the combinatorial organization of transcriptional regulation. In this paper we combine these two observations and suggest that TEs, besides rewiring the network, also played a central role in the evolution of particular patterns of combinatorial gene regulation. Results To address this issue we searched for TEs overlapping Estrogen Receptor α (ERα) binding peaks in two publicly available ChIP-seq datasets from the MCF7 cell line corresponding to different modalities of exposure to estrogen. We found a remarkable enrichment of a few specific classes of Transposons. Among these a prominent role was played by MIR (Mammalian Interspersed Repeats) transposons. These TEs underwent a dramatic expansion at the beginning of the mammalian radiation and then stabilized. We conjecture that the special affinity of ERα for the MIR class of TEs could be at the origin of the important role assumed by ERα in Mammalians. We then searched for TFBSs within the TEs overlapping ChIP-seq peaks. We found a strong enrichment of a few precise combinations of TFBS. In several cases the corresponding Transcription Factors (TFs) were known cofactors of ERα, thus supporting the idea of a co-regulatory role of TFBS within the same TE. Moreover, most of these correlations turned out to be strictly associated to specific classes of TEs thus suggesting the presence of a well-defined "transposon code" within the regulatory network. Conclusions In this work we tried to shed light into the role of Transposable Elements (TEs) in shaping the regulatory network of higher eukaryotes. To test this idea we focused on a particular transcription factor: the Estrogen Receptor α (ERα) and we found that ERα preferentially targets a well defined set of TEs and that these TEs host combinations of transcriptional regulators involving several of known co-regulators of ERα. Moreover, a significant number of these TEs turned out to be conserved between human and mouse and located in the vicinity (and thus candidate to be regulators) of important estrogen-related genes. PMID:22897927

Translational profiling of B cells infected with the Epstein-Barr virus reveals 5' leader ribosome recruitment through upstream open reading frames.

PubMed

Bencun, Maja; Klinke, Olaf; Hotz-Wagenblatt, Agnes; Klaus, Severina; Tsai, Ming-Han; Poirey, Remy; Delecluse, Henri-Jacques

2018-04-06

The Epstein-Barr virus (EBV) genome encodes several hundred transcripts. We have used ribosome profiling to characterize viral translation in infected cells and map new translation initiation sites. We show here that EBV transcripts are translated with highly variable efficiency, owing to variable transcription and translation rates, variable ribosome recruitment to the leader region and coverage by monosomes versus polysomes. Some transcripts were hardly translated, others mainly carried monosomes, showed ribosome accumulation in leader regions and most likely represent non-coding RNAs. A similar process was visible for a subset of lytic genes including the key transactivators BZLF1 and BRLF1 in cells infected with weakly replicating EBV strains. This suggests that ribosome trapping, particularly in the leader region, represents a new checkpoint for the repression of lytic replication. We could identify 25 upstream open reading frames (uORFs) located upstream of coding transcripts that displayed 5' leader ribosome trapping, six of which were located in the leader region shared by many latent transcripts. These uORFs repressed viral translation and are likely to play an important role in the regulation of EBV translation.

Transcriptional Changes in the Transition from Vegetative Cells to Asexual Development in the Model Fungus Aspergillus nidulans

PubMed Central

Garzia, Aitor; Etxebeste, Oier; Rodríguez-Romero, Julio; Fischer, Reinhard; Espeso, Eduardo A.

2013-01-01

Morphogenesis encompasses programmed changes in gene expression that lead to the development of specialized cell types. In the model fungus Aspergillus nidulans, asexual development involves the formation of characteristic cell types, collectively known as the conidiophore. With the aim of determining the transcriptional changes that occur upon induction of asexual development, we have applied massive mRNA sequencing to compare the expression pattern of 19-h-old submerged vegetative cells (hyphae) with that of similar hyphae after exposure to the air for 5 h. We found that the expression of 2,222 (20.3%) of the predicted 10,943 A. nidulans transcripts was significantly modified after air exposure, 2,035 being downregulated and 187 upregulated. The activation during this transition of genes that belong specifically to the asexual developmental pathway was confirmed. Another remarkable quantitative change occurred in the expression of genes involved in carbon or nitrogen primary metabolism. Genes participating in polar growth or sexual development were transcriptionally repressed, as were those belonging to the HogA/SakA stress response mitogen-activated protein (MAP) kinase pathway. We also identified significant expression changes in several genes purportedly involved in redox balance, transmembrane transport, secondary metabolite production, or transcriptional regulation, mainly binuclear-zinc cluster transcription factors. Genes coding for these four activities were usually grouped in metabolic clusters, which may bring regulatory implications for the induction of asexual development. These results provide a blueprint for further stage-specific gene expression studies during conidiophore development. PMID:23264642

Increased levels of B1 and B2 SINE transcripts in mouse fibroblast cells due to minute virus of mice infection.

PubMed

Williams, Warren P; Tamburic, Lillian; Astell, Caroline R

2004-10-01

Minute virus of mice (MVM), an autonomous parvovirus, has served as a model for understanding parvovirus infection including host cell response to infection. In this paper, we report the effect of MVM infection on host cell gene expression in mouse fibroblast cells (LA9 cells), analyzed by differential display. Somewhat surprisingly, our data reveal that few cellular protein-coding genes appear to be up- or downregulated and identify the murine B1 and B2 short interspersed element (SINE) transcripts as being increased upon MVM infection. Primer extension assays confirm the effect of MVM infection on SINE expression and demonstrate that both SINEs are upregulated in a roughly linear fashion throughout MVM infection. They also demonstrate that the SINE response was due to RNA polymerase III transcription and not contaminating DNA or RNA polymerase II transcription. Furthermore, expression of MVM NS1, the major nonstructural protein, by transient transfection also leads to an increase in both murine SINEs. We believe this is the first time that the B1 and B2 SINEs have been shown to be altered by viral infection and the first time parvovirus infection has been shown to increase SINE expression. The increase in SINE transcripts caused by MVM infection does not appear to be due to an increase in either of the basal transcription factors TFIIIC110 or 220, in contrast to that which has been shown for other viruses.

Expression of a non-coding RNA in ectromelia virus is required for normal plaque formation.

PubMed

Esteban, David J; Upton, Chris; Bartow-McKenney, Casey; Buller, R Mark L; Chen, Nanhai G; Schriewer, Jill; Lefkowitz, Elliot J; Wang, Chunlin

2014-02-01

Poxviruses are dsDNA viruses with large genomes. Many genes in the genome remain uncharacterized, and recent studies have demonstrated that the poxvirus transcriptome includes numerous so-called anomalous transcripts not associated with open reading frames. Here, we characterize the expression and role of an apparently non-coding RNA in orthopoxviruses, which we call viral hairpin RNA (vhRNA). Using a bioinformatics approach, we predicted expression of a transcript not associated with an open reading frame that is likely to form a stem-loop structure due to the presence of a 21 nt palindromic sequence. Expression of the transcript as early as 2 h post-infection was confirmed by northern blot and analysis of publicly available vaccinia virus infected cell transcriptomes. The transcription start site was determined by RACE PCE and transcriptome analysis, and early and late promoter sequences were identified. Finally, to test the function of the transcript we generated an ectromelia virus knockout, which failed to form plaques in cell culture. The important role of the transcript in viral replication was further demonstrated using siRNA. Although the function of the transcript remains unknown, our work contributes to evidence of an increasingly complex poxvirus transcriptome, suggesting that transcripts such as vhRNA not associated with an annotated open reading frame can play an important role in viral replication.

Global isoform-specific transcript alterations and deregulated networks in clear cell renal cell carcinoma

PubMed Central

Hamilton, Michael J.; Girke, Thomas; Martinez, Ernest

2018-01-01

Extensive genome-wide analyses of deregulated gene expression have now been performed for many types of cancer. However, most studies have focused on deregulation at the gene-level, which may overlook the alterations of specific transcripts for a given gene. Clear cell renal cell carcinoma (ccRCC) is one of the best-characterized and most pervasive renal cancers, and ccRCCs are well-documented to have aberrant RNA processing. In the present study, we examine the extent of aberrant isoform-specific RNA expression by reporting a comprehensive transcript-level analysis, using the new kallisto-sleuth-RATs pipeline, investigating coding and non-coding differential transcript expression in ccRCC. We analyzed 50 ccRCC tumors and their matched normal samples from The Cancer Genome Altas datasets. We identified 7,339 differentially expressed transcripts and 94 genes exhibiting differential transcript isoform usage in ccRCC. Additionally, transcript-level coexpression network analyses identified vasculature development and the tricarboxylic acid cycle as the most significantly deregulated networks correlating with ccRCC progression. These analyses uncovered several uncharacterized transcripts, including lncRNAs FGD5-AS1 and AL035661.1, as potential regulators of the tricarboxylic acid cycle associated with ccRCC progression. As ccRCC still presents treatment challenges, our results provide a new resource of potential therapeutics targets and highlight the importance of exploring alternative methodologies in transcriptome-wide studies.

From policy to practice: implementation of physical activity and food policies in schools

PubMed Central

2013-01-01

Purpose Public policies targeting the school setting are increasingly being used to address childhood obesity; however, their effectiveness depends on their implementation. This study explores the factors which impeded or facilitated the implementation of publicly mandated school-based physical activity and nutrition guidelines in the province of British Columbia (BC), Canada. Methods Semi-structured interviews were conducted with 50 school informants (17 principals - 33 teacher/school informants) to examine the factors associated with the implementation of the mandated Daily Physical Activity (DPA) and Food and Beverage Sales in Schools (FBSS) guidelines. Coding used a constructivist grounded theory approach. The first five transcripts and every fifth transcript thereafter were coded by two independent coders with discrepancies reconciled by a third coder. Data was coded and analysed in the NVivo 9 software. Concept maps were developed and current theoretical perspectives were integrated in the later stages of analysis. Results The Diffusion of Innovations Model provided an organizing framework to present emergent themes. With the exception of triability (not relevant in the context of mandated guidelines/policies), the key attributes of the Diffusion of Innovations Model (relative advantage, compatibility, complexity, and observability) provided a robust framework for understanding themes associated with implementation of mandated guidelines. Specifically, implementation of the DPA and FBSS guidelines was facilitated by perceptions that they: were relatively advantageous compared to status quo; were compatible with school mandates and teaching philosophies; had observable positive impacts and impeded when perceived as complex to understand and implement. In addition, a number of contextual factors including availability of resources facilitated implementation. Conclusions The enactment of mandated policies/guidelines for schools is considered an essential step in improving physical activity and healthy eating. However, policy makers need to: monitor whether schools are able to implement the guidelines, support schools struggling with implementation, and document the impact of the guidelines on students’ behaviors. To facilitate the implementation of mandated guidelines/policies, the Diffusion of Innovations Model provides an organizational framework for planning interventions. Changing the school environment is a process which cannot be undertaken solely by passive means as we know that such approaches have not resulted in adequate implementation. PMID:23731803

From policy to practice: implementation of physical activity and food policies in schools.

PubMed

Mâsse, Louise C; Naiman, Daniel; Naylor, Patti-Jean

2013-06-03

Public policies targeting the school setting are increasingly being used to address childhood obesity; however, their effectiveness depends on their implementation. This study explores the factors which impeded or facilitated the implementation of publicly mandated school-based physical activity and nutrition guidelines in the province of British Columbia (BC), Canada. Semi-structured interviews were conducted with 50 school informants (17 principals - 33 teacher/school informants) to examine the factors associated with the implementation of the mandated Daily Physical Activity (DPA) and Food and Beverage Sales in Schools (FBSS) guidelines. Coding used a constructivist grounded theory approach. The first five transcripts and every fifth transcript thereafter were coded by two independent coders with discrepancies reconciled by a third coder. Data was coded and analysed in the NVivo 9 software. Concept maps were developed and current theoretical perspectives were integrated in the later stages of analysis. The Diffusion of Innovations Model provided an organizing framework to present emergent themes. With the exception of triability (not relevant in the context of mandated guidelines/policies), the key attributes of the Diffusion of Innovations Model (relative advantage, compatibility, complexity, and observability) provided a robust framework for understanding themes associated with implementation of mandated guidelines. Specifically, implementation of the DPA and FBSS guidelines was facilitated by perceptions that they: were relatively advantageous compared to status quo; were compatible with school mandates and teaching philosophies; had observable positive impacts and impeded when perceived as complex to understand and implement. In addition, a number of contextual factors including availability of resources facilitated implementation. The enactment of mandated policies/guidelines for schools is considered an essential step in improving physical activity and healthy eating. However, policy makers need to: monitor whether schools are able to implement the guidelines, support schools struggling with implementation, and document the impact of the guidelines on students' behaviors. To facilitate the implementation of mandated guidelines/policies, the Diffusion of Innovations Model provides an organizational framework for planning interventions. Changing the school environment is a process which cannot be undertaken solely by passive means as we know that such approaches have not resulted in adequate implementation.

The agents of natural genome editing.

PubMed

Witzany, Guenther

2011-06-01

The DNA serves as a stable information storage medium and every protein which is needed by the cell is produced from this blueprint via an RNA intermediate code. More recently it was found that an abundance of various RNA elements cooperate in a variety of steps and substeps as regulatory and catalytic units with multiple competencies to act on RNA transcripts. Natural genome editing on one side is the competent agent-driven generation and integration of meaningful DNA nucleotide sequences into pre-existing genomic content arrangements, and the ability to (re-)combine and (re-)regulate them according to context-dependent (i.e. adaptational) purposes of the host organism. Natural genome editing on the other side designates the integration of all RNA activities acting on RNA transcripts without altering DNA-encoded genes. If we take the genetic code seriously as a natural code, there must be agents that are competent to act on this code because no natural code codes itself as no natural language speaks itself. As code editing agents, viral and subviral agents have been suggested because there are several indicators that demonstrate viruses competent in both RNA and DNA natural genome editing.

A transcription factor links growth rate and metabolism in the hypersaline adapted archaeon Halobacterium salinarum.

PubMed

Todor, Horia; Dulmage, Keely; Gillum, Nicholas; Bain, James R; Muehlbauer, Michael J; Schmid, Amy K

2014-09-01

Co-ordinating metabolism and growth is a key challenge for all organisms. Despite fluctuating environments, cells must produce the same metabolic outputs to thrive. The mechanisms underlying this 'growth homeostasis' are known in bacteria and eukaryotes, but remain unexplored in archaea. In the model archaeon Halobacterium salinarum, the transcription factor TrmB regulates enzyme-coding genes in diverse metabolic pathways in response to glucose. However, H. salinarum is thought not to catabolize glucose. To resolve this discrepancy, we demonstrate that TrmB regulates the gluconeogenic production of sugars incorporated into the cell surface S-layer glycoprotein. Additionally, we show that TrmB-DNA binding correlates with instantaneous growth rate, likely because S-layer glycosylation is proportional to growth. This suggests that TrmB transduces a growth rate signal to co-regulated metabolic pathways including amino acid, purine, and cobalamin biosynthesis. Remarkably, the topology and function of this growth homeostatic network appear conserved across domains despite extensive alterations in protein components. © 2014 The Authors. Molecular Microbiology published by John Wiley & Sons Ltd.

Transcriptional landscapes of Axolotl (Ambystoma mexicanum).

PubMed

Caballero-Pérez, Juan; Espinal-Centeno, Annie; Falcon, Francisco; García-Ortega, Luis F; Curiel-Quesada, Everardo; Cruz-Hernández, Andrés; Bako, Laszlo; Chen, Xuemei; Martínez, Octavio; Alberto Arteaga-Vázquez, Mario; Herrera-Estrella, Luis; Cruz-Ramírez, Alfredo

2018-01-15

The axolotl (Ambystoma mexicanum) is the vertebrate model system with the highest regeneration capacity. Experimental tools established over the past 100 years have been fundamental to start unraveling the cellular and molecular basis of tissue and limb regeneration. In the absence of a reference genome for the Axolotl, transcriptomic analysis become fundamental to understand the genetic basis of regeneration. Here we present one of the most diverse transcriptomic data sets for Axolotl by profiling coding and non-coding RNAs from diverse tissues. We reconstructed a population of 115,906 putative protein coding mRNAs as full ORFs (including isoforms). We also identified 352 conserved miRNAs and 297 novel putative mature miRNAs. Systematic enrichment analysis of gene expression allowed us to identify tissue-specific protein-coding transcripts. We also found putative novel and conserved microRNAs which potentially target mRNAs which are reported as important disease candidates in heart and liver. Copyright © 2017 Elsevier Inc. All rights reserved.

De Novo Transcriptome Analysis of Medicinally Important Plantago ovata Using RNA-Seq

PubMed Central

Kotwal, Shivanjali; Kaul, Sanjana; Sharma, Pooja; Gupta, Mehak; Shankar, Rama; Jain, Mukesh; Dhar, Manoj K.

2016-01-01

Plantago ovata is an economically and medicinally important plant of the family Plantaginaceae. It is used extensively for the production of seed husk for its application in pharmaceutical, food and cosmetic industries. In the present study, the transcriptome of P. ovata ovary was sequenced using Illumina Genome Analyzer platform to characterize the mucilage biosynthesis pathway in the plant. De novo assembly was carried out using Oases followed by velvet. A total of 46,955 non-redundant transcripts (≥100 bp) using ~29 million high-quality paired end reads were generated. Functional categorization of these transcripts revealed the presence of several genes involved in various biological processes like metabolic pathways, mucilage biosynthesis, biosynthesis of secondary metabolites and antioxidants. In addition, simple sequence-repeat motifs, non-coding RNAs and transcription factors were also identified. Expression profiling of some genes involved in mucilage biosynthetic pathway was performed in different tissues of P. ovata using Real time PCR analysis. The study has resulted in a valuable resource for further studies on gene expression, genomics and functional genomics in P. ovata. PMID:26943165

Identification of Regulatory Elements That Control PPARγ Expression in Adipocyte Progenitors

PubMed Central

Chou, Wen-Ling; Galmozzi, Andrea; Partida, David; Kwan, Kevin; Yeung, Hui; Su, Andrew I.; Saez, Enrique

2013-01-01

Adipose tissue renewal and obesity-driven expansion of fat cell number are dependent on proliferation and differentiation of adipose progenitors that reside in the vasculature that develops in coordination with adipose depots. The transcriptional events that regulate commitment of progenitors to the adipose lineage are poorly understood. Because expression of the nuclear receptor PPARγ defines the adipose lineage, isolation of elements that control PPARγ expression in adipose precursors may lead to discovery of transcriptional regulators of early adipocyte determination. Here, we describe the identification and validation in transgenic mice of 5 highly conserved non-coding sequences from the PPARγ locus that can drive expression of a reporter gene in a manner that recapitulates the tissue-specific pattern of PPARγ expression. Surprisingly, these 5 elements appear to control PPARγ expression in adipocyte precursors that are associated with the vasculature of adipose depots, but not in mature adipocytes. Characterization of these five PPARγ regulatory sequences may enable isolation of the transcription factors that bind these cis elements and provide insight into the molecular regulation of adipose tissue expansion in normal and pathological states. PMID:24009687

BAG3 promotes stem cell-like phenotype in breast cancer by upregulation of CXCR4 via interaction with its transcript.

PubMed

Liu, Bao-Qin; Zhang, Song; Li, Si; An, Ming-Xin; Li, Chao; Yan, Jing; Wang, Jia-Mei; Wang, Hua-Qin

2017-07-13

BAG3 is an evolutionarily conserved co-chaperone expressed at high levels and has a prosurvival role in many tumor types. The current study reported that BAG3 was induced under specific floating culture conditions that enrich breast cancer stem cell (BCSC)-like cells in spheres. Ectopic BAG3 overexpression increased CD44 + /CD24 - CSC subpopulations, first-generation and second-generation mammosphere formation, indicating that BAG3 promotes CSC self-renewal and maintenance in breast cancer. We further demonstrated that mechanically, BAG3 upregulated CXCR4 expression at the post-transcriptional level. Further studies showed that BAG3 interacted with CXCR4 mRNA and promoted its expression via its coding and 3'-untranslational regions. BAG3 was also found to be positively correlated with CXCR4 expression and unfavorable prognosis in patients with breast cancer. Taken together, our data demonstrate that BAG3 promotes BCSC-like phenotype through CXCR4 via interaction with its transcript. Therefore, this study establishes BAG3 as a potential adverse prognostic factor and a therapeutic target of breast cancer.

Transcriptome-wide identification of RNA-binding protein and microRNA target sites by PAR-CLIP

PubMed Central

Hafner, Markus; Landthaler, Markus; Burger, Lukas; Khorshid, Mohsen; Hausser, Jean; Berninger, Philipp; Rothballer, Andrea; Ascano, Manuel; Jungkamp, Anna-Carina; Munschauer, Mathias; Ulrich, Alexander; Wardle, Greg S.; Dewell, Scott; Zavolan, Mihaela; Tuschl, Thomas

2010-01-01

Summary RNA transcripts are subject to post-transcriptional gene regulation involving hundreds of RNA-binding proteins (RBPs) and microRNA-containing ribonucleoprotein complexes (miRNPs) expressed in a cell-type dependent fashion. We developed a cell-based crosslinking approach to determine at high resolution and transcriptome-wide the binding sites of cellular RBPs and miRNPs. The crosslinked sites are revealed by thymidine to cytidine transitions in the cDNAs prepared from immunopurified RNPs of 4-thiouridine-treated cells. We determined the binding sites and regulatory consequences for several intensely studied RBPs and miRNPs, including PUM2, QKI, IGF2BP1-3, AGO/EIF2C1-4 and TNRC6A-C. Our study revealed that these factors bind thousands of sites containing defined sequence motifs and have distinct preferences for exonic versus intronic or coding versus untranslated transcript regions. The precise mapping of binding sites across the transcriptome will be critical to the interpretation of the rapidly emerging data on genetic variation between individuals and how these variations contribute to complex genetic diseases. PMID:20371350

Dynamic Metabolite Profiling in an Archaeon Connects Transcriptional Regulation to Metabolic Consequences.

PubMed

Todor, Horia; Gooding, Jessica; Ilkayeva, Olga R; Schmid, Amy K

2015-01-01

Previous work demonstrated that the TrmB transcription factor is responsible for regulating the expression of many enzyme-coding genes in the hypersaline-adapted archaeon Halobacterium salinarum via a direct interaction with a cis-regulatory sequence in their promoters. This interaction is abolished in the presence of glucose. Although much is known about the effects of TrmB at the transcriptional level, it remains unclear whether and to what extent changes in mRNA levels directly affect metabolite levels. In order to address this question, here we performed a high-resolution metabolite profiling time course during a change in nutrients using a combination of targeted and untargeted methods in wild-type and ΔtrmB strain backgrounds. We found that TrmB-mediated transcriptional changes resulted in widespread and significant changes to metabolite levels across the metabolic network. Additionally, the pattern of growth complementation using various purines suggests that the mis-regulation of gluconeogenesis in the ΔtrmB mutant strain in the absence of glucose results in low phosphoribosylpyrophosphate (PRPP) levels. We confirmed these low PRPP levels using a quantitative mass spectrometric technique and found that they are associated with a metabolic block in de novo purine synthesis, which is partially responsible for the growth defect of the ΔtrmB mutant strain in the absence of glucose. In conclusion, we show how transcriptional regulation of metabolism affects metabolite levels and ultimately, phenotypes.

Extensive Evolutionary Changes in Regulatory Element Activity during Human Origins Are Associated with Altered Gene Expression and Positive Selection

PubMed Central

Fedrigo, Olivier; Babbitt, Courtney C.; Wortham, Matthew; Tewari, Alok K.; London, Darin; Song, Lingyun; Lee, Bum-Kyu; Iyer, Vishwanath R.; Parker, Stephen C. J.; Margulies, Elliott H.; Wray, Gregory A.; Furey, Terrence S.; Crawford, Gregory E.

2012-01-01

Understanding the molecular basis for phenotypic differences between humans and other primates remains an outstanding challenge. Mutations in non-coding regulatory DNA that alter gene expression have been hypothesized as a key driver of these phenotypic differences. This has been supported by differential gene expression analyses in general, but not by the identification of specific regulatory elements responsible for changes in transcription and phenotype. To identify the genetic source of regulatory differences, we mapped DNaseI hypersensitive (DHS) sites, which mark all types of active gene regulatory elements, genome-wide in the same cell type isolated from human, chimpanzee, and macaque. Most DHS sites were conserved among all three species, as expected based on their central role in regulating transcription. However, we found evidence that several hundred DHS sites were gained or lost on the lineages leading to modern human and chimpanzee. Species-specific DHS site gains are enriched near differentially expressed genes, are positively correlated with increased transcription, show evidence of branch-specific positive selection, and overlap with active chromatin marks. Species-specific sequence differences in transcription factor motifs found within these DHS sites are linked with species-specific changes in chromatin accessibility. Together, these indicate that the regulatory elements identified here are genetic contributors to transcriptional and phenotypic differences among primate species. PMID:22761590

Gene network reconstruction from transcriptional dynamics under kinetic model uncertainty: a case for the second derivative

PubMed Central

Bickel, David R.; Montazeri, Zahra; Hsieh, Pei-Chun; Beatty, Mary; Lawit, Shai J.; Bate, Nicholas J.

2009-01-01

Motivation: Measurements of gene expression over time enable the reconstruction of transcriptional networks. However, Bayesian networks and many other current reconstruction methods rely on assumptions that conflict with the differential equations that describe transcriptional kinetics. Practical approximations of kinetic models would enable inferring causal relationships between genes from expression data of microarray, tag-based and conventional platforms, but conclusions are sensitive to the assumptions made. Results: The representation of a sufficiently large portion of genome enables computation of an upper bound on how much confidence one may place in influences between genes on the basis of expression data. Information about which genes encode transcription factors is not necessary but may be incorporated if available. The methodology is generalized to cover cases in which expression measurements are missing for many of the genes that might control the transcription of the genes of interest. The assumption that the gene expression level is roughly proportional to the rate of translation led to better empirical performance than did either the assumption that the gene expression level is roughly proportional to the protein level or the Bayesian model average of both assumptions. Availability: http://www.oisb.ca points to R code implementing the methods (R Development Core Team 2004). Contact: dbickel@uottawa.ca Supplementary information: http://www.davidbickel.com PMID:19218351

Transcriptome discovery in non-model wild fish species for the development of quantitative transcript abundance assays

USGS Publications Warehouse

Hahn, Cassidy M.; Iwanowicz, Luke R.; Cornman, Robert S.; Mazik, Patricia M.; Blazer, Vicki S.

2016-01-01

Environmental studies increasingly identify the presence of both contaminants of emerging concern (CECs) and legacy contaminants in aquatic environments; however, the biological effects of these compounds on resident fishes remain largely unknown. High throughput methodologies were employed to establish partial transcriptomes for three wild-caught, non-model fish species; smallmouth bass (Micropterus dolomieu), white sucker (Catostomus commersonii) and brown bullhead (Ameiurus nebulosus). Sequences from these transcriptome databases were utilized in the development of a custom nCounter CodeSet that allowed for direct multiplexed measurement of 50 transcript abundance endpoints in liver tissue. Sequence information was also utilized in the development of quantitative real-time PCR (qPCR) primers. Cross-species hybridization allowed the smallmouth bass nCounter CodeSet to be used for quantitative transcript abundance analysis of an additional non-model species, largemouth bass (Micropterus salmoides). We validated the nCounter analysis data system with qPCR for a subset of genes and confirmed concordant results. Changes in transcript abundance biomarkers between sexes and seasons were evaluated to provide baseline data on transcript modulation for each species of interest.

The role of alternative Polyadenylation in regulation of rhythmic gene expression.

PubMed

Ptitsyna, Natalia; Boughorbel, Sabri; El Anbari, Mohammed; Ptitsyn, Andrey

2017-08-04

Alternative transcription is common in eukaryotic cells and plays important role in regulation of cellular processes. Alternative polyadenylation results from ambiguous PolyA signals in 3' untranslated region (UTR) of a gene. Such alternative transcripts share the same coding part, but differ by a stretch of UTR that may contain important functional sites. The methodoogy of this study is based on mathematical modeling, analytical solution, and subsequent validation by datamining in multiple independent experimental data from previously published studies. In this study we propose a mathematical model that describes the population dynamics of alternatively polyadenylated transcripts in conjunction with rhythmic expression such as transcription oscillation driven by circadian or metabolic oscillators. Analysis of the model shows that alternative transcripts with different turnover rates acquire a phase shift if the transcript decay rate is different. Difference in decay rate is one of the consequences of alternative polyadenylation. Phase shift can reach values equal to half the period of oscillation, which makes alternative transcripts oscillate in abundance in counter-phase to each other. Since counter-phased transcripts share the coding part, the rate of translation becomes constant. We have analyzed a few data sets collected in circadian timeline for the occurrence of transcript behavior that fits the mathematical model. Alternative transcripts with different turnover rate create the effect of rectifier. This "molecular diode" moderates or completely eliminates oscillation of individual transcripts and stabilizes overall protein production rate. In our observation this phenomenon is very common in different tissues in plants, mice, and humans. The occurrence of counter-phased alternative transcripts is also tissue-specific and affects functions of multiple biological pathways. Accounting for this mechanism is important for understanding the natural and engineering the synthetic cellular circuits.

Molecular characterization of Quercus suber MYB1, a transcription factor up-regulated in cork tissues.

PubMed

Almeida, Tânia; Menéndez, Esther; Capote, Tiago; Ribeiro, Teresa; Santos, Conceição; Gonçalves, Sónia

2013-01-15

The molecular processes associated with cork development in Quercus suber L. are poorly understood. A previous molecular approach identified a list of genes potentially important for cork formation and differentiation, providing a new basis for further molecular studies. This report is the first molecular characterization of one of these candidate genes, QsMYB1, coding for an R2R3-MYB transcription factor. The R2R3-MYB gene sub-family has been described as being involved in the phenylpropanoid and lignin pathways, both involved in cork biosynthesis. The results showed that the expression of QsMYB1 is putatively mediated by an alternative splicing (AS) mechanism that originates two different transcripts (QsMYB1.1 and QsMYB1.2), differing only in the 5'-untranslated region, due to retention of the first intron in one of the variants. Moreover, within the retained intron, a simple sequence repeat (SSR) was identified. The upstream regulatory region of QsMYB1 was extended by a genome walking approach, which allowed the identification of the putative gene promoter region. The relative expression pattern of QsMYB1 transcripts determined by reverse transcription quantitative polymerase chain reaction (RT-qPCR) revealed that both transcripts were up-regulated in cork tissues; the detected expression was several times higher in newly formed cork harvested from trees producing virgin, second or reproduction cork when compared with wood. Moreover, the expression analysis of QsMYB1 in several Q. suber organs showed very low expression in young branches and roots, whereas in leaves, immature acorns or male flowers, no expression was detected. These preliminary results suggest that QsMYB1 may be related to secondary growth and, in particular, with the cork biosynthesis process with a possible alternative splicing mechanism associated with its regulatory function. Copyright © 2012 Elsevier GmbH. All rights reserved.

Intrinsic disorder in transcription factors†

PubMed Central

Liu, Jiangang; Perumal, Narayanan B.; Oldfield, Christopher J.; Su, Eric W.; Uversky, Vladimir N.; Dunker, A. Keith

2008-01-01

Intrinsic disorder (ID) is highly abundant in eukaryotes, which reflect the greater need for disorder-associated signaling and transcriptional regulation in nucleated cells. Although several well-characterized examples of intrinsically disordered proteins in transcriptional regulation have been reported, no systematic analysis has been reported so far. To test for a general prevalence of intrinsic disorder in transcriptional regulation, we used the Predictor Of Natural Disorder Regions (PONDR) to analyze the abundance of intrinsic disorder in three transcription factor datasets and two control sets. This analysis revealed that from 94.13% to 82.63% of transcription factors posses extended regions of intrinsic disorder, relative to 54.51% and 18.64% of the proteins in two control datasets, which indicates the significant prevalence of intrinsic disorder in transcription factors. This propensity of transcription factors for intrinsic disorder was confirmed by cumulative distribution function analysis and charge-hydropathy plots. The amino acid composition analysis showed that all three transcription factor datasets were substantially depleted in order-promoting residues, and significantly enriched in disorder-promoting residues. Our analysis of the distribution of disorder within the transcription factor datasets revealed that: (a) The AT-hooks and basic regions of transcription factor DNA-binding domains are highly disordered; (b) The degree of disorder in transcription factor activation regions is much higher than that in DNA-binding domains; (c) The degree of disorder is significantly higher in eukaryotic transcription factors than in prokaryotic transcription factors; (d) The level of α-MoRFs (molecular recognition feature) prediction is much higher in transcription factors. Overall, our data reflected the fact that the eukaryotes with well-developed gene transcription machinery require transcription factor flexibility to be more efficient. PMID:16734424

Identification of differentially expressed small non-coding RNAs in the legume endosymbiont Sinorhizobium meliloti by comparative genomics

PubMed Central

del Val, Coral; Rivas, Elena; Torres-Quesada, Omar; Toro, Nicolás; Jiménez-Zurdo, José I

2007-01-01

Bacterial small non-coding RNAs (sRNAs) are being recognized as novel widespread regulators of gene expression in response to environmental signals. Here, we present the first search for sRNA-encoding genes in the nitrogen-fixing endosymbiont Sinorhizobium meliloti, performed by a genome-wide computational analysis of its intergenic regions. Comparative sequence data from eight related α-proteobacteria were obtained, and the interspecies pairwise alignments were scored with the programs eQRNA and RNAz as complementary predictive tools to identify conserved and stable secondary structures corresponding to putative non-coding RNAs. Northern experiments confirmed that eight of the predicted loci, selected among the original 32 candidates as most probable sRNA genes, expressed small transcripts. This result supports the combined use of eQRNA and RNAz as a robust strategy to identify novel sRNAs in bacteria. Furthermore, seven of the transcripts accumulated differentially in free-living and symbiotic conditions. Experimental mapping of the 5′-ends of the detected transcripts revealed that their encoding genes are organized in autonomous transcription units with recognizable promoter and, in most cases, termination signatures. These findings suggest novel regulatory functions for sRNAs related to the interactions of α-proteobacteria with their eukaryotic hosts. PMID:17971083

Parsimony and complexity: Cell fate assignment in the developing Drosophila eye.

PubMed

Mavromatakis, Yannis Emmanuel; Tomlinson, Andrew

2017-07-03

The specification of the R7 photoreceptor in the Drosophila eye has become a classic model for understanding how cell fates are assigned in developing systems. R7 is derived from a group of cells that also gives rise to the R1/6 photoreceptor class and the non-photoreceptor cone cells. Our studies examine the signals and cellular information that direct each of these cell types. The cell fates are directed by the combined actions of the Receptor Tyrosine Kinase (RTK) and Notch (N) signaling pathways. The RTK pathway acts to remove the transcription factor Tramtrack (Ttk) which represses the photoreceptor fate. If a cell receives an RTK signal sufficient to remove Ttk then the photoreceptor fate is specified; if not, the cone cell fate results. If Ttk is removed from a cell and its N activity is high then it is specified as an R7, but if its N activity is low then it becomes an R1/6 class photoreceptor. Thus, a remarkably simple molecular code underlies the specification of the fates: 1. Ttk degraded or not: 2. N activity high or low. In the R1/6 and cone cell precursors the molecular codes are achieved with relative simplicity but in the R7 precursor, manifold interactions occur between the RTK and N pathways, and to-date we have identified 4 distinct roles played by N in R7 fate specification. In this review we detail this molecular complexity, and describe how the RTK/N pathway crosstalk eventually leads to the simple molecular code of Tramtrack removed and N activity high. Furthermore, we describe the role played by the transcription factor Lozenge (Lz) in directing retinal precursor fates, and how the RTK/N signals specify different retinal cell types depending on the presence or absence of Lz.

Molecular Evolution of the Non-Coding Eosinophil Granule Ontogeny Transcript

PubMed Central

Rose, Dominic; Stadler, Peter F.

2011-01-01

Eukaryotic genomes are pervasively transcribed. A large fraction of the transcriptional output consists of long, mRNA-like, non-protein-coding transcripts (mlncRNAs). The evolutionary history of mlncRNAs is still largely uncharted territory. In this contribution, we explore in detail the evolutionary traces of the eosinophil granule ontogeny transcript (EGOT), an experimentally confirmed representative of an abundant class of totally intronic non-coding transcripts (TINs). EGOT is located antisense to an intron of the ITPR1 gene. We computationally identify putative EGOT orthologs in the genomes of 32 different amniotes, including orthologs from primates, rodents, ungulates, carnivores, afrotherians, and xenarthrans, as well as putative candidates from basal amniotes, such as opossum or platypus. We investigate the EGOT gene phylogeny, analyze patterns of sequence conservation, and the evolutionary conservation of the EGOT gene structure. We show that EGO-B, the spliced isoform, may be present throughout the placental mammals, but most likely dates back even further. We demonstrate here for the first time that the whole EGOT locus is highly structured, containing several evolutionary conserved, and thermodynamic stable secondary structures. Our analyses allow us to postulate novel functional roles of a hitherto poorly understood region at the intron of EGO-B which is highly conserved at the sequence level. The region contains a novel ITPR1 exon and also conserved RNA secondary structures together with a conserved TATA-like element, which putatively acts as a promoter of an independent regulatory element. PMID:22303364

A comprehensive catalog of human KRAB-associated zinc finger genes: Insights into the evolutionary history of a large family of transcriptional repressors

PubMed Central

Huntley, Stuart; Baggott, Daniel M.; Hamilton, Aaron T.; Tran-Gyamfi, Mary; Yang, Shan; Kim, Joomyeong; Gordon, Laurie; Branscomb, Elbert; Stubbs, Lisa

2006-01-01

Krüppel-type zinc finger (ZNF) motifs are prevalent components of transcription factor proteins in all eukaryotes. KRAB-ZNF proteins, in which a potent repressor domain is attached to a tandem array of DNA-binding zinc-finger motifs, are specific to tetrapod vertebrates and represent the largest class of ZNF proteins in mammals. To define the full repertoire of human KRAB-ZNF proteins, we searched the genome sequence for key motifs and then constructed and manually curated gene models incorporating those sequences. The resulting gene catalog contains 423 KRAB-ZNF protein-coding loci, yielding alternative transcripts that altogether predict at least 742 structurally distinct proteins. Active rounds of segmental duplication, involving single genes or larger regions and including both tandem and distributed duplication events, have driven the expansion of this mammalian gene family. Comparisons between the human genes and ZNF loci mined from the draft mouse, dog, and chimpanzee genomes not only identified 103 KRAB-ZNF genes that are conserved in mammals but also highlighted a substantial level of lineage-specific change; at least 136 KRAB-ZNF coding genes are primate specific, including many recent duplicates. KRAB-ZNF genes are widely expressed and clustered genes are typically not coregulated, indicating that paralogs have evolved to fill roles in many different biological processes. To facilitate further study, we have developed a Web-based public resource with access to gene models, sequences, and other data, including visualization tools to provide genomic context and interaction with other public data sets. PMID:16606702

The involvement of hypoxia-inducible transcription factor-1-dependent pathway in nickel carcinogenesis.

PubMed

Salnikow, Konstantin; Davidson, Todd; Zhang, Qunwei; Chen, Lung Chi; Su, Weichen; Costa, Max

2003-07-01

Nickel is a potent environmental pollutant in industrial countries. Because nickel compounds are carcinogenic, exposure to nickel represents a serious hazard to human health. The understanding of how nickel exerts its toxic and carcinogenic effects at a molecular level may be important in risk assessment, as well as in the treatment and prevention of occupational diseases. Previously, using human and rodent cells in vitro, we showed that hypoxia-inducible signaling pathway was activated by carcinogenic nickel compounds. Acute exposure to nickel resulted in the accumulation of hypoxia-inducible transcription factor (HIF)-1, which strongly activated hypoxia-inducible genes, including the recently discovered tumor marker NDRG1 (Cap43). To further identify HIF-1-dependent nickel-inducible genes and to understand the role of the HIF-dependent signaling pathway in nickel-induced transformation, we used the Affymetrix GeneChip to compare the gene expression profiles in wild-type cells or in cells from HIF-1 alpha knockout mouse embryos exposed to nickel chloride. As expected, when we examined 12,000 genes for expression changes, we found that genes coding for glycolytic enzymes and glucose transporters, known to be regulated by HIF-1 transcription factor, were induced by nickel only in HIF-1 alpha-proficient cells. In addition, we found a number of other hypoxia-inducible genes up-regulated by nickel in a HIF-dependent manner including BCL-2-binding protein Nip3, EGLN1, hypoxia-inducible gene 1 (HIG1), and prolyl 4-hydroxylase. Additionally, we found a number of genes induced by nickel in a HIF-independent manner, suggesting that Ni activated other signaling pathways besides HIF-1. Finally, we found that in HIF-1 alpha knockout cells, nickel strongly induced the expression of the whole group of genes that were not expressed in the presence of HIF-1. Because the majority of modulated genes were induced or suppressed by nickel in a HIF-1-dependent manner, we elucidated the role of HIF-1 transcription factor in cell transformation. In HIF-1 alpha-proficient cells, nickel exposure increased soft agar growth, whereas it decreased soft agar growth in HIF-1 alpha-deficient cells. We hypothesize that the induction of HIF-1 transcription factor by nickel may be important during the nickel-induced carcinogenic process.

Long Non-Coding RNA Malat1 Regulates Angiogenesis in Hindlimb Ischemia.

PubMed

Zhang, Xuejing; Tang, Xuelian; Hamblin, Milton H; Yin, Ke-Jie

2018-06-11

Angiogenesis is a complex process that depends on the delicate regulation of gene expression. Dysregulation of transcription during angiogenesis often leads to various human diseases. Emerging evidence has recently begun to show that long non-coding RNAs (lncRNAs) may mediate angiogenesis in both physiological and pathological conditions; concurrently, underlying molecular mechanisms are largely unexplored. Previously, our lab identified metastasis associates lung adenocarcinoma transcript 1 ( Malat1 ) as an oxygen-glucose deprivation (OGD)-responsive endothelial lncRNA. Here we reported that genetic deficiency of Malat1 leads to reduced blood vessel formation and local blood flow perfusion in mouse hind limbs at one to four weeks after hindlimb ischemia. Malat1 and vascular endothelial growth factor receptor 2 ( VEGFR2 ) levels were found to be increased in both cultured mouse primary skeletal muscle microvascular endothelial cells (SMMECs) after 16 h OGD followed by 24 h reperfusion and in mouse gastrocnemius muscle that underwent hindlimb ischemia followed by 28 days of reperfusion. Moreover, Malat1 silencing by locked nucleic acid (LNA)-GapmeRs significantly reduced tube formation, cell migration, and cell proliferation in SMMEC cultures. Mechanistically, RNA subcellular isolation and RNA-immunoprecipitation experiments demonstrate that Malat1 directly targets VEGFR2 to facilitate angiogenesis. The results suggest that Malat1 regulates cell-autonomous angiogenesis through direct regulation of VEGFR2.

Programmable DNA-binding proteins from Burkholderia provide a fresh perspective on the TALE-like repeat domain.

PubMed

de Lange, Orlando; Wolf, Christina; Dietze, Jörn; Elsaesser, Janett; Morbitzer, Robert; Lahaye, Thomas

2014-06-01

The tandem repeats of transcription activator like effectors (TALEs) mediate sequence-specific DNA binding using a simple code. Naturally, TALEs are injected by Xanthomonas bacteria into plant cells to manipulate the host transcriptome. In the laboratory TALE DNA binding domains are reprogrammed and used to target a fused functional domain to a genomic locus of choice. Research into the natural diversity of TALE-like proteins may provide resources for the further improvement of current TALE technology. Here we describe TALE-like proteins from the endosymbiotic bacterium Burkholderia rhizoxinica, termed Bat proteins. Bat repeat domains mediate sequence-specific DNA binding with the same code as TALEs, despite less than 40% sequence identity. We show that Bat proteins can be adapted for use as transcription factors and nucleases and that sequence preferences can be reprogrammed. Unlike TALEs, the core repeats of each Bat protein are highly polymorphic. This feature allowed us to explore alternative strategies for the design of custom Bat repeat arrays, providing novel insights into the functional relevance of non-RVD residues. The Bat proteins offer fertile grounds for research into the creation of improved programmable DNA-binding proteins and comparative insights into TALE-like evolution. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

A network of epigenetic regulators guides developmental haematopoiesis in vivo.

PubMed

Huang, Hsuan-Ting; Kathrein, Katie L; Barton, Abby; Gitlin, Zachary; Huang, Yue-Hua; Ward, Thomas P; Hofmann, Oliver; Dibiase, Anthony; Song, Anhua; Tyekucheva, Svitlana; Hide, Winston; Zhou, Yi; Zon, Leonard I

2013-12-01

The initiation of cellular programs is orchestrated by key transcription factors and chromatin regulators that activate or inhibit target gene expression. To generate a compendium of chromatin factors that establish the epigenetic code during developmental haematopoiesis, a large-scale reverse genetic screen was conducted targeting orthologues of 425 human chromatin factors in zebrafish. A set of chromatin regulators was identified that target different stages of primitive and definitive blood formation, including factors not previously implicated in haematopoiesis. We identified 15 factors that regulate development of primitive erythroid progenitors and 29 factors that regulate development of definitive haematopoietic stem and progenitor cells. These chromatin factors are associated with SWI/SNF and ISWI chromatin remodelling, SET1 methyltransferase, CBP-p300-HBO1-NuA4 acetyltransferase, HDAC-NuRD deacetylase, and Polycomb repressive complexes. Our work provides a comprehensive view of how specific chromatin factors and their associated complexes play a major role in the establishment of haematopoietic cells in vivo.

Transcription of the cottontail rabbit papillomavirus early region and identification of two E6 polypeptides in COS-7 cells.

PubMed Central

Barbosa, M S; Wettstein, F O

1987-01-01

Cottontail rabbit papillomavirus (CRPV) early proteins are present at very low levels in virus-induced tumors and cannot be detected by immunological methods. Furthermore, cells in culture are not readily transformed by the virus. To overcome these difficulties in identifying and characterizing the putative transforming protein(s) coded by the E6 open reading frame, the early cottontail rabbit papillomavirus region was expressed under the control of the late simian virus 40 promoter. Mapping of the transcripts in transiently transfected COS-7 cells indicated that transcription was initiated in the late region of simian virus 40. Two E6-coded polypeptides were identified, representing translation products initiated at the first and second AUG codons. Images PMID:3039182

The expanding regulatory universe of p53 in gastrointestinal cancer.

PubMed

Fesler, Andrew; Zhang, Ning; Ju, Jingfang

2016-01-01

Tumor suppresser gene TP53 is one of the most frequently deleted or mutated genes in gastrointestinal cancers. As a transcription factor, p53 regulates a number of important protein coding genes to control cell cycle, cell death, DNA damage/repair, stemness, differentiation and other key cellular functions. In addition, p53 is also able to activate the expression of a number of small non-coding microRNAs (miRNAs) through direct binding to the promoter region of these miRNAs.  Many miRNAs have been identified to be potential tumor suppressors by regulating key effecter target mRNAs. Our understanding of the regulatory network of p53 has recently expanded to include long non-coding RNAs (lncRNAs). Like miRNA, lncRNAs have been found to play important roles in cancer biology.  With our increased understanding of the important functions of these non-coding RNAs and their relationship with p53, we are gaining exciting new insights into the biology and function of cells in response to various growth environment changes. In this review we summarize the current understanding of the ever expanding involvement of non-coding RNAs in the p53 regulatory network and its implications for our understanding of gastrointestinal cancer.

Evolution of the unspliced transcriptome.

PubMed

Engelhardt, Jan; Stadler, Peter F

2015-08-20

Despite their abundance, unspliced EST data have received little attention as a source of information on non-coding RNAs. Very little is know, therefore, about the genomic distribution of unspliced non-coding transcripts and their relationship with the much better studied regularly spliced products. In particular, their evolution has remained virtually unstudied. We systematically study the evidence on unspliced transcripts available in EST annotation tracks for human and mouse, comprising 104,980 and 66,109 unspliced EST clusters, respectively. Roughly one third of these are located totally inside introns of known genes (TINs) and another third overlaps exonic regions (PINs). Eleven percent are "intergenic", far away from any annotated gene. Direct evidence for the independent transcription of many PINs and TINs is obtained from CAGE tag and chromatin data. We predict more than 2000 3'UTR-associated RNA candidates for each human and mouse. Fifteen to twenty percent of the unspliced EST cluster are conserved between human and mouse. With the exception of TINs, the sequences of unspliced EST clusters evolve significantly slower than genomic background. Furthermore, like spliced lincRNAs, they show highly tissue-specific expression patterns. Unspliced long non-coding RNAs are an important, rapidly evolving, component of mammalian transcriptomes. Their analysis is complicated by their preferential association with complex transcribed loci that usually also harbor a plethora of spliced transcripts. Unspliced EST data, although typically disregarded in transcriptome analysis, can be used to gain insights into this rarely investigated transcriptome component. The frequently postulated connection between lack of splicing and nuclear retention and the surprising overlap of chromatin-associated transcripts suggests that this class of transcripts might be involved in chromatin organization and possibly other mechanisms of epigenetic control.

Characteristics and significance of intergenic polyadenylated RNA transcription in Arabidopsis.

PubMed

Moghe, Gaurav D; Lehti-Shiu, Melissa D; Seddon, Alex E; Yin, Shan; Chen, Yani; Juntawong, Piyada; Brandizzi, Federica; Bailey-Serres, Julia; Shiu, Shin-Han

2013-01-01

The Arabidopsis (Arabidopsis thaliana) genome is the most well-annotated plant genome. However, transcriptome sequencing in Arabidopsis continues to suggest the presence of polyadenylated (polyA) transcripts originating from presumed intergenic regions. It is not clear whether these transcripts represent novel noncoding or protein-coding genes. To understand the nature of intergenic polyA transcription, we first assessed its abundance using multiple messenger RNA sequencing data sets. We found 6,545 intergenic transcribed fragments (ITFs) occupying 3.6% of Arabidopsis intergenic space. In contrast to transcribed fragments that map to protein-coding and RNA genes, most ITFs are significantly shorter, are expressed at significantly lower levels, and tend to be more data set specific. A surprisingly large number of ITFs (32.1%) may be protein coding based on evidence of translation. However, our results indicate that these "translated" ITFs tend to be close to and are likely associated with known genes. To investigate if ITFs are under selection and are functional, we assessed ITF conservation through cross-species as well as within-species comparisons. Our analysis reveals that 237 ITFs, including 49 with translation evidence, are under strong selective constraint and relatively distant from annotated features. These ITFs are likely parts of novel genes. However, the selective pressure imposed on most ITFs is similar to that of randomly selected, untranscribed intergenic sequences. Our findings indicate that despite the prevalence of ITFs, apart from the possibility of genomic contamination, many may be background or noisy transcripts derived from "junk" DNA, whose production may be inherent to the process of transcription and which, on rare occasions, may act as catalysts for the creation of novel genes.

An Integrated Encyclopedia of DNA Elements in the Human Genome

PubMed Central

2012-01-01

Summary The human genome encodes the blueprint of life, but the function of the vast majority of its nearly three billion bases is unknown. The Encyclopedia of DNA Elements (ENCODE) project has systematically mapped regions of transcription, transcription factor association, chromatin structure, and histone modification. These data enabled us to assign biochemical functions for 80% of the genome, in particular outside of the well-studied protein-coding regions. Many discovered candidate regulatory elements are physically associated with one another and with expressed genes, providing new insights into the mechanisms of gene regulation. The newly identified elements also show a statistical correspondence to sequence variants linked to human disease, and can thereby guide interpretation of this variation. Overall the project provides new insights into the organization and regulation of our genes and genome, and an expansive resource of functional annotations for biomedical research. PMID:22955616

Transcriptional diversity during lineage commitment of human blood progenitors.

PubMed

Chen, Lu; Kostadima, Myrto; Martens, Joost H A; Canu, Giovanni; Garcia, Sara P; Turro, Ernest; Downes, Kate; Macaulay, Iain C; Bielczyk-Maczynska, Ewa; Coe, Sophia; Farrow, Samantha; Poudel, Pawan; Burden, Frances; Jansen, Sjoert B G; Astle, William J; Attwood, Antony; Bariana, Tadbir; de Bono, Bernard; Breschi, Alessandra; Chambers, John C; Consortium, Bridge; Choudry, Fizzah A; Clarke, Laura; Coupland, Paul; van der Ent, Martijn; Erber, Wendy N; Jansen, Joop H; Favier, Rémi; Fenech, Matthew E; Foad, Nicola; Freson, Kathleen; van Geet, Chris; Gomez, Keith; Guigo, Roderic; Hampshire, Daniel; Kelly, Anne M; Kerstens, Hindrik H D; Kooner, Jaspal S; Laffan, Michael; Lentaigne, Claire; Labalette, Charlotte; Martin, Tiphaine; Meacham, Stuart; Mumford, Andrew; Nürnberg, Sylvia; Palumbo, Emilio; van der Reijden, Bert A; Richardson, David; Sammut, Stephen J; Slodkowicz, Greg; Tamuri, Asif U; Vasquez, Louella; Voss, Katrin; Watt, Stephen; Westbury, Sarah; Flicek, Paul; Loos, Remco; Goldman, Nick; Bertone, Paul; Read, Randy J; Richardson, Sylvia; Cvejic, Ana; Soranzo, Nicole; Ouwehand, Willem H; Stunnenberg, Hendrik G; Frontini, Mattia; Rendon, Augusto

2014-09-26

Blood cells derive from hematopoietic stem cells through stepwise fating events. To characterize gene expression programs driving lineage choice, we sequenced RNA from eight primary human hematopoietic progenitor populations representing the major myeloid commitment stages and the main lymphoid stage. We identified extensive cell type-specific expression changes: 6711 genes and 10,724 transcripts, enriched in non-protein-coding elements at early stages of differentiation. In addition, we found 7881 novel splice junctions and 2301 differentially used alternative splicing events, enriched in genes involved in regulatory processes. We demonstrated experimentally cell-specific isoform usage, identifying nuclear factor I/B (NFIB) as a regulator of megakaryocyte maturation-the platelet precursor. Our data highlight the complexity of fating events in closely related progenitor populations, the understanding of which is essential for the advancement of transplantation and regenerative medicine. Copyright © 2014, American Association for the Advancement of Science.

An Autonomous BMP2 Regulatory Element in Mesenchymal Cells

PubMed Central

Kruithof, Boudewijn P.T.; Fritz, David T.; Liu, Yijun; Garsetti, Diane E.; Frank, David B.; Pregizer, Steven K.; Gaussin, Vinciane; Mortlock, Douglas P.; Rogers, Melissa B.

2014-01-01

BMP2 is a morphogen that controls mesenchymal cell differentiation and behavior. For example, BMP2 concentration controls the differentiation of mesenchymal precursors into myocytes, adipocytes, chondrocytes, and osteoblasts. Sequences within the 3′untranslated region (UTR) of the Bmp2 mRNA mediate a post-transcriptional block of protein synthesis. Interaction of cell and developmental stage-specific trans-regulatory factors with the 3′UTR is a nimble and versatile mechanism for modulating this potent morphogen in different cell types. We show here, that an ultra-conserved sequence in the 3′UTR functions independently of promoter, coding region, and 3′UTR context in primary and immortalized tissue culture cells and in transgenic mice. Our findings indicate that the ultra-conserved sequence is an autonomously functioning post-transcriptional element that may be used to modulate the level of BMP2 and other proteins while retaining tissue specific regulatory elements. PMID:21268088

Integration of multi-omics data of a genome-reduced bacterium: Prevalence of post-transcriptional regulation and its correlation with protein abundances

PubMed Central

Chen, Wei-Hua; van Noort, Vera; Lluch-Senar, Maria; Hennrich, Marco L.; H. Wodke, Judith A.; Yus, Eva; Alibés, Andreu; Roma, Guglielmo; Mende, Daniel R.; Pesavento, Christina; Typas, Athanasios; Gavin, Anne-Claude; Serrano, Luis; Bork, Peer

2016-01-01

We developed a comprehensive resource for the genome-reduced bacterium Mycoplasma pneumoniae comprising 1748 consistently generated ‘-omics’ data sets, and used it to quantify the power of antisense non-coding RNAs (ncRNAs), lysine acetylation, and protein phosphorylation in predicting protein abundance (11%, 24% and 8%, respectively). These factors taken together are four times more predictive of the proteome abundance than of mRNA abundance. In bacteria, post-translational modifications (PTMs) and ncRNA transcription were both found to increase with decreasing genomic GC-content and genome size. Thus, the evolutionary forces constraining genome size and GC-content modify the relative contributions of the different regulatory layers to proteome homeostasis, and impact more genomic and genetic features than previously appreciated. Indeed, these scaling principles will enable us to develop more informed approaches when engineering minimal synthetic genomes. PMID:26773059

Disentangling the many layers of eukaryotic transcriptional regulation.

PubMed

Lelli, Katherine M; Slattery, Matthew; Mann, Richard S

2012-01-01

Regulation of gene expression in eukaryotes is an extremely complex process. In this review, we break down several critical steps, emphasizing new data and techniques that have expanded current gene regulatory models. We begin at the level of DNA sequence where cis-regulatory modules (CRMs) provide important regulatory information in the form of transcription factor (TF) binding sites. In this respect, CRMs function as instructional platforms for the assembly of gene regulatory complexes. We discuss multiple mechanisms controlling complex assembly, including cooperative DNA binding, combinatorial codes, and CRM architecture. The second section of this review places CRM assembly in the context of nucleosomes and condensed chromatin. We discuss how DNA accessibility and histone modifications contribute to TF function. Lastly, new advances in chromosomal mapping techniques have provided increased understanding of intra- and interchromosomal interactions. We discuss how these topological maps influence gene regulatory models.

Expression, regulation and functional assessment of the 80 amino acid Small Adipocyte Factor 1 (Smaf1) protein in adipocytes.

PubMed

Ren, Gang; Eskandari, Parisa; Wang, Siqian; Smas, Cynthia M

2016-01-15

The gene for Small Adipocyte Factor 1, Smaf1 (also known as adipogenin, ADIG), encodes a ∼600 base transcript that is highly upregulated during 3T3-L1 in vitro adipogenesis and markedly enriched in adipose tissues. Based on the lack of an obvious open reading frame in the Smaf1 transcript, it is not known if the Smaf1 gene is protein coding or non-coding RNA. Using a peptide from a putative open reading frame of Smaf1 as antigen, we generated antibodies for western analysis. Our studies prove that Smaf1 encodes an adipose-enriched protein which in western blot analysis migrates at ∼10 kDa. Rapid induction of Smaf1 protein occurs during in vitro adipogenesis and its expression in 3T3-L1 adipocytes is positively regulated by insulin and glucose. Moreover, siRNA studies reveal that expression of Smaf1 in adipocytes is wholly dependent on PPARγ. On the other hand, use of siRNA for Smaf1 to nearly abolish its protein expression in adipocytes revealed that Smaf1 does not have a major role in adipocyte triglyceride accumulation, lipolysis or insulin-stimulated pAkt induction. However, immunolocalization studies using HA-tagged Smaf1 reveal enrichment at adipocyte lipid droplets. Together our findings show that Smaf1 is a novel small protein endogenous to adipocytes and that Smaf1 expression is closely tied to PPARγ-mediated signals and the adipocyte phenotype. Copyright © 2015 Elsevier Inc. All rights reserved.

CINCINNATA in Antirrhinum majus directly modulates genes involved in cytokinin and auxin signaling.

PubMed

Das Gupta, Mainak; Aggarwal, Pooja; Nath, Utpal

2014-12-01

Mutations in the CINCINNATA (CIN) gene in Antirrhinum majus and its orthologs in Arabidopsis result in crinkly leaves as a result of excess growth towards the leaf margin. CIN homologs code for TCP (TEOSINTE-BRANCHED 1, CYCLOIDEA, PROLIFERATING CELL FACTOR 1 AND 2) transcription factors and are expressed in a broad zone in a growing leaf distal to the proliferation zone where they accelerate cell maturation. Although a few TCP targets are known, the functional basis of CIN-mediated leaf morphogenesis remains unclear. We compared the global transcription profiles of wild-type and the cin mutant of A. majus to identify the targets of CIN. We cloned and studied the direct targets using RNA in situ hybridization, DNA-protein interaction, chromatin immunoprecipitation and reporter gene analysis. Many of the genes involved in the auxin and cytokinin signaling pathways showed altered expression in the cin mutant. Further, we showed that CIN binds to genomic regions and directly promotes the transcription of a cytokinin receptor homolog HISTIDINE KINASE 4 (AmHK4) and an IAA3/SHY2 (INDOLE-3-ACETIC ACID INDUCIBLE 3/SHORT HYPOCOTYL 2) homolog in A. majus. Our results suggest that CIN limits excess cell proliferation and maintains the flatness of the leaf surface by directly modulating the hormone pathways involved in patterning cell proliferation and differentiation during leaf growth. © 2014 The Authors. New Phytologist © 2014 New Phytologist Trust.

Genome-wide transcriptome analysis of Clavibacter michiganensis subsp. michiganensis grown in xylem mimicking medium.

PubMed

Hiery, Eva; Adam, Susanne; Reid, Stephen; Hofmann, Jörg; Sonnewald, Sophia; Burkovski, Andreas

2013-12-01

The interaction between Clavibacter michiganensis subsp. michiganensis with its host, the tomato plant (Solanum lycopersicum), is poorly understood and only few virulence factors are known. While studying of the bacteria in planta is time-consuming and difficult, the analysis in vitro would facilitate research. Therefore, a xylem mimicking medium (XMM) for C. michiganensis subsp. michiganensis was established in this study based on an apoplast medium for Xanthomonas campestris pv. vesicatoria. In contrast to the apoplast medium, XMM contains no sugars, but amino acids which serve as nitrogen and carbon source. As a result, growth in XMM induced transcriptional changes of genes encoding putative sugar, amino acid and iron uptake systems. In summary, mRNA levels of about 8% of all C. michiganensis subsp. michiganensis genes were changed when XMM-grown bacteria were compared to M9 minimal medium-grown cells. Almost no transcriptional changes of genes encoding hydrolytic enzymes were detected, leading to the idea that XMM reflects the situation in the beginning of infection and therefore allows the characterization of virulence factors in this early stage of infection. The addition of the plant wound substance acetosyringone to the XMM medium led to a change in transcript amount, including genes coding for proteins involved in protein transport, iron uptake and regulation processes. Copyright © 2013 Elsevier B.V. All rights reserved.

Exploration of jasmonate signalling via automated and standardized transient expression assays in tobacco cells.

PubMed

De Sutter, Valerie; Vanderhaeghen, Rudy; Tilleman, Sofie; Lammertyn, Freya; Vanhoutte, Isabelle; Karimi, Mansour; Inzé, Dirk; Goossens, Alain; Hilson, Pierre

2005-12-01

Although sequence information and genome annotation are improving at an impressive pace, functional ontology is still non-existent or rudimentary for most genes. In this regard, transient expression assays are very valuable for identification of short functional segments in particular pathways, because they can be performed rapidly and at a scale unattainable in stably transformed tissues. Vectors were constructed and protocols developed for systematic transient assays in plant protoplasts. To enhance throughput and reproducibility, protoplast treatments were performed entirely by a liquid-handling robot in multiwell plates, including polyethylene glycol/Ca2+ cell transfection with plasmid mixtures, washes and lysis. All transcriptional readouts were measured using a dual firefly/Renilla luciferase assay, in which the former was controlled by a reporter promoter and the latter by the 35S CaMV promoter, which served as internal normalization standard. The automated protocols were suitable for transient assays in protoplasts prepared from cell cultures of Nicotiana tabacum Bright Yellow-2 and Arabidopsis thaliana. They were implemented in a screen to discover potential regulators of genes coding for key enzymes in nicotine biosynthesis. Two novel tobacco transcription factors were found, NtORC1 and NtJAP1, that positively regulate the putrescine N-methyltransferase (PMT) promoter. In addition, combinatorial tests showed that these two factors act synergistically to induce PMT transcriptional activity. The development and use of high-throughput plant transient expression assays are discussed.

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

PubMed

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

2015-12-01

The majority of the human genome consists of non-coding regions that have been called junk DNA. However, recent studies have unveiled that these regions contain cis-regulatory elements, such as promoters, enhancers, silencers, insulators, etc. These regulatory elements can play crucial roles in controlling gene expressions in specific cell types, conditions, and developmental stages. Disruption to these regions could contribute to phenotype changes. Precisely identifying regulatory elements is key to deciphering the mechanisms underlying transcriptional regulation. Cis-regulatory events are complex processes that involve chromatin accessibility, transcription factor binding, DNA methylation, histone modifications, and the interactions between them. The development of next-generation sequencing techniques has allowed us to capture these genomic features in depth. Applied analysis of genome sequences for clinical genetics has increased the urgency for detecting these regions. However, the complexity of cis-regulatory events and the deluge of sequencing data require accurate and efficient computational approaches, in particular, machine learning techniques. In this review, we describe machine learning approaches for predicting transcription factor binding sites, enhancers, and promoters, primarily driven by next-generation sequencing data. Data sources are provided in order to facilitate testing of novel methods. The purpose of this review is to attract computational experts and data scientists to advance this field. Crown Copyright © 2015. Published by Elsevier Ireland Ltd. All rights reserved.

The bromodomain protein BRD4 regulates splicing during heat shock

PubMed Central

Hussong, Michelle; Kaehler, Christian; Kerick, Martin; Grimm, Christina; Franz, Alexandra; Timmermann, Bernd; Welzel, Franziska; Isensee, Jörg; Hucho, Tim; Krobitsch, Sylvia; Schweiger, Michal R.

2017-01-01

The cellular response to heat stress is an ancient and evolutionarily highly conserved defence mechanism characterised by the transcriptional up-regulation of cyto-protective genes and a partial inhibition of splicing. These features closely resemble the proteotoxic stress response during tumor development. The bromodomain protein BRD4 has been identified as an integral member of the oxidative stress as well as of the inflammatory response, mainly due to its role in the transcriptional regulation process. In addition, there are also several lines of evidence implicating BRD4 in the splicing process. Using RNA-sequencing we found a significant increase in splicing inhibition, in particular intron retentions (IR), following heat treatment in BRD4-depleted cells. This leads to a decrease of mRNA abundancy of the affected transcripts, most likely due to premature termination codons. Subsequent experiments revealed that BRD4 interacts with the heat shock factor 1 (HSF1) such that under heat stress BRD4 is recruited to nuclear stress bodies and non-coding SatIII RNA transcripts are up-regulated. These findings implicate BRD4 as an important regulator of splicing during heat stress. Our data which links BRD4 to the stress induced splicing process may provide novel mechanisms of BRD4 inhibitors in regard to anti-cancer therapies. PMID:27536004

MicroRNAs: regulators of gene expression and cell differentiation

PubMed Central

Shivdasani, Ramesh A.

2006-01-01

The existence and roles of a class of abundant regulatory RNA molecules have recently come into sharp focus. Micro-RNAs (miRNAs) are small (approximately 22 bases), non–protein-coding RNAs that recognize target sequences of imperfect complementarity in cognate mRNAs and either destabilize them or inhibit protein translation. Although mechanisms of miRNA biogenesis have been elucidated in some detail, there is limited appreciation of their biological functions. Reported examples typically focus on miRNA regulation of a single tissue-restricted transcript, often one encoding a transcription factor, that controls a specific aspect of development, cell differentiation, or physiology. However, computational algorithms predict up to hundreds of putative targets for individual miRNAs, single transcripts may be regulated by multiple miRNAs, and miRNAs may either eliminate target gene expression or serve to finetune transcript and protein levels. Theoretical considerations and early experimental results hence suggest diverse roles for miRNAs as a class. One appealing possibility, that miRNAs eliminate low-level expression of unwanted genes and hence refine unilineage gene expression, may be especially amenable to evaluation in models of hematopoiesis. This review summarizes current understanding of miRNA mechanisms, outlines some of the important outstanding questions, and describes studies that attempt to define miRNA functions in hematopoiesis. PMID:16882713

Competition between histone and transcription factor binding regulates the onset of transcription in zebrafish embryos

PubMed Central

Joseph, Shai R; Pálfy, Máté; Hilbert, Lennart; Kumar, Mukesh; Karschau, Jens; Zaburdaev, Vasily; Shevchenko, Andrej; Vastenhouw, Nadine L

2017-01-01

Upon fertilization, the genome of animal embryos remains transcriptionally inactive until the maternal-to-zygotic transition. At this time, the embryo takes control of its development and transcription begins. How the onset of zygotic transcription is regulated remains unclear. Here, we show that a dynamic competition for DNA binding between nucleosome-forming histones and transcription factors regulates zebrafish genome activation. Taking a quantitative approach, we found that the concentration of non-DNA-bound core histones sets the time for the onset of transcription. The reduction in nuclear histone concentration that coincides with genome activation does not affect nucleosome density on DNA, but allows transcription factors to compete successfully for DNA binding. In agreement with this, transcription factor binding is sensitive to histone levels and the concentration of transcription factors also affects the time of transcription. Our results demonstrate that the relative levels of histones and transcription factors regulate the onset of transcription in the embryo. DOI: http://dx.doi.org/10.7554/eLife.23326.001 PMID:28425915

Competition between histone and transcription factor binding regulates the onset of transcription in zebrafish embryos.

PubMed

Joseph, Shai R; Pálfy, Máté; Hilbert, Lennart; Kumar, Mukesh; Karschau, Jens; Zaburdaev, Vasily; Shevchenko, Andrej; Vastenhouw, Nadine L

2017-04-20

Upon fertilization, the genome of animal embryos remains transcriptionally inactive until the maternal-to-zygotic transition. At this time, the embryo takes control of its development and transcription begins. How the onset of zygotic transcription is regulated remains unclear. Here, we show that a dynamic competition for DNA binding between nucleosome-forming histones and transcription factors regulates zebrafish genome activation. Taking a quantitative approach, we found that the concentration of non-DNA-bound core histones sets the time for the onset of transcription. The reduction in nuclear histone concentration that coincides with genome activation does not affect nucleosome density on DNA, but allows transcription factors to compete successfully for DNA binding. In agreement with this, transcription factor binding is sensitive to histone levels and the concentration of transcription factors also affects the time of transcription. Our results demonstrate that the relative levels of histones and transcription factors regulate the onset of transcription in the embryo.

Universal evolutionary selection for high dimensional silent patterns of information hidden in the redundancy of viral genetic code.

PubMed

Goz, Eli; Zafrir, Zohar; Tuller, Tamir

2018-04-30

Understanding how viruses co-evolve with their hosts and adapt various genomic level strategies in order to ensure their fitness may have essential implications in unveiling the secrets of viral evolution, and in developing new vaccines and therapeutic approaches. Here, based on a novel genomic analysis of 2,625 different viruses and 439 corresponding host organisms, we provide evidence of universal evolutionary selection for high dimensional 'silent' patterns of information hidden in the redundancy of viral genetic code. Our model suggests that long substrings of nucleotides in the coding regions of viruses from all classes, often also repeat in the corresponding viral hosts from all domains of life. Selection for these substrings cannot be explained only by such phenomena as codon usage bias, horizontal gene transfer, and the encoded proteins. Genes encoding structural proteins responsible for building the core of the viral particles were found to include more host-repeating substrings, and these substrings tend to appear in the middle parts of the viral coding regions. In addition, in human viruses these substrings tend to be enriched with motives related to transcription factors and RNA binding proteins. The host-repeating substrings are possibly related to the evolutionary pressure on the viruses to effectively interact with host's intracellular factors and to efficiently escape from the host's immune system. tamirtul@post.tau.ac.il (TT). Supplementary data are available at Bioinformatics online.

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

PubMed

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.

Bombyx mori Transcription Factors: Genome-Wide Identification, Expression Profiles and Response to Pathogens by Microarray Analysis

PubMed Central

Huang, Lulin; Cheng, Tingcai; Xu, Pingzhen; Fang, Ting; Xia, Qingyou

2012-01-01

Transcription factors are present in all living organisms, and play vital roles in a wide range of biological processes. Studies of transcription factors will help reveal the complex regulation mechanism of organisms. So far, hundreds of domains have been identified that show transcription factor activity. Here, 281 reported transcription factor domains were used as seeds to search the transcription factors in genomes of Bombyx mori L. (Lepidoptera: Bombycidae) and four other model insects. Overall, 666 transcription factors including 36 basal factors and 630 other factors were identified in B. mori genome, which accounted for 4.56% of its genome. The silkworm transcription factors' expression profiles were investigated in relation to multiple tissues, developmental stages, sexual dimorphism, and responses to oral infection by pathogens and direct bacterial injection. These all provided rich clues for revealing the transcriptional regulation mechanism of silkworm organ differentiation, growth and development, sexual dimorphism, and response to pathogen infection. PMID:22943524

Fatty Acid–Regulated Transcription Factors in the Liver

PubMed Central

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

The impact of online therapeutic feedback on outcome measures in Internet-CBTI for adolescents with insomnia.

PubMed

de Bruin, Eduard Jan; Meijer, Anne Marie

2017-01-01

Guided Internet cognitive behavioral therapy for insomnia (CBTI) offers an effective treatment for adolescents, but little is known about the active ingredients of therapeutic feedback on outcomes. This study aims to identify which factors can be distinguished in written therapeutic feedback in Internet CBTI, and examine whether these factors and participation in a chat session contribute to sleep outcomes. Internet CBTI was applied to 57 adolescents (mean age 15.43 years, SD 1.74, 82.5% girls). Symptoms of insomnia and chronic sleep reduction, and total sleep time, time in bed, and sleep efficiency from seven day sleep logs were measured at baseline, post-treatment, and at two month follow-up. With a coding instrument developed for this study, two independent researchers coded transcripts of the written therapeutic feedback of the Internet CBTI sessions with an event sampling method. Principal component analysis of the initial 17 items from the coding instrument yielded four distinct factors of therapeutic feedback, of which only Sleep expertise seemed to contribute to improvements after Internet CBTI. The other factors, indicating forms of encouragement, and participation in a chat session seemed counterproductive. This first longitudinal study into effects of therapeutic feedback in adolescent Internet CBTI indicated that emphasizing knowledge about sleep might contribute to insomnia improvement. The structured nature of the preprogrammed treatment content, delay of therapeutic feedback due to standardized timing, and unintentional reinforcement of undesirable behavior by giving attention to failures might explain the negative results of encouraging behavior. Further research to identify effective therapeutic factors in Internet therapy is warranted. Copyright © 2016 Elsevier B.V. All rights reserved.

A model of self-directed learning in internal medicine residency: a qualitative study using grounded theory.

PubMed

Sawatsky, Adam P; Ratelle, John T; Bonnes, Sara L; Egginton, Jason S; Beckman, Thomas J

2017-02-02

Existing theories of self-directed learning (SDL) have emphasized the importance of process, personal, and contextual factors. Previous medical education research has largely focused on the process of SDL. We explored the experience with and perception of SDL among internal medicine residents to gain understanding of the personal and contextual factors of SDL in graduate medical education. Using a constructivist grounded theory approach, we conducted 7 focus group interviews with 46 internal medicine residents at an academic medical center. We processed the data by using open coding and writing analytic memos. Team members organized open codes to create axial codes, which were applied to all transcripts. Guided by a previous model of SDL, we developed a theoretical model that was revised through constant comparison with new data as they were collected, and we refined the theory until it had adequate explanatory power and was appropriately grounded in the experiences of residents. We developed a theoretical model of SDL to explain the process, personal, and contextual factors affecting SDL during residency training. The process of SDL began with a trigger that uncovered a knowledge gap. Residents progressed to formulating learning objectives, using resources, applying knowledge, and evaluating learning. Personal factors included motivations, individual characteristics, and the change in approach to SDL over time. Contextual factors included the need for external guidance, the influence of residency program structure and culture, and the presence of contextual barriers. We developed a theoretical model of SDL in medical education that can be used to promote and assess resident SDL through understanding the process, person, and context of SDL.