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Sample records for genome transcriptional analysis

  1. Arabidopsis transcription factors: genome-wide comparative analysis among eukaryotes.

    Science.gov (United States)

    Riechmann, J L; Heard, J; Martin, G; Reuber, L; Jiang, C; Keddie, J; Adam, L; Pineda, O; Ratcliffe, O J; Samaha, R R; Creelman, R; Pilgrim, M; Broun, P; Zhang, J Z; Ghandehari, D; Sherman, B K; Yu, G

    2000-12-15

    The completion of the Arabidopsis thaliana genome sequence allows a comparative analysis of transcriptional regulators across the three eukaryotic kingdoms. Arabidopsis dedicates over 5% of its genome to code for more than 1500 transcription factors, about 45% of which are from families specific to plants. Arabidopsis transcription factors that belong to families common to all eukaryotes do not share significant similarity with those of the other kingdoms beyond the conserved DNA binding domains, many of which have been arranged in combinations specific to each lineage. The genome-wide comparison reveals the evolutionary generation of diversity in the regulation of transcription.

  2. In silico comparative genomic analysis of GABAA receptor transcriptional regulation

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    Joyce Christopher J

    2007-06-01

    Full Text Available Abstract Background Subtypes of the GABAA receptor subunit exhibit diverse temporal and spatial expression patterns. In silico comparative analysis was used to predict transcriptional regulatory features in individual mammalian GABAA receptor subunit genes, and to identify potential transcriptional regulatory components involved in the coordinate regulation of the GABAA receptor gene clusters. Results Previously unreported putative promoters were identified for the β2, γ1, γ3, ε, θ and π subunit genes. Putative core elements and proximal transcriptional factors were identified within these predicted promoters, and within the experimentally determined promoters of other subunit genes. Conserved intergenic regions of sequence in the mammalian GABAA receptor gene cluster comprising the α1, β2, γ2 and α6 subunits were identified as potential long range transcriptional regulatory components involved in the coordinate regulation of these genes. A region of predicted DNase I hypersensitive sites within the cluster may contain transcriptional regulatory features coordinating gene expression. A novel model is proposed for the coordinate control of the gene cluster and parallel expression of the α1 and β2 subunits, based upon the selective action of putative Scaffold/Matrix Attachment Regions (S/MARs. Conclusion The putative regulatory features identified by genomic analysis of GABAA receptor genes were substantiated by cross-species comparative analysis and now require experimental verification. The proposed model for the coordinate regulation of genes in the cluster accounts for the head-to-head orientation and parallel expression of the α1 and β2 subunit genes, and for the disruption of transcription caused by insertion of a neomycin gene in the close vicinity of the α6 gene, which is proximal to a putative critical S/MAR.

  3. Genome-wide identification of the regulatory targets of a transcription factor using biochemical characterization and computational genomic analysis

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    Jolly Emmitt R

    2005-11-01

    Full Text Available Abstract Background A major challenge in computational genomics is the development of methodologies that allow accurate genome-wide prediction of the regulatory targets of a transcription factor. We present a method for target identification that combines experimental characterization of binding requirements with computational genomic analysis. Results Our method identified potential target genes of the transcription factor Ndt80, a key transcriptional regulator involved in yeast sporulation, using the combined information of binding affinity, positional distribution, and conservation of the binding sites across multiple species. We have also developed a mathematical approach to compute the false positive rate and the total number of targets in the genome based on the multiple selection criteria. Conclusion We have shown that combining biochemical characterization and computational genomic analysis leads to accurate identification of the genome-wide targets of a transcription factor. The method can be extended to other transcription factors and can complement other genomic approaches to transcriptional regulation.

  4. Broad genomic and transcriptional analysis reveals a highly derived genome in dinoflagellate mitochondria

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    Keeling Patrick J

    2007-09-01

    Full Text Available Abstract Background Dinoflagellates comprise an ecologically significant and diverse eukaryotic phylum that is sister to the phylum containing apicomplexan endoparasites. The mitochondrial genome of apicomplexans is uniquely reduced in gene content and size, encoding only three proteins and two ribosomal RNAs (rRNAs within a highly compacted 6 kb DNA. Dinoflagellate mitochondrial genomes have been comparatively poorly studied: limited available data suggest some similarities with apicomplexan mitochondrial genomes but an even more radical type of genomic organization. Here, we investigate structure, content and expression of dinoflagellate mitochondrial genomes. Results From two dinoflagellates, Crypthecodinium cohnii and Karlodinium micrum, we generated over 42 kb of mitochondrial genomic data that indicate a reduced gene content paralleling that of mitochondrial genomes in apicomplexans, i.e., only three protein-encoding genes and at least eight conserved components of the highly fragmented large and small subunit rRNAs. Unlike in apicomplexans, dinoflagellate mitochondrial genes occur in multiple copies, often as gene fragments, and in numerous genomic contexts. Analysis of cDNAs suggests several novel aspects of dinoflagellate mitochondrial gene expression. Polycistronic transcripts were found, standard start codons are absent, and oligoadenylation occurs upstream of stop codons, resulting in the absence of termination codons. Transcripts of at least one gene, cox3, are apparently trans-spliced to generate full-length mRNAs. RNA substitutional editing, a process previously identified for mRNAs in dinoflagellate mitochondria, is also implicated in rRNA expression. Conclusion The dinoflagellate mitochondrial genome shares the same gene complement and fragmentation of rRNA genes with its apicomplexan counterpart. However, it also exhibits several unique characteristics. Most notable are the expansion of gene copy numbers and their arrangements

  5. Rice-arsenate interactions in hydroponics: whole genome transcriptional analysis.

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    Norton, Gareth J; Lou-Hing, Daniel E; Meharg, Andrew A; Price, Adam H

    2008-01-01

    Rice (Oryza sativa) varieties that are arsenate-tolerant (Bala) and -sensitive (Azucena) were used to conduct a transcriptome analysis of the response of rice seedlings to sodium arsenate (AsV) in hydroponic solution. RNA extracted from the roots of three replicate experiments of plants grown for 1 week in phosphate-free nutrient with or without 13.3 muM AsV was used to challenge the Affymetrix (52K) GeneChip Rice Genome array. A total of 576 probe sets were significantly up-regulated at least 2-fold in both varieties, whereas 622 were down-regulated. Ontological classification is presented. As expected, a large number of transcription factors, stress proteins, and transporters demonstrated differential expression. Striking is the lack of response of classic oxidative stress-responsive genes or phytochelatin synthases/synthatases. However, the large number of responses from genes involved in glutathione synthesis, metabolism, and transport suggests that glutathione conjugation and arsenate methylation may be important biochemical responses to arsenate challenge. In this report, no attempt is made to dissect differences in the response of the tolerant and sensitive variety, but analysis in a companion article will link gene expression to the known tolerance loci available in the BalaxAzucena mapping population.

  6. Rice–arsenate interactions in hydroponics: whole genome transcriptional analysis

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    Norton, Gareth J.; Lou-Hing, Daniel E.; Meharg, Andrew A.; Price, Adam H.

    2008-01-01

    Rice (Oryza sativa) varieties that are arsenate-tolerant (Bala) and -sensitive (Azucena) were used to conduct a transcriptome analysis of the response of rice seedlings to sodium arsenate (AsV) in hydroponic solution. RNA extracted from the roots of three replicate experiments of plants grown for 1 week in phosphate-free nutrient with or without 13.3 μM AsV was used to challenge the Affymetrix (52K) GeneChip Rice Genome array. A total of 576 probe sets were significantly up-regulated at least 2-fold in both varieties, whereas 622 were down-regulated. Ontological classification is presented. As expected, a large number of transcription factors, stress proteins, and transporters demonstrated differential expression. Striking is the lack of response of classic oxidative stress-responsive genes or phytochelatin synthases/synthatases. However, the large number of responses from genes involved in glutathione synthesis, metabolism, and transport suggests that glutathione conjugation and arsenate methylation may be important biochemical responses to arsenate challenge. In this report, no attempt is made to dissect differences in the response of the tolerant and sensitive variety, but analysis in a companion article will link gene expression to the known tolerance loci available in the Bala×Azucena mapping population. PMID:18453530

  7. Gene prediction and RFX transcriptional regulation analysis using comparative genomics

    OpenAIRE

    Chu, Jeffrey Shih Chieh

    2011-01-01

    Regulatory Factor X (RFX) is a family of transcription factors (TF) that is conserved in all metazoans, in some fungi, and in only a few single-cellular organisms. Seven members are found in mammals, nine in fishes, three in fruit flies, and a single member in nematodes and fungi. RFX is involved in many different roles in humans, but a particular function that is conserved in many metazoans is its regulation of ciliogenesis. Probing over 150 genomes for the presence of RFX and ciliary genes ...

  8. Comparative transcriptional and genomic analysis of Plasmodium falciparum field isolates.

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    Margaret J Mackinnon

    2009-10-01

    Full Text Available Mechanisms for differential regulation of gene expression may underlie much of the phenotypic variation and adaptability of malaria parasites. Here we describe transcriptional variation among culture-adapted field isolates of Plasmodium falciparum, the species responsible for most malarial disease. It was found that genes coding for parasite protein export into the red cell cytosol and onto its surface, and genes coding for sexual stage proteins involved in parasite transmission are up-regulated in field isolates compared with long-term laboratory isolates. Much of this variability was associated with the loss of small or large chromosomal segments, or other forms of gene copy number variation that are prevalent in the P. falciparum genome (copy number variants, CNVs. Expression levels of genes inside these segments were correlated to that of genes outside and adjacent to the segment boundaries, and this association declined with distance from the CNV boundary. This observation could not be explained by copy number variation in these adjacent genes. This suggests a local-acting regulatory role for CNVs in transcription of neighboring genes and helps explain the chromosomal clustering that we observed here. Transcriptional co-regulation of physical clusters of adaptive genes may provide a way for the parasite to readily adapt to its highly heterogeneous and strongly selective environment.

  9. Genome wide analysis of stress responsive WRKY transcription factors in Arabidopsis thaliana

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    Shaiq Sultan

    2016-04-01

    Full Text Available WRKY transcription factors are a class of DNA-binding proteins that bind with a specific sequence C/TTGACT/C known as W-Box found in promoters of genes which are regulated by these WRKYs. From previous studies, 43 different stress responsive WRKY transcription factors in Arabidopsis thaliana, identified and then categorized in three groups viz., abiotic, biotic and both of these stresses. A comprehensive genome wide analysis including chromosomal localization, gene structure analysis, multiple sequence alignment, phylogenetic analysis and promoter analysis of these WRKY genes was carried out in this study to determine the functional homology in Arabidopsis. This analysis led to the classification of these WRKY family members into 3 major groups and subgroups and showed evolutionary relationship among these groups on the base of their functional WRKY domain, chromosomal localization and intron/exon structure. The proposed groups of these stress responsive WRKY genes and annotation based on their position on chromosomes can also be explored to determine their functional homology in other plant species in relation to different stresses. The result of the present study provides indispensable genomic information for the stress responsive WRKY transcription factors in Arabidopsis and will pave the way to explain the precise role of various AtWRKYs in plant growth and development under stressed conditions.

  10. Comparative Genomics and Transcriptional Analysis of Prophages Identified in the Genomes of Lactobacillus gasseri, Lactobacillus salivarius, and Lactobacillus casei†

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    Ventura, Marco; Canchaya, Carlos; Bernini, Valentina; Altermann, Eric; Barrangou, Rodolphe; McGrath, Stephen; Claesson, Marcus J.; Li, Yin; Leahy, Sinead; Walker, Carey D.; Zink, Ralf; Neviani, Erasmo; Steele, Jim; Broadbent, Jeff; Klaenhammer, Todd R.; Fitzgerald, Gerald F.; O'Toole, Paul W.; van Sinderen, Douwe

    2006-01-01

    Lactobacillus gasseri ATCC 33323, Lactobacillus salivarius subsp. salivarius UCC 118, and Lactobacillus casei ATCC 334 contain one (LgaI), four (Sal1, Sal2, Sal3, Sal4), and one (Lca1) distinguishable prophage sequences, respectively. Sequence analysis revealed that LgaI, Lca1, Sal1, and Sal2 prophages belong to the group of Sfi11-like pac site and cos site Siphoviridae, respectively. Phylogenetic investigation of these newly described prophage sequences revealed that they have not followed an evolutionary development similar to that of their bacterial hosts and that they show a high degree of diversity, even within a species. The attachment sites were determined for all these prophage elements; LgaI as well as Sal1 integrates in tRNA genes, while prophage Sal2 integrates in a predicted arginino-succinate lyase-encoding gene. In contrast, Lca1 and the Sal3 and Sal4 prophage remnants are integrated in noncoding regions in the L. casei ATCC 334 and L. salivarius UCC 118 genomes. Northern analysis showed that large parts of the prophage genomes are transcriptionally silent and that transcription is limited to genome segments located near the attachment site. Finally, pulsed-field gel electrophoresis followed by Southern blot hybridization with specific prophage probes indicates that these prophage sequences are narrowly distributed within lactobacilli. PMID:16672450

  11. Genome-wide analysis of the WRKY transcription factors in aegilops tauschii.

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    Ma, Jianhui; Zhang, Daijing; Shao, Yun; Liu, Pei; Jiang, Lina; Li, Chunxi

    2014-01-01

    The WRKY transcription factors (TFs) play important roles in responding to abiotic and biotic stress in plants. However, due to its unfinished genome sequencing, relatively few WRKY TFs with full-length coding sequences (CDSs) have been identified in wheat. Instead, the Aegilops tauschii genome, which is the D-genome progenitor of the hexaploid wheat genome, provides important resources for the discovery of new genes. In this study, we performed a bioinformatics analysis to identify WRKY TFs with full-length CDSs from the A. tauschii genome. A detailed evolutionary analysis for all these TFs was conducted, and quantitative real-time PCR was carried out to investigate the expression patterns of the abiotic stress-related WRKY TFs under different abiotic stress conditions in A. tauschii seedlings. A total of 93 WRKY TFs were identified from A. tauschii, and 79 of them were found to be newly discovered genes compared with wheat. Gene phylogeny, gene structure and chromosome location of the 93 WRKY TFs were fully analyzed. These studies provide a global view of the WRKY TFs from A. tauschii and a firm foundation for further investigations in both A. tauschii and wheat. © 2015 S. Karger AG, Basel.

  12. Genomic localization, sequence analysis, and transcription of the putative human cytomegalovirus DNA polymerase gene

    International Nuclear Information System (INIS)

    Heilbronn, T.; Jahn, G.; Buerkle, A.; Freese, U.K.; Fleckenstein, B.; Zur Hausen, H.

    1987-01-01

    The human cytomegalovirus (HCMV)-induced DNA polymerase has been well characterized biochemically and functionally, but its genomic location has not yet been assigned. To identify the coding sequence, cross-hybridization with the herpes simplex virus type 1 (HSV-1) polymerase gene was used, as suggested by the close similarity of the herpes group virus-induced DNA polymerases to the HCMV DNA polymerase. A cosmid and plasmid library of the entire HCMV genome was screened with the BamHI Q fragment of HSF-1 at different stringency conditions. One PstI-HincII restriction fragment of 850 base pairs mapping within the EcoRI M fragment of HCMV cross-hybridized at T/sub m/ - 25/degrees/C. Sequence analysis revealed one open reading frame spanning the entire sequence. The amino acid sequence showed a highly conserved domain of 133 amino acids shared with the HSV and putative Esptein-Barr virus polymerase sequences. This domain maps within the C-terminal part of the HSV polymerase gene, which has been suggested to contain part of the catalytic center of the enzyme. Transcription analysis revealed one 5.4-kilobase early transcript in the sense orientation with respect to the open reading frame identified. This transcript appears to code for the 140-kilodalton HCMV polymerase protein

  13. Genome-wide analysis of a Wnt1-regulated transcriptional network implicates neurodegenerative pathways.

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    Wexler, Eric M; Rosen, Ezra; Lu, Daning; Osborn, Gregory E; Martin, Elizabeth; Raybould, Helen; Geschwind, Daniel H

    2011-10-04

    Wnt proteins are critical to mammalian brain development and function. The canonical Wnt signaling pathway involves the stabilization and nuclear translocation of β-catenin; however, Wnt also signals through alternative, noncanonical pathways. To gain a systems-level, genome-wide view of Wnt signaling, we analyzed Wnt1-stimulated changes in gene expression by transcriptional microarray analysis in cultured human neural progenitor (hNP) cells at multiple time points over a 72-hour time course. We observed a widespread oscillatory-like pattern of changes in gene expression, involving components of both the canonical and the noncanonical Wnt signaling pathways. A higher-order, systems-level analysis that combined independent component analysis, waveform analysis, and mutual information-based network construction revealed effects on pathways related to cell death and neurodegenerative disease. Wnt effectors were tightly clustered with presenilin1 (PSEN1) and granulin (GRN), which cause dominantly inherited forms of Alzheimer's disease and frontotemporal dementia (FTD), respectively. We further explored a potential link between Wnt1 and GRN and found that Wnt1 decreased GRN expression by hNPs. Conversely, GRN knockdown increased WNT1 expression, demonstrating that Wnt and GRN reciprocally regulate each other. Finally, we provided in vivo validation of the in vitro findings by analyzing gene expression data from individuals with FTD. These unbiased and genome-wide analyses provide evidence for a connection between Wnt signaling and the transcriptional regulation of neurodegenerative disease genes.

  14. Genome-wide classification and expression analysis of MYB transcription factor families in rice and Arabidopsis

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    2012-01-01

    Background The MYB gene family comprises one of the richest groups of transcription factors in plants. Plant MYB proteins are characterized by a highly conserved MYB DNA-binding domain. MYB proteins are classified into four major groups namely, 1R-MYB, 2R-MYB, 3R-MYB and 4R-MYB based on the number and position of MYB repeats. MYB transcription factors are involved in plant development, secondary metabolism, hormone signal transduction, disease resistance and abiotic stress tolerance. A comparative analysis of MYB family genes in rice and Arabidopsis will help reveal the evolution and function of MYB genes in plants. Results A genome-wide analysis identified at least 155 and 197 MYB genes in rice and Arabidopsis, respectively. Gene structure analysis revealed that MYB family genes possess relatively more number of introns in the middle as compared with C- and N-terminal regions of the predicted genes. Intronless MYB-genes are highly conserved both in rice and Arabidopsis. MYB genes encoding R2R3 repeat MYB proteins retained conserved gene structure with three exons and two introns, whereas genes encoding R1R2R3 repeat containing proteins consist of six exons and five introns. The splicing pattern is similar among R1R2R3 MYB genes in Arabidopsis. In contrast, variation in splicing pattern was observed among R1R2R3 MYB members of rice. Consensus motif analysis of 1kb upstream region (5′ to translation initiation codon) of MYB gene ORFs led to the identification of conserved and over-represented cis-motifs in both rice and Arabidopsis. Real-time quantitative RT-PCR analysis showed that several members of MYBs are up-regulated by various abiotic stresses both in rice and Arabidopsis. Conclusion A comprehensive genome-wide analysis of chromosomal distribution, tandem repeats and phylogenetic relationship of MYB family genes in rice and Arabidopsis suggested their evolution via duplication. Genome-wide comparative analysis of MYB genes and their expression analysis

  15. [Genome-wide identification and analysis of WRKY transcription factors in Medicago truncatula].

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    Song, Hui; Nan, Zhibiao

    2014-02-01

    WRKY gene family plays important roles in plant by involving in transcriptional regulations during various physiologically processes such as development, metabolism and responses to biotic and abiotic stresses. WRKY genes have been identified in various plants. However, only few WRKY genes in Medicago truncatula have been identified with systematic analysis and comparison. In this study, we identified 93 WRKY genes through analyses of M. truncatula genome. These genes include 19 type-I genes, 49 type II genes and 13 type-III genes, and 12 non-regular type genes. All of these genes were characterized through analyses of gene duplication, chromosomal locations, structural diversity, conserved protein motifs and phylogenetic relations. The results showed that 11 times of gene duplication event occurred in WRKY gene family involving 24 genes. WRKY genes, containing 6 gene clusters, are unevenly distributed into chromosome 1 to 6, and there is the purifying selection pressure in WRKY group III genes.

  16. Genome-wide analysis of WRKY transcription factors in Solanum lycopersicum.

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    Huang, Shengxiong; Gao, Yongfeng; Liu, Jikai; Peng, Xiaoli; Niu, Xiangli; Fei, Zhangjun; Cao, Shuqing; Liu, Yongsheng

    2012-06-01

    The WRKY transcription factors have been implicated in multiple biological processes in plants, especially in regulating defense against biotic and abiotic stresses. However, little information is available about the WRKYs in tomato (Solanum lycopersicum). The recent release of the whole-genome sequence of tomato allowed us to perform a genome-wide investigation for tomato WRKY proteins, and to compare these positively identified proteins with their orthologs in model plants, such as Arabidopsis and rice. In the present study, based on the recently released tomato whole-genome sequences, we identified 81 SlWRKY genes that were classified into three main groups, with the second group further divided into five subgroups. Depending on WRKY domains' sequences derived from tomato, Arabidopsis and rice, construction of a phylogenetic tree demonstrated distinct clustering and unique gene expansion of WRKY genes among the three species. Genome mapping analysis revealed that tomato WRKY genes were enriched on several chromosomes, especially on chromosome 5, and 16 % of the family members were tandemly duplicated genes. The tomato WRKYs from each group were shown to share similar motif compositions. Furthermore, tomato WRKY genes showed distinct temporal and spatial expression patterns in different developmental processes and in response to various biotic and abiotic stresses. The expression of 18 selected tomato WRKY genes in response to drought and salt stresses and Pseudomonas syringae invasion, respectively, was validated by quantitative RT-PCR. Our results will provide a platform for functional identification and molecular breeding study of WRKY genes in tomato and probably other Solanaceae plants.

  17. Large-scale analysis of antisense transcription in wheat using the Affymetrix GeneChip Wheat Genome Array

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    Settles Matthew L

    2009-05-01

    Full Text Available Abstract Background Natural antisense transcripts (NATs are transcripts of the opposite DNA strand to the sense-strand either at the same locus (cis-encoded or a different locus (trans-encoded. They can affect gene expression at multiple stages including transcription, RNA processing and transport, and translation. NATs give rise to sense-antisense transcript pairs and the number of these identified has escalated greatly with the availability of DNA sequencing resources and public databases. Traditionally, NATs were identified by the alignment of full-length cDNAs or expressed sequence tags to genome sequences, but an alternative method for large-scale detection of sense-antisense transcript pairs involves the use of microarrays. In this study we developed a novel protocol to assay sense- and antisense-strand transcription on the 55 K Affymetrix GeneChip Wheat Genome Array, which is a 3' in vitro transcription (3'IVT expression array. We selected five different tissue types for assay to enable maximum discovery, and used the 'Chinese Spring' wheat genotype because most of the wheat GeneChip probe sequences were based on its genomic sequence. This study is the first report of using a 3'IVT expression array to discover the expression of natural sense-antisense transcript pairs, and may be considered as proof-of-concept. Results By using alternative target preparation schemes, both the sense- and antisense-strand derived transcripts were labeled and hybridized to the Wheat GeneChip. Quality assurance verified that successful hybridization did occur in the antisense-strand assay. A stringent threshold for positive hybridization was applied, which resulted in the identification of 110 sense-antisense transcript pairs, as well as 80 potentially antisense-specific transcripts. Strand-specific RT-PCR validated the microarray observations, and showed that antisense transcription is likely to be tissue specific. For the annotated sense

  18. Draft genome sequence and transcriptional analysis of Rosellinia necatrix infected with a virulent mycovirus.

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    Shimizu, Takeo; Kanematsu, Satoko; Yaegashi, Hajime

    2018-04-24

    Understanding the molecular mechanisms of pathogenesis is useful in developing effective control methods for fungal diseases. The white root rot fungus Rosellinia necatrix is a soil-borne pathogen that causes serious economic losses in various crops, including fruit trees, worldwide. Here, using next-generation sequencing techniques, we first produced a 44-Mb draft genome sequence of R. necatrix strain W97, an isolate from Japan, in which 12,444 protein-coding genes were predicted. To survey differentially expressed genes (DEGs) associated with the pathogenesis of the fungus, the hypovirulent W97 strain infected with Rosellinia necatrix megabirnavirus 1 (RnMBV1) was used for a comprehensive transcriptome analysis. In total, 545 and 615 genes are up- and down-regulated, respectively, in R. necatrix infected with RnMBV1. Gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses of the DEGs suggested that primary and secondary metabolism would be greatly disturbed in R. necatrix infected with RnMBV1. The genes encoding transcriptional regulators, plant cell wall-degrading enzymes, and toxin production, such as cytochalasin E, were also found in the DEGs. The genetic resources provided in this study will accelerate the discovery of genes associated with pathogenesis and other biological characteristics of R. necatrix, thus contributing to disease control.

  19. Transcriptional and phylogenetic analysis of five complete ambystomatid salamander mitochondrial genomes.

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    Samuels, Amy K; Weisrock, David W; Smith, Jeramiah J; France, Katherine J; Walker, John A; Putta, Srikrishna; Voss, S Randal

    2005-04-11

    We report on a study that extended mitochondrial transcript information from a recent EST project to obtain complete mitochondrial genome sequence for 5 tiger salamander complex species (Ambystoma mexicanum, A. t. tigrinum, A. andersoni, A. californiense, and A. dumerilii). We describe, for the first time, aspects of mitochondrial transcription in a representative amphibian, and then use complete mitochondrial sequence data to examine salamander phylogeny at both deep and shallow levels of evolutionary divergence. The available mitochondrial ESTs for A. mexicanum (N=2481) and A. t. tigrinum (N=1205) provided 92% and 87% coverage of the mitochondrial genome, respectively. Complete mitochondrial sequences for all species were rapidly obtained by using long distance PCR and DNA sequencing. A number of genome structural characteristics (base pair length, base composition, gene number, gene boundaries, codon usage) were highly similar among all species and to other distantly related salamanders. Overall, mitochondrial transcription in Ambystoma approximated the pattern observed in other vertebrates. We inferred from the mapping of ESTs onto mtDNA that transcription occurs from both heavy and light strand promoters and continues around the entire length of the mtDNA, followed by post-transcriptional processing. However, the observation of many short transcripts corresponding to rRNA genes indicates that transcription may often terminate prematurely to bias transcription of rRNA genes; indeed an rRNA transcription termination signal sequence was observed immediately following the 16S rRNA gene. Phylogenetic analyses of salamander family relationships consistently grouped Ambystomatidae in a clade containing Cryptobranchidae and Hynobiidae, to the exclusion of Salamandridae. This robust result suggests a novel alternative hypothesis because previous studies have consistently identified Ambystomatidae and Salamandridae as closely related taxa. Phylogenetic analyses of tiger

  20. Construction of a genomic library of the human cytomegalovirus genome and analysis of late transcription of its inverted internal repeat region

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    Silva, K.F.S.T.

    1989-01-01

    The investigations described in this dissertation were designed to determine the transcriptionally active DNA sequences of IIR region and to identify the viral mRNA transcribed from the transcriptionally most active DNA sequences of that region during late phase of HCMV Towne infection. Preliminary transcriptional studies which included the hybridization of a southern blot of XbaI digested entire HCMV genome to 32 P-labelled late phase infected cell A + RNA, indicated that late viral transcripts homologous to XbaI Q fragment of IIR region were very highly abundant while XbaI Q fragment showed a very low transcriptional activity. To facilitate further analysis of late transcription of IIR region, the entire DNA sequences of IIR region were molecularly cloned as U, S, and H BamHI fragments in pACYC-184 plasmid vector. In addition, to be used in future studies on other regions of the genome, except for y and c' smaller fragments the entire 240 kb HCMV genome was cloned as BamHI fragments in the same vector. Furthermore, the U, S, and H BamHI fragments were mapped with six other restriction enzymes in order to use that mapping data in subsequent transcriptional analysis of the IIR region. Further localization of transcriptionally active DNA sequences within IIR region was achieved by hybridization of southern blots of restricted U, S, and H BamHI fragments with 3' 32 P-labelled infected cell late A + RNA. The 1.5 kb EcooRI subfragments of S BamHI fragment and the adjoining 0.72 kb XhoI subfragment of H BamHI fragment revealed the highest level of transcription, although the remainder of the S fragment was also transcribed at a substantial level. The U fragment and the remainder of the H fragment was transcribed at a very low level

  1. Genome-wide analysis of EgEVE_1, a transcriptionally active endogenous viral element associated to small RNAs in Eucalyptus genomes

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    Helena Sanches Marcon

    2017-02-01

    Full Text Available Abstract Endogenous viral elements (EVEs are the result of heritable horizontal gene transfer from viruses to hosts. In the last years, several EVE integration events were reported in plants by the exponential availability of sequenced genomes. Eucalyptus grandis is a forest tree species with a sequenced genome that is poorly studied in terms of evolution and mobile genetic elements composition. Here we report the characterization of E. grandis endogenous viral element 1 (EgEVE_1, a transcriptionally active EVE with a size of 5,664 bp. Phylogenetic analysis and genomic distribution demonstrated that EgEVE_1 is a newly described member of the Caulimoviridae family, distinct from the recently characterized plant Florendoviruses. Genomic distribution of EgEVE_1 and Florendovirus is also distinct. EgEVE_1 qPCR quantification in Eucalyptus urophylla suggests that this genome has more EgEVE_1 copies than E. grandis. EgEVE_1 transcriptional activity was demonstrated by RT-qPCR in five Eucalyptus species and one intrageneric hybrid. We also identified that Eucalyptus EVEs can generate small RNAs (sRNAs,that might be involved in de novo DNA methylation and virus resistance. Our data suggest that EVE families in Eucalyptus have distinct properties, and we provide the first comparative analysis of EVEs in Eucalyptus genomes.

  2. Whole-genome transcriptional analysis of Escherichia coli during heat inactivation processes related to industrial cooking.

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    Guernec, A; Robichaud-Rincon, P; Saucier, L

    2013-08-01

    Escherichia coli K-12 was grown to the stationary phase, for maximum physiological resistance, in brain heart infusion (BHI) broth at 37°C. Cells were then heated at 58°C or 60°C to reach a process lethality value \\[\\mathbf{\\left(}{{\\mathit{F}}^{\\mathit{o}}}_{\\mathbf{70}}^{\\mathbf{10}}\\mathbf{\\right)} \\] of 2 or 3 or to a core temperature of 71°C (control industrial cooking temperature). Growth recovery and cell membrane integrity were evaluated immediately after heating, and a global transcription analysis was performed using gene expression microarrays. Only cells heated at 58°C with F(o) = 2 were still able to grow on liquid or solid BHI broth after heat treatment. However, their transcriptome did not differ from that of bacteria heated at 58°C with F(o) = 3 (P value for the false discovery rate [P-FDR] > 0.01), where no growth recovery was observed posttreatment. Genome-wide transcriptomic data obtained at 71°C were distinct from those of the other treatments without growth recovery. Quantification of heat shock gene expression by real-time PCR revealed that dnaK and groEL mRNA levels decreased significantly above 60°C to reach levels similar to those of control cells at 37°C (P citE, glyS, oppB, and asd, whose expression was upregulated at 71°C, may be worth investigating as good biomarkers for accurately determining the efficiency of heat treatments, especially when cells are too injured to be enumerated using growth media.

  3. Whole-genome transcriptional analysis of heavy metal stresses inCaulobacter crescentus

    Energy Technology Data Exchange (ETDEWEB)

    Hu, Ping; Brodie, Eoin L.; Suzuki, Yohey; McAdams, Harley H.; Andersen, Gary L.

    2005-09-21

    The bacterium Caulobacter crescentus and related stalkbacterial species are known for their distinctive ability to live in lownutrient environments, a characteristic of most heavy metal contaminatedsites. Caulobacter crescentus is a model organism for studying cell cycleregulation with well developed genetics. We have identified the pathwaysresponding to heavy metal toxicity in C. crescentus to provide insightsfor possible application of Caulobacter to environmental restoration. Weexposed C. crescentus cells to four heavy metals (chromium, cadmium,selenium and uranium) and analyzed genome wide transcriptional activitiespost exposure using a Affymetrix GeneChip microarray. C. crescentusshowed surprisingly high tolerance to uranium, a possible mechanism forwhich may be formation of extracellular calcium-uranium-phosphateprecipitates. The principal response to these metals was protectionagainst oxidative stress (up-regulation of manganese-dependent superoxidedismutase, sodA). Glutathione S-transferase, thioredoxin, glutaredoxinsand DNA repair enzymes responded most strongly to cadmium and chromate.The cadmium and chromium stress response also focused on reducing theintracellular metal concentration, with multiple efflux pumps employed toremove cadmium while a sulfate transporter was down-regulated to reducenon-specific uptake of chromium. Membrane proteins were also up-regulatedin response to most of the metals tested. A two-component signaltransduction system involved in the uranium response was identified.Several differentially regulated transcripts from regions previously notknown to encode proteins were identified, demonstrating the advantage ofevaluating the transcriptome using whole genome microarrays.

  4. Genome-wide cloning, identification, classification and functional analysis of cotton heat shock transcription factors in cotton (Gossypium hirsutum).

    Science.gov (United States)

    Wang, Jun; Sun, Na; Deng, Ting; Zhang, Lida; Zuo, Kaijing

    2014-11-06

    Heat shock transcriptional factors (Hsfs) play important roles in the processes of biotic and abiotic stresses as well as in plant development. Cotton (Gossypium hirsutum, 2n=4x=(AD)2=52) is an important crop for natural fiber production. Due to continuous high temperature and intermittent drought, heat stress is becoming a handicap to improve cotton yield and lint quality. Recently, the related wild diploid species Gossypium raimondii genome (2n=2x=(D5)2=26) has been fully sequenced. In order to analyze the functions of different Hsfs at the genome-wide level, detailed characterization and analysis of the Hsf gene family in G. hirsutum is indispensable. EST assembly and genome-wide analyses were applied to clone and identify heat shock transcription factor (Hsf) genes in Upland cotton (GhHsf). Forty GhHsf genes were cloned, identified and classified into three main classes (A, B and C) according to the characteristics of their domains. Analysis of gene duplications showed that GhHsfs have occurred more frequently than reported in plant genomes such as Arabidopsis and Populus. Quantitative real-time PCR (qRT-PCR) showed that all GhHsf transcripts are expressed in most cotton plant tissues including roots, stems, leaves and developing fibers, and abundantly in developing ovules. Three expression patterns were confirmed in GhHsfs when cotton plants were exposed to high temperature for 1 h. GhHsf39 exhibited the most immediate response to heat shock. Comparative analysis of Hsfs expression differences between the wild-type and fiberless mutant suggested that Hsfs are involved in fiber development. Comparative genome analysis showed that Upland cotton D-subgenome contains 40 Hsf members, and that the whole genome of Upland cotton contains more than 80 Hsf genes due to genome duplication. The expression patterns in different tissues in response to heat shock showed that GhHsfs are important for heat stress as well as fiber development. These results provide an improved

  5. The prophages of Lactobacillus johnsonii NCC 533: comparative genomics and transcription analysis

    International Nuclear Information System (INIS)

    Ventura, Marco; Canchaya, Carlos; Pridmore, R. David; Bruessow, Harald

    2004-01-01

    Two non-inducible, but apparently complete prophages were identified in the genome of the sequenced Lactobacillus johnsonii strain NCC 533. The 38- and 40-kb-long prophages Lj928 and Lj965 represent distinct lineages of Sfi11-like pac-site Siphoviridae unrelated at the DNA sequence level. The deduced structural proteins from Lj928 demonstrated aa sequence identity with Lactococcus lactis phage TP901-1, while Lj965 shared sequence links with Streptococcus thermophilus phage O1205. With the exception of tRNA genes, inserted between DNA replication and DNA packaging genes, the transcription of the prophage was restricted to the genome segments near both attachment sites. Transcribed genes unrelated to phage functions were inserted between the phage repressor and integrase genes; one group of genes shared sequence relatedness with a mobile DNA element in Staphylococcus aureus. A short, but highly transcribed region was located between the phage lysin and right attachment site; it lacked a protein-encoding function in one prophage

  6. Genomic identification of WRKY transcription factors in carrot (Daucus carota) and analysis of evolution and homologous groups for plants.

    Science.gov (United States)

    Li, Meng-Yao; Xu, Zhi-Sheng; Tian, Chang; Huang, Ying; Wang, Feng; Xiong, Ai-Sheng

    2016-03-15

    WRKY transcription factors belong to one of the largest transcription factor families. These factors possess functions in plant growth and development, signal transduction, and stress response. Here, we identified 95 DcWRKY genes in carrot based on the carrot genomic and transcriptomic data, and divided them into three groups. Phylogenetic analysis of WRKY proteins from carrot and Arabidopsis divided these proteins into seven subgroups. To elucidate the evolution and distribution of WRKY transcription factors in different species, we constructed a schematic of the phylogenetic tree and compared the WRKY family factors among 22 species, which including plants, slime mold and protozoan. An in-depth study was performed to clarify the homologous factor groups of nine divergent taxa in lower and higher plants. Based on the orthologous factors between carrot and Arabidopsis, 38 DcWRKY proteins were calculated to interact with other proteins in the carrot genome. Yeast two-hybrid assay showed that DcWRKY20 can interact with DcMAPK1 and DcMAPK4. The expression patterns of the selected DcWRKY genes based on transcriptome data and qRT-PCR suggested that those selected DcWRKY genes are involved in root development, biotic and abiotic stress response. This comprehensive analysis provides a basis for investigating the evolution and function of WRKY genes.

  7. Genome-Wide Identification and Structural Analysis of bZIP Transcription Factor Genes in Brassica napus.

    Science.gov (United States)

    Zhou, Yan; Xu, Daixiang; Jia, Ledong; Huang, Xiaohu; Ma, Guoqiang; Wang, Shuxian; Zhu, Meichen; Zhang, Aoxiang; Guan, Mingwei; Lu, Kun; Xu, Xinfu; Wang, Rui; Li, Jiana; Qu, Cunmin

    2017-10-24

    The basic region/leucine zipper motif (bZIP) transcription factor family is one of the largest families of transcriptional regulators in plants. bZIP genes have been systematically characterized in some plants, but not in rapeseed ( Brassica napus ). In this study, we identified 247 BnbZIP genes in the rapeseed genome, which we classified into 10 subfamilies based on phylogenetic analysis of their deduced protein sequences. The BnbZIP genes were grouped into functional clades with Arabidopsis genes with similar putative functions, indicating functional conservation. Genome mapping analysis revealed that the BnbZIPs are distributed unevenly across all 19 chromosomes, and that some of these genes arose through whole-genome duplication and dispersed duplication events. All expression profiles of 247 bZIP genes were extracted from RNA-sequencing data obtained from 17 different B . napus ZS11 tissues with 42 various developmental stages. These genes exhibited different expression patterns in various tissues, revealing that these genes are differentially regulated. Our results provide a valuable foundation for functional dissection of the different BnbZIP homologs in B . napus and its parental lines and for molecular breeding studies of bZIP genes in B . napus .

  8. Genome-wide screening and transcriptional profile analysis of desaturase genes in the European corn borer moth

    Institute of Scientific and Technical Information of China (English)

    Bingye Xue; Alejandro P. Rooney; Wendell L. Roelofs

    2012-01-01

    Acyl-coenzyme A (Acyl-CoA) desaturases play a key role in the biosynthesis of female moth sex pheromones.Desaturase genes are encoded by a large multigene family,and they have been divided into five subgroups on the basis of biochemical functionality and phylogenetic affinity.In this study both copy numbers and transcriptional levels of desaturase genes in the European corn borer (ECB),Ostrinia nubilalis,were investigated.The results from genome-wide screening of ECB bacterial artificial chromosome (BAC)library indicated there are many copies of some desaturase genes in the genome.An open reading frame (ORF) has been isolated for the novel desaturase gene ECB ezi-△11β from ECB gland complementary DNA and its functionality has been analyzed by two yeast expression systems.No functional activities have been detected for it.The expression levels of the four desaturase genes both in the pheromone gland and fat body of ECB and Asian corn borer (ACB),O.furnacalis,were determined by real-time polymerase chain reaction.In the ECB gland,△ 11 is the most abundant,although the amount of △14 is also considerable.In the ACB gland,△14 is the most abundant and is 100 times more abundant than all the other three combined.The results from the analysis of evolution of desaturase gene transcription in the ECB,ACB and other moths indicate that the pattern of △ 11 gene transcription is significantly different from the transcriptional patterns of other desaturase genes and this difference is tied to the underlying nucleotide composition bias of the genome.

  9. Genome-wide DNA methylation patterns and transcription analysis in sheep muscle.

    Directory of Open Access Journals (Sweden)

    Christine Couldrey

    Full Text Available DNA methylation plays a central role in regulating many aspects of growth and development in mammals through regulating gene expression. The development of next generation sequencing technologies have paved the way for genome-wide, high resolution analysis of DNA methylation landscapes using methodology known as reduced representation bisulfite sequencing (RRBS. While RRBS has proven to be effective in understanding DNA methylation landscapes in humans, mice, and rats, to date, few studies have utilised this powerful method for investigating DNA methylation in agricultural animals. Here we describe the utilisation of RRBS to investigate DNA methylation in sheep Longissimus dorsi muscles. RRBS analysis of ∼1% of the genome from Longissimus dorsi muscles provided data of suitably high precision and accuracy for DNA methylation analysis, at all levels of resolution from genome-wide to individual nucleotides. Combining RRBS data with mRNAseq data allowed the sheep Longissimus dorsi muscle methylome to be compared with methylomes from other species. While some species differences were identified, many similarities were observed between DNA methylation patterns in sheep and other more commonly studied species. The RRBS data presented here highlights the complexity of epigenetic regulation of genes. However, the similarities observed across species are promising, in that knowledge gained from epigenetic studies in human and mice may be applied, with caution, to agricultural species. The ability to accurately measure DNA methylation in agricultural animals will contribute an additional layer of information to the genetic analyses currently being used to maximise production gains in these species.

  10. Comparative analysis of regulatory elements between Escherichia coli and Klebsiella pneumoniae by genome-wide transcription start site profiling.

    Directory of Open Access Journals (Sweden)

    Donghyuk Kim

    Full Text Available Genome-wide transcription start site (TSS profiles of the enterobacteria Escherichia coli and Klebsiella pneumoniae were experimentally determined through modified 5' RACE followed by deep sequencing of intact primary mRNA. This identified 3,746 and 3,143 TSSs for E. coli and K. pneumoniae, respectively. Experimentally determined TSSs were then used to define promoter regions and 5' UTRs upstream of coding genes. Comparative analysis of these regulatory elements revealed the use of multiple TSSs, identical sequence motifs of promoter and Shine-Dalgarno sequence, reflecting conserved gene expression apparatuses between the two species. In both species, over 70% of primary transcripts were expressed from operons having orthologous genes during exponential growth. However, expressed orthologous genes in E. coli and K. pneumoniae showed a strikingly different organization of upstream regulatory regions with only 20% identical promoters with TSSs in both species. Over 40% of promoters had TSSs identified in only one species, despite conserved promoter sequences existing in the other species. 662 conserved promoters having TSSs in both species resulted in the same number of comparable 5' UTR pairs, and that regulatory element was found to be the most variant region in sequence among promoter, 5' UTR, and ORF. In K. pneumoniae, 48 sRNAs were predicted and 36 of them were expressed during exponential growth. Among them, 34 orthologous sRNAs between two species were analyzed in depth, and the analysis showed that many sRNAs of K. pneumoniae, including pleiotropic sRNAs such as rprA, arcZ, and sgrS, may work in the same way as in E. coli. These results reveal a new dimension of comparative genomics such that a comparison of two genomes needs to be comprehensive over all levels of genome organization.

  11. Papillomavirus genomes in human cervical carcinoma: Analysis of their integration and transcriptional activity

    International Nuclear Information System (INIS)

    Matulic, M.; Soric, J.

    1994-01-01

    Eighty-four biopsies derived from cervical tissues were analyzed for the presence of human papillomavirus (HPV) DNA types 6, 16 and 18 using Southern blot hybridization. HPV 6 was found in none of the cervical biopsies, and HPV types 16 and 18 were found in 44% of them. The rate of HPV 16/18 positive samples increased proportionally to the severity of the lesion. In normal tissue there were no positive samples, in mild and moderate dysplasia HPV 16/18 was present in 20% and in severe dysplasia and invasive carcinomas in 37 and 50%, respectively. In biopsies from 13 cases with squamous cell carcinoma of the uterine cervix and CIN III lesions HPV 16 was integrated within the host genome. It was concluded that the virus could be integrated at variable, presumably randomly selected chromosomal loci and with different number of copies. Transcription of HPV 16 and 18 was detected in one cervical cancer in HeLa cells, respectively. These results imply that HPV types 16 and 18 play an etiological role in the carcinogenesis of human cervical epithelial cells. (author)

  12. Genomic organization, transcript variants and comparative analysis of the human nucleoporin 155 (NUP155) gene

    DEFF Research Database (Denmark)

    Zhang, X.; Yang, J.; Yu, J.

    2002-01-01

    Nucleoporin 155 (Nup155) is a major component of the nuclear pore complex (NPC) involved in cellular nucleo-cytoplasmic transport. We have acquired the complete sequence and interpreted the genomic organization of the Nup155 orthologos from human (Homo sapiens) and pufferfish (Fugu rubripes), which...... complementary to RNAs of the Nup155 orthologs from Fugu and mouse. Comparative analysis of the Nup155 orthologs in many species, including H. sapiens, Mus musculus, Rattus norvegicus, F. rubripes, Arabidopsis thaliana, Drosophila melanogaster, and Saccharomyces cerevisiae, has revealed two paralogs in S...

  13. Genome-Wide Phylogenetic Comparative Analysis of Plant Transcriptional Regulation: A Timeline of Loss, Gain, Expansion, and Correlation with Complexity

    OpenAIRE

    Lang, Daniel; Weiche, Benjamin; Timmerhaus, Gerrit; Richardt, Sandra; Ria?o-Pach?n, Diego M.; Corr?a, Luiz G. G.; Reski, Ralf; Mueller-Roeber, Bernd; Rensing, Stefan A.

    2010-01-01

    Evolutionary retention of duplicated genes encoding transcription-associated proteins (TAPs, comprising transcription factors and other transcriptional regulators) has been hypothesized to be positively correlated with increasing morphological complexity and paleopolyploidizations, especially within the plant kingdom. Here, we present the most comprehensive set of classification rules for TAPs and its application for genome-wide analyses of plants and algae. Using a dated species tree and phy...

  14. Genome-wide analysis of differential transcriptional and epigenetic variability across human immune cell types

    DEFF Research Database (Denmark)

    Ecker, Simone; Chen, Lu; Pancaldi, Vera

    2017-01-01

    Background: A healthy immune system requires immune cells that adapt rapidly to environmental challenges. This phenotypic plasticity can be mediated by transcriptional and epigenetic variability. Results: We apply a novel analytical approach to measure and compare transcriptional and epigenetic v...

  15. Genome-Wide Identification and Expression Analysis of WRKY Transcription Factors under Multiple Stresses in Brassica napus.

    Science.gov (United States)

    He, Yajun; Mao, Shaoshuai; Gao, Yulong; Zhu, Liying; Wu, Daoming; Cui, Yixin; Li, Jiana; Qian, Wei

    2016-01-01

    WRKY transcription factors play important roles in responses to environmental stress stimuli. Using a genome-wide domain analysis, we identified 287 WRKY genes with 343 WRKY domains in the sequenced genome of Brassica napus, 139 in the A sub-genome and 148 in the C sub-genome. These genes were classified into eight groups based on phylogenetic analysis. In the 343 WRKY domains, a total of 26 members showed divergence in the WRKY domain, and 21 belonged to group I. This finding suggested that WRKY genes in group I are more active and variable compared with genes in other groups. Using genome-wide identification and analysis of the WRKY gene family in Brassica napus, we observed genome duplication, chromosomal/segmental duplications and tandem duplication. All of these duplications contributed to the expansion of the WRKY gene family. The duplicate segments that were detected indicated that genome duplication events occurred in the two diploid progenitors B. rapa and B. olearecea before they combined to form B. napus. Analysis of the public microarray database and EST database for B. napus indicated that 74 WRKY genes were induced or preferentially expressed under stress conditions. According to the public QTL data, we identified 77 WRKY genes in 31 QTL regions related to various stress tolerance. We further evaluated the expression of 26 BnaWRKY genes under multiple stresses by qRT-PCR. Most of the genes were induced by low temperature, salinity and drought stress, indicating that the WRKYs play important roles in B. napus stress responses. Further, three BnaWRKY genes were strongly responsive to the three multiple stresses simultaneously, which suggests that these 3 WRKY may have multi-functional roles in stress tolerance and can potentially be used in breeding new rapeseed cultivars. We also found six tandem repeat pairs exhibiting similar expression profiles under the various stress conditions, and three pairs were mapped in the stress related QTL regions

  16. Genome-Wide Identification and Expression Analysis of WRKY Transcription Factors under Multiple Stresses in Brassica napus.

    Directory of Open Access Journals (Sweden)

    Yajun He

    Full Text Available WRKY transcription factors play important roles in responses to environmental stress stimuli. Using a genome-wide domain analysis, we identified 287 WRKY genes with 343 WRKY domains in the sequenced genome of Brassica napus, 139 in the A sub-genome and 148 in the C sub-genome. These genes were classified into eight groups based on phylogenetic analysis. In the 343 WRKY domains, a total of 26 members showed divergence in the WRKY domain, and 21 belonged to group I. This finding suggested that WRKY genes in group I are more active and variable compared with genes in other groups. Using genome-wide identification and analysis of the WRKY gene family in Brassica napus, we observed genome duplication, chromosomal/segmental duplications and tandem duplication. All of these duplications contributed to the expansion of the WRKY gene family. The duplicate segments that were detected indicated that genome duplication events occurred in the two diploid progenitors B. rapa and B. olearecea before they combined to form B. napus. Analysis of the public microarray database and EST database for B. napus indicated that 74 WRKY genes were induced or preferentially expressed under stress conditions. According to the public QTL data, we identified 77 WRKY genes in 31 QTL regions related to various stress tolerance. We further evaluated the expression of 26 BnaWRKY genes under multiple stresses by qRT-PCR. Most of the genes were induced by low temperature, salinity and drought stress, indicating that the WRKYs play important roles in B. napus stress responses. Further, three BnaWRKY genes were strongly responsive to the three multiple stresses simultaneously, which suggests that these 3 WRKY may have multi-functional roles in stress tolerance and can potentially be used in breeding new rapeseed cultivars. We also found six tandem repeat pairs exhibiting similar expression profiles under the various stress conditions, and three pairs were mapped in the stress related

  17. Genome-wide analysis of WRKY transcription factors in white pear (Pyrus bretschneideri) reveals evolution and patterns under drought stress.

    Science.gov (United States)

    Huang, Xiaosan; Li, Kongqing; Xu, Xiaoyong; Yao, Zhenghong; Jin, Cong; Zhang, Shaoling

    2015-12-24

    WRKY transcription factors (TFs) constitute one of the largest protein families in higher plants, and its members contain one or two conserved WRKY domains, about 60 amino acid residues with the WRKYGQK sequence followed by a C2H2 or C2HC zinc finger motif. WRKY proteins play significant roles in plant development, and in responses to biotic and abiotic stresses. Pear (Pyrus bretschneideri) is one of the most important fruit crops in the world and is frequently threatened by abiotic stress, such as drought, affecting growth, development and productivity. Although the pear genome sequence has been released, little is known about the WRKY TFs in pear, especially in respond to drought stress at the genome-wide level. We identified a total of 103 WRKY TFs in the pear genome. Based on the structural features of WRKY proteins and topology of the phylogenetic tree, the pear WRKY (PbWRKY) family was classified into seven groups (Groups 1, 2a-e, and 3). The microsyteny analysis indicated that 33 (32%) PbWRKY genes were tandemly duplicated and 57 genes (55.3%) were segmentally duplicated. RNA-seq experiment data and quantitative real-time reverse transcription PCR revealed that PbWRKY genes in different groups were induced by drought stress, and Group 2a and 3 were mainly involved in the biological pathways in response to drought stress. Furthermore, adaptive evolution analysis detected a significant positive selection for Pbr001425 in Group 3, and its expression pattern differed from that of other members in this group. The present study provides a solid foundation for further functional dissection and molecular evolution of WRKY TFs in pear, especially for improving the water-deficient resistance of pear through manipulation of the PbWRKYs.

  18. Transcriptional Analysis Allows Genome Reannotation and Reveals that Cryptococcus gattii VGII Undergoes Nutrient Restriction during Infection

    Directory of Open Access Journals (Sweden)

    Patrícia Aline Gröhs Ferrareze

    2017-08-01

    Full Text Available Cryptococcus gattii is a human and animal pathogen that infects healthy hosts and caused the Pacific Northwest outbreak of cryptococcosis. The inhalation of infectious propagules can lead to internalization of cryptococcal cells by alveolar macrophages, a niche in which C. gattii cells can survive and proliferate. Although the nutrient composition of macrophages is relatively unknown, the high induction of amino acid transporter genes inside the phagosome indicates a preference for amino acid uptake instead of synthesis. However, the presence of countable errors in the R265 genome annotation indicates significant inhibition of transcriptomic analysis in this hypervirulent strain. Thus, we analyzed RNA-Seq data from in vivo and in vitro cultures of C. gattii R265 to perform the reannotation of the genome. In addition, based on in vivo transcriptomic data, we identified highly expressed genes and pathways of amino acid metabolism that would enable C. gattii to survive and proliferate in vivo. Importantly, we identified high expression in three APC amino acid transporters as well as the GABA permease. The use of amino acids as carbon and nitrogen sources, releasing ammonium and generating carbohydrate metabolism intermediaries, also explains the high expression of components of several degradative pathways, since glucose starvation is an important host defense mechanism.

  19. Genome-wide analysis and expression profiling of the ERF transcription factor family in potato (Solanum tuberosum L.).

    Science.gov (United States)

    Charfeddine, Mariam; Saïdi, Mohamed Najib; Charfeddine, Safa; Hammami, Asma; Gargouri Bouzid, Radhia

    2015-04-01

    The ERF transcription factors belong to the AP2/ERF superfamily, one of the largest transcription factor families in plants. They play important roles in plant development processes, as well as in the response to biotic, abiotic, and hormone signaling. In the present study, 155 putative ERF transcription factor genes were identified from the potato (Solanum tuberosum) genome database, and compared with those from Arabidopsis thaliana. The StERF proteins are divided into ten phylogenetic groups. Expression analyses of five StERFs were carried out by semi-quantitative RT-PCR and compared with published RNA-seq data. These latter analyses were used to distinguish tissue-specific, biotic, and abiotic stress genes as well as hormone-responsive StERF genes. The results are of interest to better understand the role of the AP2/ERF genes in response to diverse types of stress in potatoes. A comprehensive analysis of the physiological functions and biological roles of the ERF family genes in S. tuberosum is required to understand crop stress tolerance mechanisms.

  20. Genome-wide transcriptional analysis of two soybean genotypes under dehydration and rehydration conditions

    Science.gov (United States)

    2013-01-01

    Background Soybean is an important crop that provides valuable proteins and oils for human use. Because soybean growth and development is extremely sensitive to water deficit, quality and crop yields are severely impacted by drought stress. In the face of limited water resources, drought-responsive genes are therefore of interest. Identification and analysis of dehydration- and rehydration-inducible differentially expressed genes (DEGs) would not only aid elucidation of molecular mechanisms of stress response, but also enable improvement of crop stress tolerance via gene transfer. Using Digital Gene Expression Tag profiling (DGE), a new technique based on Illumina sequencing, we analyzed expression profiles between two soybean genotypes to identify drought-responsive genes. Results Two soybean genotypes—drought-tolerant Jindou21 and drought-sensitive Zhongdou33—were subjected to dehydration and rehydration conditions. For analysis of DEGs under dehydration conditions, 20 cDNA libraries were generated from roots and leaves at two different time points under well-watered and dehydration conditions. We also generated eight libraries for analysis under rehydration conditions. Sequencing of the 28 libraries produced 25,000–33,000 unambiguous tags, which were mapped to reference sequences for annotation of expressed genes. Many genes exhibited significant expression differences among the libraries. DEGs in the drought-tolerant genotype were identified by comparison of DEGs among treatments and genotypes. In Jindou21, 518 and 614 genes were differentially expressed under dehydration in leaves and roots, respectively, with 24 identified both in leaves and roots. The main functional categories enriched in these DEGs were metabolic process, response to stresses, plant hormone signal transduction, protein processing, and plant-pathogen interaction pathway; the associated genes primarily encoded transcription factors, protein kinases, and other regulatory proteins. The

  1. Genome-wide analysis of transcription factors during somatic embryogenesis in banana (Musa spp.) cv. Grand Naine.

    Science.gov (United States)

    Shivani; Awasthi, Praveen; Sharma, Vikrant; Kaur, Navjot; Kaur, Navneet; Pandey, Pankaj; Tiwari, Siddharth

    2017-01-01

    Transcription factors BABY BOOM (BBM), WUSCHEL (WUS), BSD, LEAFY COTYLEDON (LEC), LEAFY COTYLEDON LIKE (LIL), VIVIPAROUS1 (VP1), CUP SHAPED COTYLEDONS (CUC), BOLITA (BOL), and AGAMOUS LIKE (AGL) play a crucial role in somatic embryogenesis. In this study, we identified eighteen genes of these nine transcription factors families from the banana genome database. All genes were analyzed for their structural features, subcellular, and chromosomal localization. Protein sequence analysis indicated the presence of characteristic conserved domains in these transcription factors. Phylogenetic analysis revealed close evolutionary relationship among most transcription factors of various monocots. The expression patterns of eighteen genes in embryogenic callus containing somatic embryos (precisely isolated by Laser Capture Microdissection), non-embryogenic callus, and cell suspension cultures of banana cultivar Grand Naine were analyzed. The application of 2, 4-dichlorophenoxyacetic acid (2, 4-D) in the callus induction medium enhanced the expression of MaBBM1, MaBBM2, MaWUS2, and MaVP1 in the embryogenic callus. It suggested 2, 4-D acts as an inducer for the expression of these genes. The higher expression of MaBBM2 and MaWUS2 in embryogenic cell suspension (ECS) as compared to non-embryogenic cells suspension (NECS), suggested that these genes may play a crucial role in banana somatic embryogenesis. MaVP1 showed higher expression in both ECS and NECS, whereas MaLEC2 expression was significantly higher in NECS. It suggests that MaLEC2 has a role in the development of non-embryogenic cells. We postulate that MaBBM2 and MaWUS2 can be served as promising molecular markers for the embryogencity in banana.

  2. Genome-wide analysis of the Dof transcription factor gene family reveals soybean-specific duplicable and functional characteristics.

    Directory of Open Access Journals (Sweden)

    Yong Guo

    Full Text Available The Dof domain protein family is a classic plant-specific zinc-finger transcription factor family involved in a variety of biological processes. There is great diversity in the number of Dof genes in different plants. However, there are only very limited reports on the characterization of Dof transcription factors in soybean (Glycine max. In the present study, 78 putative Dof genes were identified from the whole-genome sequence of soybean. The predicted GmDof genes were non-randomly distributed within and across 19 out of 20 chromosomes and 97.4% (38 pairs were preferentially retained duplicate paralogous genes located in duplicated regions of the genome. Soybean-specific segmental duplications contributed significantly to the expansion of the soybean Dof gene family. These Dof proteins were phylogenetically clustered into nine distinct subgroups among which the gene structure and motif compositions were considerably conserved. Comparative phylogenetic analysis of these Dof proteins revealed four major groups, similar to those reported for Arabidopsis and rice. Most of the GmDofs showed specific expression patterns based on RNA-seq data analyses. The expression patterns of some duplicate genes were partially redundant while others showed functional diversity, suggesting the occurrence of sub-functionalization during subsequent evolution. Comprehensive expression profile analysis also provided insights into the soybean-specific functional divergence among members of the Dof gene family. Cis-regulatory element analysis of these GmDof genes suggested diverse functions associated with different processes. Taken together, our results provide useful information for the functional characterization of soybean Dof genes by combining phylogenetic analysis with global gene-expression profiling.

  3. Genome-wide identification and analysis of the B3 superfamily of transcription factors in Brassicaceae and major crop plants.

    Science.gov (United States)

    Peng, Fred Y; Weselake, Randall J

    2013-05-01

    The plant-specific B3 superfamily of transcription factors has diverse functions in plant growth and development. Using a genome-wide domain analysis, we identified 92, 187, 58, 90, 81, 55, and 77 B3 transcription factor genes in the sequenced genome of Arabidopsis, Brassica rapa, castor bean (Ricinus communis), cocoa (Theobroma cacao), soybean (Glycine max), maize (Zea mays), and rice (Oryza sativa), respectively. The B3 superfamily has substantially expanded during the evolution in eudicots particularly in Brassicaceae, as compared to monocots in the analysis. We observed domain duplication in some of these B3 proteins, forming more complex domain architectures than currently understood. We found that the length of B3 domains exhibits a large variation, which may affect their exact number of α-helices and β-sheets in the core structure of B3 domains, and possibly have functional implications. Analysis of the public microarray data indicated that most of the B3 gene pairs encoding Arabidopsis-rice orthologs are preferentially expressed in different tissues, suggesting their different roles in these two species. Using ESTs in crops, we identified many B3 genes preferentially expressed in reproductive tissues. In a sequence-based quantitative trait loci analysis in rice and maize, we have found many B3 genes associated with traits such as grain yield, seed weight and number, and protein content. Our results provide a framework for future studies into the function of B3 genes in different phases of plant development, especially the ones related to traits in major crops.

  4. Comparative genomic analysis of pathogenic and probiotic Enterococcus faecalis isolates, and their transcriptional responses to growth in human urine.

    Directory of Open Access Journals (Sweden)

    Heidi C Vebø

    Full Text Available Urinary tract infection (UTI is the most common infection caused by enterococci, and Enterococcus faecalis accounts for the majority of enterococcal infections. Although a number of virulence related traits have been established, no comprehensive genomic or transcriptomic studies have been conducted to investigate how to distinguish pathogenic from non-pathogenic E. faecalis in their ability to cause UTI. In order to identify potential genetic traits or gene regulatory features that distinguish pathogenic from non-pathogenic E. faecalis with respect to UTI, we have performed comparative genomic analysis, and investigated growth capacity and transcriptome profiling in human urine in vitro. Six strains of different origins were cultivated and all grew readily in human urine. The three strains chosen for transcriptional analysis showed an overall similar response with respect to energy and nitrogen metabolism, stress mechanism, cell envelope modifications, and trace metal acquisition. Our results suggest that citrate and aspartate are significant for growth of E. faecalis in human urine, and manganese appear to be a limiting factor. The majority of virulence factors were either not differentially regulated or down-regulated. Notably, a significant up-regulation of genes involved in biofilm formation was observed. Strains from different origins have similar capacity to grow in human urine. The overall similar transcriptional responses between the two pathogenic and the probiotic strain suggest that the pathogenic potential of a certain E. faecalis strain may to a great extent be determined by presence of fitness and virulence factors, rather than the level of expression of such traits.

  5. Whole genome transcription profiling of Anaplasma phagocytophilum in human and tick host cells by tiling array analysis

    Directory of Open Access Journals (Sweden)

    Chavez Adela

    2008-07-01

    Full Text Available Abstract Background Anaplasma phagocytophilum (Ap is an obligate intracellular bacterium and the agent of human granulocytic anaplasmosis, an emerging tick-borne disease. Ap alternately infects ticks and mammals and a variety of cell types within each. Understanding the biology behind such versatile cellular parasitism may be derived through the use of tiling microarrays to establish high resolution, genome-wide transcription profiles of the organism as it infects cell lines representative of its life cycle (tick; ISE6 and pathogenesis (human; HL-60 and HMEC-1. Results Detailed, host cell specific transcriptional behavior was revealed. There was extensive differential Ap gene transcription between the tick (ISE6 and the human (HL-60 and HMEC-1 cell lines, with far fewer differentially transcribed genes between the human cell lines, and all disproportionately represented by membrane or surface proteins. There were Ap genes exclusively transcribed in each cell line, apparent human- and tick-specific operons and paralogs, and anti-sense transcripts that suggest novel expression regulation processes. Seven virB2 paralogs (of the bacterial type IV secretion system showed human or tick cell dependent transcription. Previously unrecognized genes and coding sequences were identified, as were the expressed p44/msp2 (major surface proteins paralogs (of 114 total, through elevated signal produced to the unique hypervariable region of each – 2/114 in HL-60, 3/114 in HMEC-1, and none in ISE6. Conclusion Using these methods, whole genome transcription profiles can likely be generated for Ap, as well as other obligate intracellular organisms, in any host cells and for all stages of the cell infection process. Visual representation of comprehensive transcription data alongside an annotated map of the genome renders complex transcription into discernable patterns.

  6. Transcription as a Threat to Genome Integrity.

    Science.gov (United States)

    Gaillard, Hélène; Aguilera, Andrés

    2016-06-02

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

  7. Diurnal Cycling Transcription Factors of Pineapple Revealed by Genome-Wide Annotation and Global Transcriptomic Analysis.

    Science.gov (United States)

    Sharma, Anupma; Wai, Ching Man; Ming, Ray; Yu, Qingyi

    2017-09-01

    Circadian clock provides fitness advantage by coordinating internal metabolic and physiological processes to external cyclic environments. Core clock components exhibit daily rhythmic changes in gene expression, and the majority of them are transcription factors (TFs) and transcription coregulators (TCs). We annotated 1,398 TFs from 67 TF families and 80 TCs from 20 TC families in pineapple, and analyzed their tissue-specific and diurnal expression patterns. Approximately 42% of TFs and 45% of TCs displayed diel rhythmic expression, including 177 TF/TCs cycling only in the nonphotosynthetic leaf tissue, 247 cycling only in the photosynthetic leaf tissue, and 201 cycling in both. We identified 68 TF/TCs whose cycling expression was tightly coupled between the photosynthetic and nonphotosynthetic leaf tissues. These TF/TCs likely coordinate key biological processes in pineapple as we demonstrated that this group is enriched in homologous genes that form the core circadian clock in Arabidopsis and includes a STOP1 homolog. Two lines of evidence support the important role of the STOP1 homolog in regulating CAM photosynthesis in pineapple. First, STOP1 responds to acidic pH and regulates a malate channel in multiple plant species. Second, the cycling expression pattern of the pineapple STOP1 and the diurnal pattern of malate accumulation in pineapple leaf are correlated. We further examined duplicate-gene retention and loss in major known circadian genes and refined their evolutionary relationships between pineapple and other plants. Significant variations in duplicate-gene retention and loss were observed for most clock genes in both monocots and dicots. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  8. Therapeutics of Ebola hemorrhagic fever: whole-genome transcriptional analysis of successful disease mitigation.

    Science.gov (United States)

    Yen, Judy Y; Garamszegi, Sara; Geisbert, Joan B; Rubins, Kathleen H; Geisbert, Thomas W; Honko, Anna; Xia, Yu; Connor, John H; Hensley, Lisa E

    2011-11-01

    The mechanisms of Ebola (EBOV) pathogenesis are only partially understood, but the dysregulation of normal host immune responses (including destruction of lymphocytes, increases in circulating cytokine levels, and development of coagulation abnormalities) is thought to play a major role. Accumulating evidence suggests that much of the observed pathology is not the direct result of virus-induced structural damage but rather is due to the release of soluble immune mediators from EBOV-infected cells. It is therefore essential to understand how the candidate therapeutic may be interrupting the disease process and/or targeting the infectious agent. To identify genetic signatures that are correlates of protection, we used a DNA microarray-based approach to compare the host genome-wide responses of EBOV-infected nonhuman primates (NHPs) responding to candidate therapeutics. We observed that, although the overall circulating immune response was similar in the presence and absence of coagulation inhibitors, surviving NHPs clustered together. Noticeable differences in coagulation-associated genes appeared to correlate with survival, which revealed a subset of distinctly differentially expressed genes, including chemokine ligand 8 (CCL8/MCP-2), that may provide possible targets for early-stage diagnostics or future therapeutics. These analyses will assist us in understanding the pathogenic mechanisms of EBOV infection and in identifying improved therapeutic strategies.

  9. Genomic analysis of NAC transcription factors in banana (Musa acuminata) and definition of NAC orthologous groups for monocots and dicots.

    Science.gov (United States)

    Cenci, Albero; Guignon, Valentin; Roux, Nicolas; Rouard, Mathieu

    2014-05-01

    Identifying the molecular mechanisms underlying tolerance to abiotic stresses is important in crop breeding. A comprehensive understanding of the gene families associated with drought tolerance is therefore highly relevant. NAC transcription factors form a large plant-specific gene family involved in the regulation of tissue development and responses to biotic and abiotic stresses. The main goal of this study was to set up a framework of orthologous groups determined by an expert sequence comparison of NAC genes from both monocots and dicots. In order to clarify the orthologous relationships among NAC genes of different species, we performed an in-depth comparative study of four divergent taxa, in dicots and monocots, whose genomes have already been completely sequenced: Arabidopsis thaliana, Vitis vinifera, Musa acuminata and Oryza sativa. Due to independent evolution, NAC copy number is highly variable in these plant genomes. Based on an expert NAC sequence comparison, we propose forty orthologous groups of NAC sequences that were probably derived from an ancestor gene present in the most recent common ancestor of dicots and monocots. These orthologous groups provide a curated resource for large-scale protein sequence annotation of NAC transcription factors. The established orthology relationships also provide a useful reference for NAC function studies in newly sequenced genomes such as M. acuminata and other plant species.

  10. Genome-wide identification of WRKY transcription factors in kiwifruit (Actinidia spp.) and analysis of WRKY expression in responses to biotic and abiotic stresses.

    Science.gov (United States)

    Jing, Zhaobin; Liu, Zhande

    2018-04-01

    As one of the largest transcriptional factor families in plants, WRKY transcription factors play important roles in various biotic and abiotic stress responses. To date, WRKY genes in kiwifruit (Actinidia spp.) remain poorly understood. In our study, o total of 97 AcWRKY genes have been identified in the kiwifruit genome. An overview of these AcWRKY genes is analyzed, including the phylogenetic relationships, exon-intron structures, synteny and expression profiles. The 97 AcWRKY genes were divided into three groups based on the conserved WRKY domain. Synteny analysis indicated that segmental duplication events contributed to the expansion of the kiwifruit AcWRKY family. In addition, the synteny analysis between kiwifruit and Arabidopsis suggested that some of the AcWRKY genes were derived from common ancestors before the divergence of these two species. Conserved motifs outside the AcWRKY domain may reflect their functional conservation. Genome-wide segmental and tandem duplication were found, which may contribute to the expansion of AcWRKY genes. Furthermore, the analysis of selected AcWRKY genes showed a variety of expression patterns in five different organs as well as during biotic and abiotic stresses. The genome-wide identification and characterization of kiwifruit WRKY transcription factors provides insight into the evolutionary history and is a useful resource for further functional analyses of kiwifruit.

  11. Genome-wide identification and expression analysis of SBP-like transcription factor genes in Moso Bamboo (Phyllostachys edulis).

    Science.gov (United States)

    Pan, Feng; Wang, Yue; Liu, Huanglong; Wu, Min; Chu, Wenyuan; Chen, Danmei; Xiang, Yan

    2017-06-27

    The SQUAMOSA promoter binding protein-like (SPL) proteins are plant-specific transcription factors (TFs) that function in a variety of developmental processes including growth, flower development, and signal transduction. SPL proteins are encoded by a gene family, and these genes have been characterized in two model grass species, Zea mays and Oryza sativa. The SPL gene family has not been well studied in moso bamboo (Phyllostachys edulis), a woody grass species. We identified 32 putative PeSPL genes in the P. edulis genome. Phylogenetic analysis arranged the PeSPL protein sequences in eight groups. Similarly, phylogenetic analysis of the SBP-like and SBP proteins from rice and maize clustered them into eight groups analogous to those from P. edulis. Furthermore, the deduced PeSPL proteins in each group contained very similar conserved sequence motifs. Our analyses indicate that the PeSPL genes experienced a large-scale duplication event ~15 million years ago (MYA), and that divergence between the PeSPL and OsSPL genes occurred 34 MYA. The stress-response expression profiles and tissue-specificity of the putative PeSPL gene promoter regions showed that SPL genes in moso bamboo have potential biological functions in stress resistance as well as in growth and development. We therefore examined PeSPL gene expression in response to different plant hormone and drought (polyethylene glycol-6000; PEG) treatments to mimic biotic and abiotic stresses. Expression of three (PeSPL10, -12, -17), six (PeSPL1, -10, -12, -17, -20, -31), and nine (PeSPL5, -8, -9, -14, -15, -19, -20, -31, -32) genes remained relatively stable after treating with salicylic acid (SA), gibberellic acid (GA), and PEG, respectively, while the expression patterns of other genes changed. In addition, analysis of tissue-specific expression of the moso bamboo SPL genes during development showed differences in their spatiotemporal expression patterns, and many were expressed at high levels in flowers and

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

    DEFF Research Database (Denmark)

    Valen, Eivind; Sandelin, Albin Gustav

    2011-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Kang Keunsoo

    2013-01-01

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

  14. Reconstructing transcriptional regulatory networks through genomics data

    OpenAIRE

    Sun, Ning; Zhao, Hongyu

    2009-01-01

    One central problem in biology is to understand how gene expression is regulated under different conditions. Microarray gene expression data and other high throughput data have made it possible to dissect transcriptional regulatory networks at the genomics level. Owing to the very large number of genes that need to be studied, the relatively small number of data sets available, the noise in the data and the different natures of the distinct data types, network inference presents great challen...

  15. Discovering transcription factor binding sites in highly repetitive regions of genomes with multi-read analysis of ChIP-Seq data.

    Science.gov (United States)

    Chung, Dongjun; Kuan, Pei Fen; Li, Bo; Sanalkumar, Rajendran; Liang, Kun; Bresnick, Emery H; Dewey, Colin; Keleş, Sündüz

    2011-07-01

    Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) is rapidly replacing chromatin immunoprecipitation combined with genome-wide tiling array analysis (ChIP-chip) as the preferred approach for mapping transcription-factor binding sites and chromatin modifications. The state of the art for analyzing ChIP-seq data relies on using only reads that map uniquely to a relevant reference genome (uni-reads). This can lead to the omission of up to 30% of alignable reads. We describe a general approach for utilizing reads that map to multiple locations on the reference genome (multi-reads). Our approach is based on allocating multi-reads as fractional counts using a weighted alignment scheme. Using human STAT1 and mouse GATA1 ChIP-seq datasets, we illustrate that incorporation of multi-reads significantly increases sequencing depths, leads to detection of novel peaks that are not otherwise identifiable with uni-reads, and improves detection of peaks in mappable regions. We investigate various genome-wide characteristics of peaks detected only by utilization of multi-reads via computational experiments. Overall, peaks from multi-read analysis have similar characteristics to peaks that are identified by uni-reads except that the majority of them reside in segmental duplications. We further validate a number of GATA1 multi-read only peaks by independent quantitative real-time ChIP analysis and identify novel target genes of GATA1. These computational and experimental results establish that multi-reads can be of critical importance for studying transcription factor binding in highly repetitive regions of genomes with ChIP-seq experiments.

  16. Discovering transcription factor binding sites in highly repetitive regions of genomes with multi-read analysis of ChIP-Seq data.

    Directory of Open Access Journals (Sweden)

    Dongjun Chung

    2011-07-01

    Full Text Available Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq is rapidly replacing chromatin immunoprecipitation combined with genome-wide tiling array analysis (ChIP-chip as the preferred approach for mapping transcription-factor binding sites and chromatin modifications. The state of the art for analyzing ChIP-seq data relies on using only reads that map uniquely to a relevant reference genome (uni-reads. This can lead to the omission of up to 30% of alignable reads. We describe a general approach for utilizing reads that map to multiple locations on the reference genome (multi-reads. Our approach is based on allocating multi-reads as fractional counts using a weighted alignment scheme. Using human STAT1 and mouse GATA1 ChIP-seq datasets, we illustrate that incorporation of multi-reads significantly increases sequencing depths, leads to detection of novel peaks that are not otherwise identifiable with uni-reads, and improves detection of peaks in mappable regions. We investigate various genome-wide characteristics of peaks detected only by utilization of multi-reads via computational experiments. Overall, peaks from multi-read analysis have similar characteristics to peaks that are identified by uni-reads except that the majority of them reside in segmental duplications. We further validate a number of GATA1 multi-read only peaks by independent quantitative real-time ChIP analysis and identify novel target genes of GATA1. These computational and experimental results establish that multi-reads can be of critical importance for studying transcription factor binding in highly repetitive regions of genomes with ChIP-seq experiments.

  17. Genome-wide Analysis of RARβ Transcriptional Targets in Mouse Striatum Links Retinoic Acid Signaling with Huntington's Disease and Other Neurodegenerative Disorders.

    Science.gov (United States)

    Niewiadomska-Cimicka, Anna; Krzyżosiak, Agnieszka; Ye, Tao; Podleśny-Drabiniok, Anna; Dembélé, Doulaye; Dollé, Pascal; Krężel, Wojciech

    2017-07-01

    Retinoic acid (RA) signaling through retinoic acid receptors (RARs), known for its multiple developmental functions, emerged more recently as an important regulator of adult brain physiology. How RAR-mediated regulation is achieved is poorly known, partly due to the paucity of information on critical target genes in the brain. Also, it is not clear how reduced RA signaling may contribute to pathophysiology of diverse neuropsychiatric disorders. We report the first genome-wide analysis of RAR transcriptional targets in the brain. Using chromatin immunoprecipitation followed by high-throughput sequencing and transcriptomic analysis of RARβ-null mutant mice, we identified genomic targets of RARβ in the striatum. Characterization of RARβ transcriptional targets in the mouse striatum points to mechanisms through which RAR may control brain functions and display neuroprotective activity. Namely, our data indicate with statistical significance (FDR 0.1) a strong contribution of RARβ in controlling neurotransmission, energy metabolism, and transcription, with a particular involvement of G-protein coupled receptor (p = 5.0e -5 ), cAMP (p = 4.5e -4 ), and calcium signaling (p = 3.4e -3 ). Many identified RARβ target genes related to these pathways have been implicated in Alzheimer's, Parkinson's, and Huntington's disease (HD), raising the possibility that compromised RA signaling in the striatum may be a mechanistic link explaining the similar affective and cognitive symptoms in these diseases. The RARβ transcriptional targets were particularly enriched for transcripts affected in HD. Using the R6/2 transgenic mouse model of HD, we show that partial sequestration of RARβ in huntingtin protein aggregates may account for reduced RA signaling reported in HD.

  18. Genome-wide analysis identifies chickpea (Cicer arietinum) heat stress transcription factors (Hsfs) responsive to heat stress at the pod development stage.

    Science.gov (United States)

    Chidambaranathan, Parameswaran; Jagannadham, Prasanth Tej Kumar; Satheesh, Viswanathan; Kohli, Deshika; Basavarajappa, Santosh Halasabala; Chellapilla, Bharadwaj; Kumar, Jitendra; Jain, Pradeep Kumar; Srinivasan, R

    2018-05-01

    The heat stress transcription factors (Hsfs) play a prominent role in thermotolerance and eliciting the heat stress response in plants. Identification and expression analysis of Hsfs gene family members in chickpea would provide valuable information on heat stress responsive Hsfs. A genome-wide analysis of Hsfs gene family resulted in the identification of 22 Hsf genes in chickpea in both desi and kabuli genome. Phylogenetic analysis distinctly separated 12 A, 9 B, and 1 C class Hsfs, respectively. An analysis of cis-regulatory elements in the upstream region of the genes identified many stress responsive elements such as heat stress elements (HSE), abscisic acid responsive element (ABRE) etc. In silico expression analysis showed nine and three Hsfs were also expressed in drought and salinity stresses, respectively. Q-PCR expression analysis of Hsfs under heat stress at pod development and at 15 days old seedling stage showed that CarHsfA2, A6, and B2 were significantly upregulated in both the stages of crop growth and other four Hsfs (CarHsfA2, A6a, A6c, B2a) showed early transcriptional upregulation for heat stress at seedling stage of chickpea. These subclasses of Hsfs identified in this study can be further evaluated as candidate genes in the characterization of heat stress response in chickpea.

  19. Genome-wide transcription analyses in rice using tiling microarrays

    DEFF Research Database (Denmark)

    Li, Lei; Wang, Xiangfeng; Stolc, Viktor

    2006-01-01

    . We report here a full-genome transcription analysis of the indica rice subspecies using high-density oligonucleotide tiling microarrays. Our results provided expression data support for the existence of 35,970 (81.9%) annotated gene models and identified 5,464 unique transcribed intergenic regions...... that share similar compositional properties with the annotated exons and have significant homology to other plant proteins. Elucidating and mapping of all transcribed regions revealed an association between global transcription and cytological chromosome features, and an overall similarity of transcriptional......Sequencing and computational annotation revealed several features, including high gene numbers, unusual composition of the predicted genes and a large number of genes lacking homology to known genes, that distinguish the rice (Oryza sativa) genome from that of other fully sequenced model species...

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

    Directory of Open Access Journals (Sweden)

    Janet L Smith

    2015-05-01

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

  1. Genome-wide Expression Analysis and Metabolite Profiling Elucidate Transcriptional Regulation of Flavonoid Biosynthesis and Modulation under Abiotic Stresses in Banana.

    Science.gov (United States)

    Pandey, Ashutosh; Alok, Anshu; Lakhwani, Deepika; Singh, Jagdeep; Asif, Mehar H; Trivedi, Prabodh K

    2016-08-19

    Flavonoid biosynthesis is largely regulated at the transcriptional level due to the modulated expression of genes related to the phenylpropanoid pathway in plants. Although accumulation of different flavonoids has been reported in banana, a staple fruit crop, no detailed information is available on regulation of the biosynthesis in this important plant. We carried out genome-wide analysis of banana (Musa acuminata, AAA genome) and identified 28 genes belonging to 9 gene families associated with flavonoid biosynthesis. Expression analysis suggested spatial and temporal regulation of the identified genes in different tissues of banana. Analysis revealed enhanced expression of genes related to flavonol and proanthocyanidin (PA) biosynthesis in peel and pulp at the early developmental stages of fruit. Genes involved in anthocyanin biosynthesis were highly expressed during banana fruit ripening. In general, higher accumulation of metabolites was observed in the peel as compared to pulp tissue. A correlation between expression of genes and metabolite content was observed at the early stage of fruit development. Furthermore, this study also suggests regulation of flavonoid biosynthesis, at transcriptional level, under light and dark exposures as well as methyl jasmonate (MJ) treatment in banana.

  2. Genome-wide identification and comparative analysis of squamosa-promoter binding proteins (sbp) transcription factor family in gossypium raimondii and arabidopsis thaliana

    International Nuclear Information System (INIS)

    Ali, M.A.; Alia, K.B.; Atif, R.M.; Rasulj, I.; Nadeem, H.U.; Shahid, A.; Azeem, F

    2017-01-01

    SQUAMOSA-Promoter Binding Proteins (SBP) are class of transcription factors that play vital role in regulation of plant tissue growth and development. The genes encoding these proteins have not yet been identified in diploid cotton. Thus here, a comprehensive genome wide analysis of SBP genes/proteins was carried out to identify the genes encoding SBP proteins in Gossypium raimondii and Arabidopsis thaliana. We identified 17 SBP genes from Arabidopsis thaliana genome and 30 SBP genes from Gossypium raimondii. Chromosome localization studies revealed the uneven distribution of SBP encoding genes both in the genomes of A. thaliana and G. raimondii. In cotton, five SBP genes were located on chromosome no. 2, while no gene was found on chromosome 9. In A. thaliana, maximum seven SBP genes were identified on chromosome 9, while chromosome 4 did not have any SBP gene. Thus, the SBP gene family might have expanded as a result of segmental as well as tandem duplications in these species. The comparative phylogenetic analysis of Arabidopsis and cotton SBPs revealed the presence of eight groups. The gene structure analysis of SBP encoding genes revealed the presence of one to eleven inrons in both Arabidopsis and G. raimondii. The proteins sharing the same phyletic group mostly demonstrated the similar intron-exon occurrence pattern; and share the common conserved domains. The SBP DNA-binding domain shared 24 absolutely conserved residues in Arabidopsis. The present study can serve as a base for the functional characterization of SBP gene family in Gossypium raimondii. (author)

  3. Enriching Genomic Resources and Transcriptional Profile Analysis of Miscanthus sinensis under Drought Stress Based on RNA Sequencing

    Directory of Open Access Journals (Sweden)

    Gang Nie

    2017-01-01

    Full Text Available Miscanthus × giganteus is wildly cultivated as a potential biofuel feedstock around the world; however, the narrow genetic basis and sterile characteristics have become a limitation for its utilization. As a progenitor of M. × giganteus, M. sinensis is widely distributed around East Asia providing well abiotic stress tolerance. To enrich the M. sinensis genomic databases and resources, we sequenced and annotated the transcriptome of M. sinensis by using an Illumina HiSeq 2000 platform. Approximately 316 million high-quality trimmed reads were generated from 349 million raw reads, and a total of 114,747 unigenes were obtained after de novo assembly. Furthermore, 95,897 (83.57% unigenes were annotated to at least one database including NR, Swiss-Prot, KEGG, COG, GO, and NT, supporting that the sequences obtained were annotated properly. Differentially expressed gene analysis indicates that drought stress 15 days could be a critical period for M. sinensis response to drought stress. The high-throughput transcriptome sequencing of M. sinensis under drought stress has greatly enriched the current genomic available resources. The comparison of DEGs under different periods of drought stress identified a wealth of candidate genes involved in drought tolerance regulatory networks, which will facilitate further genetic improvement and molecular studies of the M. sinensis.

  4. Genome-wide identification and characterization of cacao WRKY transcription factors and analysis of their expression in response to witches' broom disease.

    Science.gov (United States)

    Silva Monteiro de Almeida, Dayanne; Oliveira Jordão do Amaral, Daniel; Del-Bem, Luiz-Eduardo; Bronze Dos Santos, Emily; Santana Silva, Raner José; Peres Gramacho, Karina; Vincentz, Michel; Micheli, Fabienne

    2017-01-01

    Transcriptional regulation, led by transcription factors (TFs) such as those of the WRKY family, is a mechanism used by the organism to enhance or repress gene expression in response to stimuli. Here, we report on the genome-wide analysis of the Theobroma cacao WRKY TF family and also investigate the expression of WRKY genes in cacao infected by the fungus Moniliophthora perniciosa. In the cacao genome, 61 non-redundant WRKY sequences were found and classified in three groups (I to III) according to the WRKY and zinc-finger motif types. The 61 putative WRKY sequences were distributed on the 10 cacao chromosomes and 24 of them came from duplication events. The sequences were phylogenetically organized according to the general WRKY groups. The phylogenetic analysis revealed that subgroups IIa and IIb are sister groups and share a common ancestor, as well as subgroups IId and IIe. The most divergent groups according to the plant origin were IIc and III. According to the phylogenetic analysis, 7 TcWRKY genes were selected and analyzed by RT-qPCR in susceptible and resistant cacao plants infected (or not) with M. perniciosa. Some TcWRKY genes presented interesting responses to M. perniciosa such as Tc01_p014750/Tc06_p013130/AtWRKY28, Tc09_p001530/Tc06_p004420/AtWRKY40, Tc04_p016130/AtWRKY54 and Tc10_p016570/ AtWRKY70. Our results can help to select appropriate candidate genes for further characterization in cacao or in other Theobroma species.

  5. Genome-wide identification and characterization of cacao WRKY transcription factors and analysis of their expression in response to witches' broom disease

    Science.gov (United States)

    Silva Monteiro de Almeida, Dayanne; Oliveira Jordão do Amaral, Daniel; Del-Bem, Luiz-Eduardo; Bronze dos Santos, Emily; Santana Silva, Raner José; Peres Gramacho, Karina; Vincentz, Michel

    2017-01-01

    Transcriptional regulation, led by transcription factors (TFs) such as those of the WRKY family, is a mechanism used by the organism to enhance or repress gene expression in response to stimuli. Here, we report on the genome-wide analysis of the Theobroma cacao WRKY TF family and also investigate the expression of WRKY genes in cacao infected by the fungus Moniliophthora perniciosa. In the cacao genome, 61 non-redundant WRKY sequences were found and classified in three groups (I to III) according to the WRKY and zinc-finger motif types. The 61 putative WRKY sequences were distributed on the 10 cacao chromosomes and 24 of them came from duplication events. The sequences were phylogenetically organized according to the general WRKY groups. The phylogenetic analysis revealed that subgroups IIa and IIb are sister groups and share a common ancestor, as well as subgroups IId and IIe. The most divergent groups according to the plant origin were IIc and III. According to the phylogenetic analysis, 7 TcWRKY genes were selected and analyzed by RT-qPCR in susceptible and resistant cacao plants infected (or not) with M. perniciosa. Some TcWRKY genes presented interesting responses to M. perniciosa such as Tc01_p014750/Tc06_p013130/AtWRKY28, Tc09_p001530/Tc06_p004420/AtWRKY40, Tc04_p016130/AtWRKY54 and Tc10_p016570/ AtWRKY70. Our results can help to select appropriate candidate genes for further characterization in cacao or in other Theobroma species. PMID:29084273

  6. Genome-wide identification and comparative analysis of the heat shock transcription factor family in Chinese white pear (Pyrus bretschneideri) and five other Rosaceae species.

    Science.gov (United States)

    Qiao, Xin; Li, Meng; Li, Leiting; Yin, Hao; Wu, Juyou; Zhang, Shaoling

    2015-01-21

    Heat shock transcription factors (Hsfs), which act as important transcriptional regulatory proteins in eukaryotes, play a central role in controlling the expression of heat-responsive genes. At present, the genomes of Chinese white pear ('Dangshansuli') and five other Rosaceae fruit crops have been fully sequenced. However, information about the Hsfs gene family in these Rosaceae species is limited, and the evolutionary history of the Hsfs gene family also remains unresolved. In this study, 137 Hsf genes were identified from six Rosaceae species (Pyrus bretschneideri, Malus × domestica, Prunus persica, Fragaria vesca, Prunus mume, and Pyrus communis), 29 of which came from Chinese white pear, designated as PbHsf. Based on the structural characteristics and phylogenetic analysis of these sequences, the Hsf family genes could be classified into three main groups (classes A, B, and C). Segmental and dispersed duplications were the primary forces underlying Hsf gene family expansion in the Rosaceae. Most of the PbHsf duplicated gene pairs were dated back to the recent whole-genome duplication (WGD, 30-45 million years ago (MYA)). Purifying selection also played a critical role in the evolution of Hsf genes. Transcriptome data demonstrated that the expression levels of the PbHsf genes were widely different. Six PbHsf genes were upregulated in fruit under naturally increased temperature. A comprehensive analysis of Hsf genes was performed in six Rosaceae species, and 137 full length Hsf genes were identified. The results presented here will undoubtedly be useful for better understanding the complexity of the Hsf gene family and will facilitate functional characterization in future studies.

  7. Transcript profiling of common bean (Phaseolus vulgaris L. using the GeneChip® Soybean Genome Array: optimizing analysis by masking biased probes

    Directory of Open Access Journals (Sweden)

    Gronwald John W

    2010-05-01

    Full Text Available Abstract Background Common bean (Phaseolus vulgaris L. and soybean (Glycine max both belong to the Phaseoleae tribe and share significant coding sequence homology. This suggests that the GeneChip® Soybean Genome Array (soybean GeneChip may be used for gene expression studies using common bean. Results To evaluate the utility of the soybean GeneChip for transcript profiling of common bean, we hybridized cRNAs purified from nodule, leaf, and root of common bean and soybean in triplicate to the soybean GeneChip. Initial data analysis showed a decreased sensitivity and accuracy of measuring differential gene expression in common bean cross-species hybridization (CSH GeneChip data compared to that of soybean. We employed a method that masked putative probes targeting inter-species variable (ISV regions between common bean and soybean. A masking signal intensity threshold was selected that optimized both sensitivity and accuracy of measuring differential gene expression. After masking for ISV regions, the number of differentially-expressed genes identified in common bean was increased by 2.8-fold reflecting increased sensitivity. Quantitative RT-PCR (qRT-PCR analysis of 20 randomly selected genes and purine-ureide pathway genes demonstrated an increased accuracy of measuring differential gene expression after masking for ISV regions. We also evaluated masked probe frequency per probe set to gain insight into the sequence divergence pattern between common bean and soybean. The sequence divergence pattern analysis suggested that the genes for basic cellular functions and metabolism were highly conserved between soybean and common bean. Additionally, our results show that some classes of genes, particularly those associated with environmental adaptation, are highly divergent. Conclusions The soybean GeneChip is a suitable cross-species platform for transcript profiling in common bean when used in combination with the masking protocol described. In

  8. Mycoplasma hyopneumoniae Transcription Unit Organization: Genome Survey and Prediction

    Science.gov (United States)

    Siqueira, Franciele Maboni; Schrank, Augusto; Schrank, Irene Silveira

    2011-01-01

    Mycoplasma hyopneumoniae is associated with swine respiratory diseases. Although gene organization and regulation are well known in many prokaryotic organisms, knowledge on mycoplasma is limited. This study performed a comparative analysis of three strains of M. hyopneumoniae (7448, J and 232), with a focus on genome organization and gene comparison for open read frame (ORF) cluster (OC) identification. An in silico analysis of gene organization demonstrated 117 OCs and 34 single ORFs in M. hyopneumoniae 7448 and J, while 116 OCs and 36 single ORFs were identified in M. hyopneumoniae 232. Genomic comparison revealed high synteny and conservation of gene order between the OCs defined for 7448 and J strains as well as for 7448 and 232 strains. Twenty-one OCs were chosen and experimentally confirmed by reverse transcription–PCR from M. hyopneumoniae 7448 genome, validating our prediction. A subset of the ORFs within an OC could be independently transcribed due to the presence of internal promoters. Our results suggest that transcription occurs in ‘run-on’ from an upstream promoter in M. hyopneumoniae, thus forming large ORF clusters (from 2 to 29 ORFs in the same orientation) and indicating a complex transcriptional organization. PMID:22086999

  9. Identification of genes associated with nitrogen-use efficiency by genome-wide transcriptional analysis of two soybean genotypes

    Directory of Open Access Journals (Sweden)

    Zhou Xin A

    2011-10-01

    Full Text Available Abstract Background Soybean is a valuable crop that provides protein and oil. Soybean requires a large amount of nitrogen (N to accumulate high levels of N in the seed. The yield and protein content of soybean seeds are directly affected by the N-use efficiency (NUE of the plant, and improvements in NUE will improve yields and quality of soybean products. Genetic engineering is one of the approaches to improve NUE, but at present, it is hampered by the lack of information on genes associated with NUE. Solexa sequencing is a new method for estimating gene expression in the transcription level. Here, the expression profiles were analyzed between two soybean varieties in N-limited conditions to identify genes related to NUE. Results Two soybean genotypes were grown under N-limited conditions; a low-N-tolerant variety (No.116 and a low-N-sensitive variety (No.84-70. The shoots and roots of soybeans were used for sequencing. Eight libraries were generated for analysis: 2 genotypes × 2 tissues (roots and shoots × 2 time periods [short-term (0.5 to 12 h and long-term (3 to 12 d responses] and compared the transcriptomes by high-throughput tag-sequencing analysis. 5,739,999, 5,846,807, 5,731,901, 5,970,775, 5,476,878, 5,900,343, 5,930,716, and 5,862,642 clean tags were obtained for the eight libraries: L1, 116-shoot short-term; L2 84-70-shoot short-term; L3 116-shoot long-term; L4 84-70-shoot long-term; L5 116-root short-term; L6 84-70-root short-term; L7 116-root long-term;L8 84-70-root long-term; these corresponded to 224,154, 162,415, 191,994, 181,792, 204,639, 206,998, 233,839 and 257,077 distinct tags, respectively. The clean tags were mapped to the reference sequences for annotation of expressed genes. Many genes showed substantial differences in expression among the libraries. In total, 3,231genes involved in twenty-two metabolic and signal transduction pathways were up- or down-regulated. Twenty-four genes were randomly selected and confirmed

  10. Genome-Wide Analysis of the AP2/ERF Transcription Factors Family and the Expression Patterns of DREB Genes in Moso Bamboo (Phyllostachys edulis.

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    Huili Wu

    Full Text Available The AP2/ERF transcription factor family, one of the largest families unique to plants, performs a significant role in terms of regulation of growth and development, and responses to biotic and abiotic stresses. Moso bamboo (Phyllostachys edulis is a fast-growing non-timber forest species with the highest ecological, economic and social values of all bamboos in Asia. The draft genome of moso bamboo and the available genomes of other plants provide great opportunities to research global information on the AP2/ERF family in moso bamboo. In total, 116 AP2/ERF transcription factors were identified in moso bamboo. The phylogeny analyses indicated that the 116 AP2/ERF genes could be divided into three subfamilies: AP2, RAV and ERF; and the ERF subfamily genes were divided into 11 groups. The gene structures, exons/introns and conserved motifs of the PeAP2/ERF genes were analyzed. Analysis of the evolutionary patterns and divergence showed the PeAP2/ERF genes underwent a large-scale event around 15 million years ago (MYA and the division time of AP2/ERF family genes between rice and moso bamboo was 15-23 MYA. We surveyed the putative promoter regions of the PeDREBs and showed that largely stress-related cis-elements existed in these genes. Further analysis of expression patterns of PeDREBs revealed that the most were strongly induced by drought, low-temperature and/or high salinity stresses in roots and, in contrast, most PeDREB genes had negative functions in leaves under the same respective stresses. In this study there were two main interesting points: there were fewer members of the PeDREB subfamily in moso bamboo than in other plants and there were differences in DREB gene expression profiles between leaves and roots triggered in response to abiotic stress. The information produced from this study may be valuable in overcoming challenges in cultivating moso bamboo.

  11. Genome-wide identification and characterization of WRKY transcriptional factor family in apple and analysis of their responses to waterlogging and drought stress.

    Science.gov (United States)

    Meng, Dong; Li, Yuanyuan; Bai, Yang; Li, Mingjun; Cheng, Lailiang

    2016-06-01

    As one of the largest transcriptional factor families in plants, WRKY genes play significant roles in various biotic and abiotic stress responses. Although the WRKY gene family has been characterized in a few plant species, the details remain largely unknown in the apple (Malus domestica Borkh.). In this study, we identified a total of 127 MdWRKYs from the apple genome, which were divided into four subgroups according to the WRKY domains and zinc finger motif. Most of them were mapped onto the apple's 17 chromosomes and were expressed in more than one tissue, including shoot tips, mature leaves, fruit and apple calli. We then contrasted WRKY expression patterns between calli grown in solid medium (control) and liquid medium (representing waterlogging stress) and found that 34 WRKY genes were differentially expressed between the two growing conditions. Finally, we determined the expression patterns of 10 selected WRKY genes in an apple rootstock, G41, in response to waterlogging and drought stress, which identified candidate genes involved in responses to water stress for functional analysis. Our data provide interesting candidate MdWRKYs for future functional analysis and demonstrate that apple callus is a useful system for characterizing gene expression and function in apple. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  12. Genome Binding and Gene Regulation by Stem Cell Transcription Factors

    NARCIS (Netherlands)

    J.H. Brandsma (Johan)

    2016-01-01

    markdownabstractNearly all cells of an individual organism contain the same genome. However, each cell type transcribes a different set of genes due to the presence of different sets of cell type-specific transcription factors. Such transcription factors bind to regulatory regions such as promoters

  13. Genome-wide analysis and identification of stress-responsive genes of the NAM-ATAF1,2-CUC2 transcription factor family in apple.

    Science.gov (United States)

    Su, Hongyan; Zhang, Shizhong; Yuan, Xiaowei; Chen, Changtian; Wang, Xiao-Fei; Hao, Yu-Jin

    2013-10-01

    NAC (NAM, ATAF1,2, and CUC2) proteins constitute one of the largest families of plant-specific transcription factors. To date, little is known about the NAC genes in the apple (Malus domestica). In this study, a total of 180 NAC genes were identified in the apple genome and were phylogenetically clustered into six groups (I-VI) with the NAC genes from Arabidopsis and rice. The predicted apple NAC genes were distributed across all of 17 chromosomes at various densities. Additionally, the gene structure and motif compositions of the apple NAC genes were analyzed. Moreover, the expression of 29 selected apple NAC genes was analyzed in different tissues and under different abiotic stress conditions. All of the selected genes, with the exception of four genes, were expressed in at least one of the tissues tested, which indicates that the NAC genes are involved in various aspects of the physiological and developmental processes of the apple. Encouragingly, 17 of the selected genes were found to respond to one or more of the abiotic stress treatments, and these 17 genes included not only the expected 7 genes that were clustered with the well-known stress-related marker genes in group IV but also 10 genes located in other subgroups, none of which contains members that have been reported to be stress-related. To the best of our knowledge, this report describes the first genome-wide analysis of the apple NAC gene family, and the results should provide valuable information for understanding the classification and putative functions of this family. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  14. The transcriptionally active regions in the genome of Bacillus subtilis

    DEFF Research Database (Denmark)

    Rasmussen, Simon; Nielsen, Henrik Bjørn; Jarmer, Hanne Østergaard

    2009-01-01

    The majority of all genes have so far been identified and annotated systematically through in silico gene finding. Here we report the finding of 3662 strand-specific transcriptionally active regions (TARs) in the genome of Bacillus subtilis by the use of tiling arrays. We have measured the genome...

  15. Whole-genome transcription and DNA methylation analysis of peripheral blood mononuclear cells identified aberrant gene regulation pathways in systemic lupus erythematosus.

    Science.gov (United States)

    Zhu, Honglin; Mi, Wentao; Luo, Hui; Chen, Tao; Liu, Shengxi; Raman, Indu; Zuo, Xiaoxia; Li, Quan-Zhen

    2016-07-13

    Recent achievement in genetics and epigenetics has led to the exploration of the pathogenesis of systemic lupus erythematosus (SLE). Identification of differentially expressed genes and their regulatory mechanism(s) at whole-genome level will provide a comprehensive understanding of the development of SLE and its devastating complications, lupus nephritis (LN). We performed whole-genome transcription and DNA methylation analysis in PBMC of 30 SLE patients, including 15 with LN (SLE LN(+)) and 15 without LN (SLE LN(-)), and 25 normal controls (NC) using HumanHT-12 Beadchips and Illumina Human Methy450 chips. The serum proinflammatory cytokines were quantified using Bio-plex Human Cytokine 27-plex assay. Differentially expressed genes and differentially methylated CpG were analyzed with GenomeStudio, R, and SAM software. The association between DNA methylation and gene expression were tested. Gene interaction pathways of the differentially expressed genes were analyzed by IPA software. We identified 552 upregulated genes and 550 downregulated genes in PBMC of SLE. Integration of DNA methylation and gene expression profiling showed that 334 upregulated genes were hypomethylated, and 479 downregulated genes were hypermethylated. Pathway analysis on the differential genes in SLE revealed significant enrichment in interferon (IFN) signaling and toll-like receptor (TLR) signaling pathways. Nine IFN- and seven TLR-related genes were identified and displayed step-wise increase in SLE LN(-) and SLE LN(+). Hypomethylated CpG sites were detected on these genes. The gene expressions for MX1, GPR84, and E2F2 were increased in SLE LN(+) as compared to SLE LN(-) patients. The serum levels of inflammatory cytokines, including IL17A, IP-10, bFGF, TNF-α, IL-6, IL-15, GM-CSF, IL-1RA, IL-5, and IL-12p70, were significantly elevated in SLE compared with NC. The levels of IL-15 and IL1RA correlated with their mRNA expression. The upregulation of IL-15 may be regulated by hypomethylated

  16. Zipper plot: visualizing transcriptional activity of genomic regions.

    Science.gov (United States)

    Avila Cobos, Francisco; Anckaert, Jasper; Volders, Pieter-Jan; Everaert, Celine; Rombaut, Dries; Vandesompele, Jo; De Preter, Katleen; Mestdagh, Pieter

    2017-05-02

    Reconstructing transcript models from RNA-sequencing (RNA-seq) data and establishing these as independent transcriptional units can be a challenging task. Current state-of-the-art tools for long non-coding RNA (lncRNA) annotation are mainly based on evolutionary constraints, which may result in false negatives due to the overall limited conservation of lncRNAs. To tackle this problem we have developed the Zipper plot, a novel visualization and analysis method that enables users to simultaneously interrogate thousands of human putative transcription start sites (TSSs) in relation to various features that are indicative for transcriptional activity. These include publicly available CAGE-sequencing, ChIP-sequencing and DNase-sequencing datasets. Our method only requires three tab-separated fields (chromosome, genomic coordinate of the TSS and strand) as input and generates a report that includes a detailed summary table, a Zipper plot and several statistics derived from this plot. Using the Zipper plot, we found evidence of transcription for a set of well-characterized lncRNAs and observed that fewer mono-exonic lncRNAs have CAGE peaks overlapping with their TSSs compared to multi-exonic lncRNAs. Using publicly available RNA-seq data, we found more than one hundred cases where junction reads connected protein-coding gene exons with a downstream mono-exonic lncRNA, revealing the need for a careful evaluation of lncRNA 5'-boundaries. Our method is implemented using the statistical programming language R and is freely available as a webtool.

  17. Genome-wide survey and expression analysis of the plant-specific NAC transcription factor family in soybean during development and dehydration stress.

    Science.gov (United States)

    Le, Dung Tien; Nishiyama, Rie; Watanabe, Yasuko; Mochida, Keiichi; Yamaguchi-Shinozaki, Kazuko; Shinozaki, Kazuo; Tran, Lam-Son Phan

    2011-08-01

    Plant-specific NAC transcription factors (TFs) play important roles in regulating diverse biological processes, including development, senescence, growth, cell division and responses to environmental stress stimuli. Within the soybean genome, we identified 152 full-length GmNAC TFs, including 11 membrane-bound members. In silico analysis of the GmNACs, together with their Arabidopsis and rice counterparts, revealed similar NAC architecture. Next, we explored the soybean Affymetrix array and Illumina transcriptome sequence data to analyse tissue-specific expression profiles of GmNAC genes. Phylogenetic analysis using stress-related NAC TFs from Arabidopsis and rice as seeding sequences identified 58 of the 152 GmNACs as putative stress-responsive genes, including eight previously reported dehydration-responsive GmNACs. We could design gene-specific primers for quantitative real-time PCR verification of 38 out of 50 newly predicted stress-related genes. Twenty-five and six GmNACs were found to be induced and repressed 2-fold or more, respectively, in soybean roots and/or shoots in response to dehydration. GmNAC085, whose amino acid sequence was 39%; identical to that of well-known SNAC1/ONAC2, was the most induced gene upon dehydration, showing 390-fold and 20-fold induction in shoots and roots, respectively. Our systematic analysis has identified excellent tissue-specific and/or dehydration-responsive candidate GmNAC genes for in-depth characterization and future development of improved drought-tolerant transgenic soybeans.

  18. Pervasive, Genome-Wide Transcription in the Organelle Genomes of Diverse Plastid-Bearing Protists

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    Matheus Sanitá Lima

    2017-11-01

    Full Text Available Organelle genomes are among the most sequenced kinds of chromosome. This is largely because they are small and widely used in molecular studies, but also because next-generation sequencing technologies made sequencing easier, faster, and cheaper. However, studies of organelle RNA have not kept pace with those of DNA, despite huge amounts of freely available eukaryotic RNA-sequencing (RNA-seq data. Little is known about organelle transcription in nonmodel species, and most of the available eukaryotic RNA-seq data have not been mined for organelle transcripts. Here, we use publicly available RNA-seq experiments to investigate organelle transcription in 30 diverse plastid-bearing protists with varying organelle genomic architectures. Mapping RNA-seq data to organelle genomes revealed pervasive, genome-wide transcription, regardless of the taxonomic grouping, gene organization, or noncoding content. For every species analyzed, transcripts covered ≥85% of the mitochondrial and/or plastid genomes (all of which were ≤105 kb, indicating that most of the organelle DNA—coding and noncoding—is transcriptionally active. These results follow earlier studies of model species showing that organellar transcription is coupled and ubiquitous across the genome, requiring significant downstream processing of polycistronic transcripts. Our findings suggest that noncoding organelle DNA can be transcriptionally active, raising questions about the underlying function of these transcripts and underscoring the utility of publicly available RNA-seq data for recovering complete genome sequences. If pervasive transcription is also found in bigger organelle genomes (>105 kb and across a broader range of eukaryotes, this could indicate that noncoding organelle RNAs are regulating fundamental processes within eukaryotic cells.

  19. Pervasive, Genome-Wide Transcription in the Organelle Genomes of Diverse Plastid-Bearing Protists.

    Science.gov (United States)

    Sanitá Lima, Matheus; Smith, David Roy

    2017-11-06

    Organelle genomes are among the most sequenced kinds of chromosome. This is largely because they are small and widely used in molecular studies, but also because next-generation sequencing technologies made sequencing easier, faster, and cheaper. However, studies of organelle RNA have not kept pace with those of DNA, despite huge amounts of freely available eukaryotic RNA-sequencing (RNA-seq) data. Little is known about organelle transcription in nonmodel species, and most of the available eukaryotic RNA-seq data have not been mined for organelle transcripts. Here, we use publicly available RNA-seq experiments to investigate organelle transcription in 30 diverse plastid-bearing protists with varying organelle genomic architectures. Mapping RNA-seq data to organelle genomes revealed pervasive, genome-wide transcription, regardless of the taxonomic grouping, gene organization, or noncoding content. For every species analyzed, transcripts covered ≥85% of the mitochondrial and/or plastid genomes (all of which were ≤105 kb), indicating that most of the organelle DNA-coding and noncoding-is transcriptionally active. These results follow earlier studies of model species showing that organellar transcription is coupled and ubiquitous across the genome, requiring significant downstream processing of polycistronic transcripts. Our findings suggest that noncoding organelle DNA can be transcriptionally active, raising questions about the underlying function of these transcripts and underscoring the utility of publicly available RNA-seq data for recovering complete genome sequences. If pervasive transcription is also found in bigger organelle genomes (>105 kb) and across a broader range of eukaryotes, this could indicate that noncoding organelle RNAs are regulating fundamental processes within eukaryotic cells. Copyright © 2017 Sanitá Lima and Smith.

  20. Genome-wide analysis of brain and gonad transcripts reveals changes of key sex reversal-related genes expression and signaling pathways in three stages of Monopterus albus.

    Directory of Open Access Journals (Sweden)

    Wei Chi

    Full Text Available The natural sex reversal severely affects the sex ratio and thus decreases the productivity of the rice field eel (Monopterus albus. How to understand and manipulate this process is one of the major issues for the rice field eel stocking. So far the genomics and transcriptomics data available for this species are still scarce. Here we provide a comprehensive study of transcriptomes of brain and gonad tissue in three sex stages (female, intersex and male from the rice field eel to investigate changes in transcriptional level during the sex reversal process.Approximately 195 thousand unigenes were generated and over 44.4 thousand were functionally annotated. Comparative study between stages provided multiple differentially expressed genes in brain and gonad tissue. Overall 4668 genes were found to be of unequal abundance between gonad tissues, far more than that of the brain tissues (59 genes. These genes were enriched in several different signaling pathways. A number of 231 genes were found with different levels in gonad in each stage, with several reproduction-related genes included. A total of 19 candidate genes that could be most related to sex reversal were screened out, part of these genes' expression patterns were validated by RT-qPCR. The expression of spef2, maats1, spag6 and dmc1 were abundant in testis, but was barely detected in females, while the 17β-hsd12, zpsbp3, gal3 and foxn5 were only expressed in ovary.This study investigated the complexity of brain and gonad transcriptomes in three sex stages of the rice field eel. Integrated analysis of different gene expression and changes in signaling pathways, such as PI3K-Akt pathway, provided crucial data for further study of sex transformation mechanisms.

  1. Comparative genomic reconstruction of transcriptional networks controlling central metabolism in the Shewanella genus

    Directory of Open Access Journals (Sweden)

    Kovaleva Galina

    2011-06-01

    Full Text Available Abstract Background Genome-scale prediction of gene regulation and reconstruction of transcriptional regulatory networks in bacteria is one of the critical tasks of modern genomics. The Shewanella genus is comprised of metabolically versatile gamma-proteobacteria, whose lifestyles and natural environments are substantially different from Escherichia coli and other model bacterial species. The comparative genomics approaches and computational identification of regulatory sites are useful for the in silico reconstruction of transcriptional regulatory networks in bacteria. Results To explore conservation and variations in the Shewanella transcriptional networks we analyzed the repertoire of transcription factors and performed genomics-based reconstruction and comparative analysis of regulons in 16 Shewanella genomes. The inferred regulatory network includes 82 transcription factors and their DNA binding sites, 8 riboswitches and 6 translational attenuators. Forty five regulons were newly inferred from the genome context analysis, whereas others were propagated from previously characterized regulons in the Enterobacteria and Pseudomonas spp.. Multiple variations in regulatory strategies between the Shewanella spp. and E. coli include regulon contraction and expansion (as in the case of PdhR, HexR, FadR, numerous cases of recruiting non-orthologous regulators to control equivalent pathways (e.g. PsrA for fatty acid degradation and, conversely, orthologous regulators to control distinct pathways (e.g. TyrR, ArgR, Crp. Conclusions We tentatively defined the first reference collection of ~100 transcriptional regulons in 16 Shewanella genomes. The resulting regulatory network contains ~600 regulated genes per genome that are mostly involved in metabolism of carbohydrates, amino acids, fatty acids, vitamins, metals, and stress responses. Several reconstructed regulons including NagR for N-acetylglucosamine catabolism were experimentally validated in S

  2. TIGER: Toolbox for integrating genome-scale metabolic models, expression data, and transcriptional regulatory networks

    Directory of Open Access Journals (Sweden)

    Jensen Paul A

    2011-09-01

    Full Text Available Abstract Background Several methods have been developed for analyzing genome-scale models of metabolism and transcriptional regulation. Many of these methods, such as Flux Balance Analysis, use constrained optimization to predict relationships between metabolic flux and the genes that encode and regulate enzyme activity. Recently, mixed integer programming has been used to encode these gene-protein-reaction (GPR relationships into a single optimization problem, but these techniques are often of limited generality and lack a tool for automating the conversion of rules to a coupled regulatory/metabolic model. Results We present TIGER, a Toolbox for Integrating Genome-scale Metabolism, Expression, and Regulation. TIGER converts a series of generalized, Boolean or multilevel rules into a set of mixed integer inequalities. The package also includes implementations of existing algorithms to integrate high-throughput expression data with genome-scale models of metabolism and transcriptional regulation. We demonstrate how TIGER automates the coupling of a genome-scale metabolic model with GPR logic and models of transcriptional regulation, thereby serving as a platform for algorithm development and large-scale metabolic analysis. Additionally, we demonstrate how TIGER's algorithms can be used to identify inconsistencies and improve existing models of transcriptional regulation with examples from the reconstructed transcriptional regulatory network of Saccharomyces cerevisiae. Conclusion The TIGER package provides a consistent platform for algorithm development and extending existing genome-scale metabolic models with regulatory networks and high-throughput data.

  3. Modeling Ebola Virus Genome Replication and Transcription with Minigenome Systems.

    Science.gov (United States)

    Cressey, Tessa; Brauburger, Kristina; Mühlberger, Elke

    2017-01-01

    In this chapter, we describe the minigenome system for Ebola virus (EBOV), which reconstitutes EBOV polymerase activity in cells and can be used to model viral genome replication and transcription. This protocol comprises all steps including cell culture, plasmid preparation, transfection, and luciferase reporter assay readout.

  4. Genome-wide investigation of transcription factors provides insights into transcriptional regulation in Plutella xylostella.

    Science.gov (United States)

    Zhao, Qian; Ma, Dongna; Huang, Yuping; He, Weiyi; Li, Yiying; Vasseur, Liette; You, Minsheng

    2018-04-01

    Transcription factors (TFs), which play a vital role in regulating gene expression, are prevalent in all organisms and characterization of them may provide important clues for understanding regulation in vivo. The present study reports a genome-wide investigation of TFs in the diamondback moth, Plutella xylostella (L.), a worldwide pest of crucifers. A total of 940 TFs distributed among 133 families were identified. Phylogenetic analysis of insect species showed that some of these families were found to have expanded during the evolution of P. xylostella or Lepidoptera. RNA-seq analysis showed that some of the TF families, such as zinc fingers, homeobox, bZIP, bHLH, and MADF_DNA_bdg genes, were highly expressed in certain tissues including midgut, salivary glands, fat body, and hemocytes, with an obvious sex-biased expression pattern. In addition, a number of TFs showed significant differences in expression between insecticide susceptible and resistant strains, suggesting that these TFs play a role in regulating genes related to insecticide resistance. Finally, we identified an expansion of the HOX cluster in Lepidoptera, which might be related to Lepidoptera-specific evolution. Knockout of this cluster using CRISPR/Cas9 showed that the egg cannot hatch, indicating that this cluster may be related to egg development and maturation. This is the first comprehensive study on identifying and characterizing TFs in P. xylostella. Our results suggest that some TF families are expanded in the P. xylostella genome, and these TFs may have important biological roles in growth, development, sexual dimorphism, and resistance to insecticides. The present work provides a solid foundation for understanding regulation via TFs in P. xylostella and insights into the evolution of the P. xylostella genome.

  5. Genomic context drives transcription of insertion sequences in the bacterial endosymbiont Wolbachia wVulC.

    Science.gov (United States)

    Cerveau, Nicolas; Gilbert, Clément; Liu, Chao; Garrett, Roger A; Grève, Pierre; Bouchon, Didier; Cordaux, Richard

    2015-06-10

    Transposable elements (TEs) are DNA pieces that are present in almost all the living world at variable genomic density. Due to their mobility and density, TEs are involved in a large array of genomic modifications. In eukaryotes, TE expression has been studied in detail in several species. In prokaryotes, studies of IS expression are generally linked to particular copies that induce a modification of neighboring gene expression. Here we investigated global patterns of IS transcription in the Alphaproteobacterial endosymbiont Wolbachia wVulC, using both RT-PCR and bioinformatic analyses. We detected several transcriptional promoters in all IS groups. Nevertheless, only one of the potentially functional IS groups possesses a promoter located upstream of the transposase gene, that could lead up to the production of a functional protein. We found that the majority of IS groups are expressed whatever their functional status. RT-PCR analyses indicate that the transcription of two IS groups lacking internal promoters upstream of the transposase start codon may be driven by the genomic environment. We confirmed this observation with the transcription analysis of individual copies of one IS group. These results suggest that the genomic environment is important for IS expression and it could explain, at least partly, copy number variability of the various IS groups present in the wVulC genome and, more generally, in bacterial genomes. Copyright © 2015 Elsevier B.V. All rights reserved.

  6. Targeted genome regulation via synthetic programmable transcriptional regulators

    KAUST Repository

    Piatek, Agnieszka Anna

    2016-04-19

    Regulation of gene transcription controls cellular functions and coordinates responses to developmental, physiological and environmental cues. Precise and efficient molecular tools are needed to characterize the functions of single and multiple genes in linear and interacting pathways in a native context. Modular DNA-binding domains from zinc fingers (ZFs) and transcriptional activator-like proteins (TALE) are amenable to bioengineering to bind DNA target sequences of interest. As a result, ZF and TALE proteins were used to develop synthetic programmable transcription factors. However, these systems are limited by the requirement to re-engineer proteins for each new target sequence. The clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR associated 9 (Cas9) genome editing tool was recently repurposed for targeted transcriptional regulation by inactivation of the nuclease activity of Cas9. Due to the facile engineering, simplicity, precision and amenability to library construction, the CRISPR/Cas9 system is poised to revolutionize the functional genomics field across diverse eukaryotic species. In this review, we discuss the development of synthetic customizable transcriptional regulators and provide insights into their current and potential applications, with special emphasis on plant systems, in characterization of gene functions, elucidation of molecular mechanisms and their biotechnological applications. © 2016 Informa UK Limited, trading as Taylor & Francis Group

  7. Nanobody®-based chromatin immunoprecipitation/micro-array analysis for genome-wide identification of transcription factor DNA binding sites

    Science.gov (United States)

    Nguyen-Duc, Trong; Peeters, Eveline; Muyldermans, Serge; Charlier, Daniel; Hassanzadeh-Ghassabeh, Gholamreza

    2013-01-01

    Nanobodies® are single-domain antibody fragments derived from camelid heavy-chain antibodies. Because of their small size, straightforward production in Escherichia coli, easy tailoring, high affinity, specificity, stability and solubility, nanobodies® have been exploited in various biotechnological applications. A major challenge in the post-genomics and post-proteomics era is the identification of regulatory networks involving nucleic acid–protein and protein–protein interactions. Here, we apply a nanobody® in chromatin immunoprecipitation followed by DNA microarray hybridization (ChIP-chip) for genome-wide identification of DNA–protein interactions. The Lrp-like regulator Ss-LrpB, arguably one of the best-studied specific transcription factors of the hyperthermophilic archaeon Sulfolobus solfataricus, was chosen for this proof-of-principle nanobody®-assisted ChIP. Three distinct Ss-LrpB-specific nanobodies®, each interacting with a different epitope, were generated for ChIP. Genome-wide ChIP-chip with one of these nanobodies® identified the well-established Ss-LrpB binding sites and revealed several unknown target sequences. Furthermore, these ChIP-chip profiles revealed auxiliary operator sites in the open reading frame of Ss-lrpB. Our work introduces nanobodies® as a novel class of affinity reagents for ChIP. Taking into account the unique characteristics of nanobodies®, in particular, their short generation time, nanobody®-based ChIP is expected to further streamline ChIP-chip and ChIP-Seq experiments, especially in organisms with no (or limited) possibility of genetic manipulation. PMID:23275538

  8. Identification of genome-specific transcripts in wheat–rye translocation lines

    Directory of Open Access Journals (Sweden)

    Tong Geon Lee

    2015-09-01

    Full Text Available Studying gene expression in wheat–rye translocation lines is complicated due to the presence of homeologs in hexaploid wheat and high levels of synteny between wheat and rye genomes (Naranjo and Fernandez-Rueda, 1991 [1]; Devos et al., 1995 [2]; Lee et al., 2010 [3]; Lee et al., 2013 [4]. To overcome limitations of current gene expression studies on wheat–rye translocation lines and identify genome-specific transcripts, we developed a custom Roche NimbleGen Gene Expression microarray that contains probes derived from the sequence of hexaploid wheat, diploid rye and diploid progenitors of hexaploid wheat genome (Lee et al., 2014. Using the array developed, we identified genome-specific transcripts in a wheat–rye translocation line (Lee et al., 2014. Expression data are deposited in the NCBI Gene Expression Omnibus (GEO under accession number GSE58678. Here we report the details of the methods used in the array workflow and data analysis.

  9. Hematopoietic transcriptional mechanisms: from locus-specific to genome-wide vantage points.

    Science.gov (United States)

    DeVilbiss, Andrew W; Sanalkumar, Rajendran; Johnson, Kirby D; Keles, Sunduz; Bresnick, Emery H

    2014-08-01

    Hematopoiesis is an exquisitely regulated process in which stem cells in the developing embryo and the adult generate progenitor cells that give rise to all blood lineages. Master regulatory transcription factors control hematopoiesis by integrating signals from the microenvironment and dynamically establishing and maintaining genetic networks. One of the most rudimentary aspects of cell type-specific transcription factor function, how they occupy a highly restricted cohort of cis-elements in chromatin, remains poorly understood. Transformative technologic advances involving the coupling of next-generation DNA sequencing technology with the chromatin immunoprecipitation assay (ChIP-seq) have enabled genome-wide mapping of factor occupancy patterns. However, formidable problems remain; notably, ChIP-seq analysis yields hundreds to thousands of chromatin sites occupied by a given transcription factor, and only a fraction of the sites appear to be endowed with critical, non-redundant function. It has become en vogue to map transcription factor occupancy patterns genome-wide, while using powerful statistical tools to establish correlations to inform biology and mechanisms. With the advent of revolutionary genome editing technologies, one can now reach beyond correlations to conduct definitive hypothesis testing. This review focuses on key discoveries that have emerged during the path from single loci to genome-wide analyses, specifically in the context of hematopoietic transcriptional mechanisms. Copyright © 2014 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.

  10. Regulatory hotspots in the malaria parasite genome dictate transcriptional variation.

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    Joseph M Gonzales

    2008-09-01

    Full Text Available The determinants of transcriptional regulation in malaria parasites remain elusive. The presence of a well-characterized gene expression cascade shared by different Plasmodium falciparum strains could imply that transcriptional regulation and its natural variation do not contribute significantly to the evolution of parasite drug resistance. To clarify the role of transcriptional variation as a source of stain-specific diversity in the most deadly malaria species and to find genetic loci that dictate variations in gene expression, we examined genome-wide expression level polymorphisms (ELPs in a genetic cross between phenotypically distinct parasite clones. Significant variation in gene expression is observed through direct co-hybridizations of RNA from different P. falciparum clones. Nearly 18% of genes were regulated by a significant expression quantitative trait locus. The genetic determinants of most of these ELPs resided in hotspots that are physically distant from their targets. The most prominent regulatory locus, influencing 269 transcripts, coincided with a Chromosome 5 amplification event carrying the drug resistance gene, pfmdr1, and 13 other genes. Drug selection pressure in the Dd2 parental clone lineage led not only to a copy number change in the pfmdr1 gene but also to an increased copy number of putative neighboring regulatory factors that, in turn, broadly influence the transcriptional network. Previously unrecognized transcriptional variation, controlled by polymorphic regulatory genes and possibly master regulators within large copy number variants, contributes to sweeping phenotypic evolution in drug-resistant malaria parasites.

  11. Introns Protect Eukaryotic Genomes from Transcription-Associated Genetic Instability.

    Science.gov (United States)

    Bonnet, Amandine; Grosso, Ana R; Elkaoutari, Abdessamad; Coleno, Emeline; Presle, Adrien; Sridhara, Sreerama C; Janbon, Guilhem; Géli, Vincent; de Almeida, Sérgio F; Palancade, Benoit

    2017-08-17

    Transcription is a source of genetic instability that can notably result from the formation of genotoxic DNA:RNA hybrids, or R-loops, between the nascent mRNA and its template. Here we report an unexpected function for introns in counteracting R-loop accumulation in eukaryotic genomes. Deletion of endogenous introns increases R-loop formation, while insertion of an intron into an intronless gene suppresses R-loop accumulation and its deleterious impact on transcription and recombination in yeast. Recruitment of the spliceosome onto the mRNA, but not splicing per se, is shown to be critical to attenuate R-loop formation and transcription-associated genetic instability. Genome-wide analyses in a number of distant species differing in their intron content, including human, further revealed that intron-containing genes and the intron-richest genomes are best protected against R-loop accumulation and subsequent genetic instability. Our results thereby provide a possible rationale for the conservation of introns throughout the eukaryotic lineage. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Transcriptional and chromatin regulation during fasting – The genomic era

    Science.gov (United States)

    Goldstein, Ido; Hager, Gordon L.

    2015-01-01

    An elaborate metabolic response to fasting is orchestrated by the liver and is heavily reliant upon transcriptional regulation. In response to hormones (glucagon, glucocorticoids) many transcription factors (TFs) are activated and regulate various genes involved in metabolic pathways aimed at restoring homeostasis: gluconeogenesis, fatty acid oxidation, ketogenesis and amino acid shuttling. We summarize the recent discoveries regarding fasting-related TFs with an emphasis on genome-wide binding patterns. Collectively, the summarized findings reveal a large degree of co-operation between TFs during fasting which occurs at motif-rich DNA sites bound by a combination of TFs. These new findings implicate transcriptional and chromatin regulation as major determinants of the response to fasting and unravels the complex, multi-TF nature of this response. PMID:26520657

  13. Enhancing yeast transcription analysis through integration of heterogeneous data

    DEFF Research Database (Denmark)

    Grotkjær, Thomas; Nielsen, Jens

    2004-01-01

    of Saccharomyces cerevisiae whole genome transcription data. A special focus is on the quantitative aspects of normalisation and mathematical modelling approaches, since they are expected to play an increasing role in future DNA microarray analysis studies. Data analysis is exemplified with cluster analysis......DNA microarray technology enables the simultaneous measurement of the transcript level of thousands of genes. Primary analysis can be done with basic statistical tools and cluster analysis, but effective and in depth analysis of the vast amount of transcription data requires integration with data...... from several heterogeneous data Sources, such as upstream promoter sequences, genome-scale metabolic models, annotation databases and other experimental data. In this review, we discuss how experimental design, normalisation, heterogeneous data and mathematical modelling can enhance analysis...

  14. Global transcriptional analysis of psoriatic skin and blood confirms known disease-associated pathways and highlights novel genomic "hot spots" for differentially expressed genes.

    Science.gov (United States)

    Coda, Alvin B; Icen, Murat; Smith, Jason R; Sinha, Animesh A

    2012-07-01

    There are major gaps in our knowledge regarding the exact mechanisms and genetic basis of psoriasis. To investigate the pathogenesis of psoriasis, gene expression in 10 skin (5 lesional, 5 nonlesional) and 11 blood (6 psoriatic, 5 nonpsoriatic) samples were examined using Affymetrix HG-U95A microarrays. We detected 535 (425 upregulated, 110 downregulated) DEGs in lesional skin at 1% false discovery rate (FDR). Combining nine microarray studies comparing lesional and nonlesional psoriatic skin, 34.5% of dysregulated genes were overlapped in multiple studies. We further identified 20 skin and 2 blood associated transcriptional "hot spots" at specified genomic locations. At 5% FDR, 11.8% skin and 10.4% blood DEGs in our study mapped to one of the 12 PSORS loci. DEGs that overlap with PSORS loci may offer prioritized targets for downstream genetic fine mapping studies. Novel DEG "hot spots" may provide new targets for defining susceptibility loci in future studies. Copyright © 2012 Elsevier Inc. All rights reserved.

  15. Reporter-Based Synthetic Genetic Array Analysis: A Functional Genomics Approach for Investigating Transcript or Protein Abundance Using Fluorescent Proteins in Saccharomyces cerevisiae.

    Science.gov (United States)

    Göttert, Hendrikje; Mattiazzi Usaj, Mojca; Rosebrock, Adam P; Andrews, Brenda J

    2018-01-01

    Fluorescent reporter genes have long been used to quantify various cell features such as transcript and protein abundance. Here, we describe a method, reporter synthetic genetic array (R-SGA) analysis, which allows for the simultaneous quantification of any fluorescent protein readout in thousands of yeast strains using an automated pipeline. R-SGA combines a fluorescent reporter system with standard SGA analysis and can be used to examine any array-based strain collection available to the yeast community. This protocol describes the R-SGA methodology for screening different arrays of yeast mutants including the deletion collection, a collection of temperature-sensitive strains for the assessment of essential yeast genes and a collection of inducible overexpression strains. We also present an alternative pipeline for the analysis of R-SGA output strains using flow cytometry of cells in liquid culture. Data normalization for both pipelines is discussed.

  16. Transcriptional Slippage and RNA Editing Increase the Diversity of Transcripts in Chloroplasts: Insight from Deep Sequencing of Vigna radiata Genome and Transcriptome.

    Directory of Open Access Journals (Sweden)

    Ching-Ping Lin

    Full Text Available We performed deep sequencing of the nuclear and organellar genomes of three mungbean genotypes: Vigna radiata ssp. sublobata TC1966, V. radiata var. radiata NM92 and the recombinant inbred line RIL59 derived from a cross between TC1966 and NM92. Moreover, we performed deep sequencing of the RIL59 transcriptome to investigate transcript variability. The mungbean chloroplast genome has a quadripartite structure including a pair of inverted repeats separated by two single copy regions. A total of 213 simple sequence repeats were identified in the chloroplast genomes of NM92 and RIL59; 78 single nucleotide variants and nine indels were discovered in comparing the chloroplast genomes of TC1966 and NM92. Analysis of the mungbean chloroplast transcriptome revealed mRNAs that were affected by transcriptional slippage and RNA editing. Transcriptional slippage frequency was positively correlated with the length of simple sequence repeats of the mungbean chloroplast genome (R2=0.9911. In total, 41 C-to-U editing sites were found in 23 chloroplast genes and in one intergenic spacer. No editing site that swapped U to C was found. A combination of bioinformatics and experimental methods revealed that the plastid-encoded RNA polymerase-transcribed genes psbF and ndhA are affected by transcriptional slippage in mungbean and in main lineages of land plants, including three dicots (Glycine max, Brassica rapa, and Nicotiana tabacum, two monocots (Oryza sativa and Zea mays, two gymnosperms (Pinus taeda and Ginkgo biloba and one moss (Physcomitrella patens. Transcript analysis of the rps2 gene showed that transcriptional slippage could affect transcripts at single sequence repeat regions with poly-A runs. It showed that transcriptional slippage together with incomplete RNA editing may cause sequence diversity of transcripts in chloroplasts of land plants.

  17. Evidence for site-specific occupancy of the mitochondrial genome by nuclear transcription factors.

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    Georgi K Marinov

    Full Text Available Mitochondria contain their own circular genome, with mitochondria-specific transcription and replication systems and corresponding regulatory proteins. All of these proteins are encoded in the nuclear genome and are post-translationally imported into mitochondria. In addition, several nuclear transcription factors have been reported to act in mitochondria, but there has been no comprehensive mapping of their occupancy patterns and it is not clear how many other factors may also be found in mitochondria. Here we address these questions by using ChIP-seq data from the ENCODE, mouseENCODE and modENCODE consortia for 151 human, 31 mouse and 35 C. elegans factors. We identified 8 human and 3 mouse transcription factors with strong localized enrichment over the mitochondrial genome that was usually associated with the corresponding recognition sequence motif. Notably, these sites of occupancy are often the sites with highest ChIP-seq signal intensity within both the nuclear and mitochondrial genomes and are thus best explained as true binding events to mitochondrial DNA, which exist in high copy number in each cell. We corroborated these findings by immunocytochemical staining evidence for mitochondrial localization. However, we were unable to find clear evidence for mitochondrial binding in ENCODE and other publicly available ChIP-seq data for most factors previously reported to localize there. As the first global analysis of nuclear transcription factors binding in mitochondria, this work opens the door to future studies that probe the functional significance of the phenomenon.

  18. Genome-wide transcriptional reprogramming under drought stress

    KAUST Repository

    Chen, Hao

    2012-01-01

    Soil water deficit is one of the major factors limiting plant productivity. Plants cope with this adverse environmental condition by coordinating the up- or downregulation of an array of stress responsive genes. Reprogramming the expression of these genes leads to rebalanced development and growth that are in concert with the reduced water availability and that ultimately confer enhanced stress tolerance. Currently, several techniques have been employed to monitor genome-wide transcriptional reprogramming under drought stress. The results from these high throughput studies indicate that drought stress-induced transcriptional reprogramming is dynamic, has temporal and spatial specificity, and is coupled with the circadian clock and phytohormone signaling pathways. © 2012 Springer-Verlag Berlin Heidelberg. All rights are reserved.

  19. Genome-wide identification, classification, and functional analysis of the basic helix-loop-helix transcription factors in the cattle, Bos Taurus.

    Science.gov (United States)

    Li, Fengmei; Liu, Wuyi

    2017-06-01

    The basic helix-loop-helix (bHLH) transcription factors (TFs) form a huge superfamily and play crucial roles in many essential developmental, genetic, and physiological-biochemical processes of eukaryotes. In total, 109 putative bHLH TFs were identified and categorized successfully in the genomic databases of cattle, Bos Taurus, after removing redundant sequences and merging genetic isoforms. Through phylogenetic analyses, 105 proteins among these bHLH TFs were classified into 44 families with 46, 25, 14, 3, 13, and 4 members in the high-order groups A, B, C, D, E, and F, respectively. The remaining 4 bHLH proteins were sorted out as 'orphans.' Next, these 109 putative bHLH proteins identified were further characterized as significantly enriched in 524 significant Gene Ontology (GO) annotations (corrected P value ≤ 0.05) and 21 significantly enriched pathways (corrected P value ≤ 0.05) that had been mapped by the web server KOBAS 2.0. Furthermore, 95 bHLH proteins were further screened and analyzed together with two uncharacterized proteins in the STRING online database to reconstruct the protein-protein interaction network of cattle bHLH TFs. Ultimately, 89 bHLH proteins were fully mapped in a network with 67 biological process, 13 molecular functions, 5 KEGG pathways, 12 PFAM protein domains, and 25 INTERPRO classified protein domains and features. These results provide much useful information and a good reference for further functional investigations and updated researches on cattle bHLH TFs.

  20. Human Metapneumovirus Induces Formation of Inclusion Bodies for Efficient Genome Replication and Transcription.

    Science.gov (United States)

    Cifuentes-Muñoz, Nicolás; Branttie, Jean; Slaughter, Kerri Beth; Dutch, Rebecca Ellis

    2017-12-15

    Human metapneumovirus (HMPV) causes significant upper and lower respiratory disease in all age groups worldwide. The virus possesses a negative-sense single-stranded RNA genome of approximately 13.3 kb encapsidated by multiple copies of the nucleoprotein (N), giving rise to helical nucleocapsids. In addition, copies of the phosphoprotein (P) and the large RNA polymerase (L) decorate the viral nucleocapsids. After viral attachment, endocytosis, and fusion mediated by the viral glycoproteins, HMPV nucleocapsids are released into the cell cytoplasm. To visualize the subsequent steps of genome transcription and replication, a fluorescence in situ hybridization (FISH) protocol was established to detect different viral RNA subpopulations in infected cells. The FISH probes were specific for detection of HMPV positive-sense RNA (+RNA) and viral genomic RNA (vRNA). Time course analysis of human bronchial epithelial BEAS-2B cells infected with HMPV revealed the formation of inclusion bodies (IBs) from early times postinfection. HMPV IBs were shown to be cytoplasmic sites of active transcription and replication, with the translation of viral proteins being closely associated. Inclusion body formation was consistent with an actin-dependent coalescence of multiple early replicative sites. Time course quantitative reverse transcription-PCR analysis suggested that the coalescence of inclusion bodies is a strategy to efficiently replicate and transcribe the viral genome. These results provide a better understanding of the steps following HMPV entry and have important clinical implications. IMPORTANCE Human metapneumovirus (HMPV) is a recently discovered pathogen that affects human populations of all ages worldwide. Reinfections are common throughout life, but no vaccines or antiviral treatments are currently available. In this work, a spatiotemporal analysis of HMPV replication and transcription in bronchial epithelial cell-derived immortal cells was performed. HMPV was shown to

  1. Genome Wide Identification and Characterization of Apple bHLH Transcription Factors and Expression Analysis in Response to Drought and Salt Stress.

    Science.gov (United States)

    Mao, Ke; Dong, Qinglong; Li, Chao; Liu, Changhai; Ma, Fengwang

    2017-01-01

    The bHLH (basic helix-loop-helix) transcription factor family is the second largest in plants. It occurs in all three eukaryotic kingdoms, and plays important roles in regulating growth and development. However, family members have not previously been studied in apple. Here, we identified 188 MdbHLH proteins in apple "Golden Delicious" ( Malus × domestica Borkh.), which could be classified into 18 groups. We also investigated the gene structures and 12 conserved motifs in these MdbHLH s. Coupled with expression analysis and protein interaction network prediction, we identified several genes that might be responsible for abiotic stress responses. This study provides insight and rich resources for subsequent investigations of such proteins in apple.

  2. Transcription profile of Escherichia coli: genomic SELEX search for regulatory targets of transcription factors.

    Science.gov (United States)

    Ishihama, Akira; Shimada, Tomohiro; Yamazaki, Yukiko

    2016-03-18

    Bacterial genomes are transcribed by DNA-dependent RNA polymerase (RNAP), which achieves gene selectivity through interaction with sigma factors that recognize promoters, and transcription factors (TFs) that control the activity and specificity of RNAP holoenzyme. To understand the molecular mechanisms of transcriptional regulation, the identification of regulatory targets is needed for all these factors. We then performed genomic SELEX screenings of targets under the control of each sigma factor and each TF. Here we describe the assembly of 156 SELEX patterns of a total of 116 TFs performed in the presence and absence of effector ligands. The results reveal several novel concepts: (i) each TF regulates more targets than hitherto recognized; (ii) each promoter is regulated by more TFs than hitherto recognized; and (iii) the binding sites of some TFs are located within operons and even inside open reading frames. The binding sites of a set of global regulators, including cAMP receptor protein, LeuO and Lrp, overlap with those of the silencer H-NS, suggesting that certain global regulators play an anti-silencing role. To facilitate sharing of these accumulated SELEX datasets with the research community, we compiled a database, 'Transcription Profile of Escherichia coli' (www.shigen.nig.ac.jp/ecoli/tec/). © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  3. Genomic dissection of conserved transcriptional regulation in intestinal epithelial cells.

    Directory of Open Access Journals (Sweden)

    Colin R Lickwar

    2017-08-01

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

  4. Nitrogen fixation and molecular oxygen: comparative genomic reconstruction of transcription regulation in Alphaproteobacteria

    Directory of Open Access Journals (Sweden)

    Olga V Tsoy

    2016-08-01

    Full Text Available Biological nitrogen fixation plays a crucial role in the nitrogen cycle. An ability to fix atmospheric nitrogen, reducing it to ammonium, was described for multiple species of Bacteria and Archaea. Being a complex and sensitive process, nitrogen fixation requires a complicated regulatory system, also, on the level of transcription. The transcriptional regulatory network for nitrogen fixation was extensively studied in several representatives of the class Alphaproteobacteria. This regulatory network includes the activator of nitrogen fixation NifA, working in tandem with the alternative sigma-factor RpoN as well as oxygen-responsive regulatory systems, one-component regulators FnrN/FixK and two-component system FixLJ. Here we used a comparative genomics analysis for in silico study of the transcriptional regulatory network in 50 genomes of Alphaproteobacteria. We extended the known regulons and proposed the scenario for the evolution of the nitrogen fixation transcriptional network. The reconstructed network substantially expands the existing knowledge of transcriptional regulation in nitrogen-fixing microorganisms and can be used for genetic experiments, metabolic reconstruction, and evolutionary analysis.

  5. Whole genome duplications and expansion of the vertebrate GATA transcription factor gene family

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    Bowerman Bruce

    2009-08-01

    Full Text Available Abstract Background GATA transcription factors influence many developmental processes, including the specification of embryonic germ layers. The GATA gene family has significantly expanded in many animal lineages: whereas diverse cnidarians have only one GATA transcription factor, six GATA genes have been identified in many vertebrates, five in many insects, and eleven to thirteen in Caenorhabditis nematodes. All bilaterian animal genomes have at least one member each of two classes, GATA123 and GATA456. Results We have identified one GATA123 gene and one GATA456 gene from the genomic sequence of two invertebrate deuterostomes, a cephalochordate (Branchiostoma floridae and a hemichordate (Saccoglossus kowalevskii. We also have confirmed the presence of six GATA genes in all vertebrate genomes, as well as additional GATA genes in teleost fish. Analyses of conserved sequence motifs and of changes to the exon-intron structure, and molecular phylogenetic analyses of these deuterostome GATA genes support their origin from two ancestral deuterostome genes, one GATA 123 and one GATA456. Comparison of the conserved genomic organization across vertebrates identified eighteen paralogous gene families linked to multiple vertebrate GATA genes (GATA paralogons, providing the strongest evidence yet for expansion of vertebrate GATA gene families via genome duplication events. Conclusion From our analysis, we infer the evolutionary birth order and relationships among vertebrate GATA transcription factors, and define their expansion via multiple rounds of whole genome duplication events. As the genomes of four independent invertebrate deuterostome lineages contain single copy GATA123 and GATA456 genes, we infer that the 0R (pre-genome duplication invertebrate deuterostome ancestor also had two GATA genes, one of each class. Synteny analyses identify duplications of paralogous chromosomal regions (paralogons, from single ancestral vertebrate GATA123 and GATA456

  6. Genome-wide analysis of the HD-ZIP IV transcription factor family in Gossypium arboreum and GaHDG11 involved in osmotic tolerance in transgenic Arabidopsis.

    Science.gov (United States)

    Chen, Eryong; Zhang, Xueyan; Yang, Zhaoen; Wang, Xiaoqian; Yang, Zuoren; Zhang, Chaojun; Wu, Zhixia; Kong, Depei; Liu, Zhao; Zhao, Ge; Butt, Hamama Islam; Zhang, Xianlong; Li, Fuguang

    2017-06-01

    HD-ZIP IV proteins belong to the homeodomain-leucine zipper (HD-ZIP) transcription factor family and are involved in trichome development and drought stress in plants. Although some functions of the HD-ZIP IV group are well understood in Arabidopsis, little is known about their function in cotton. In this study, HD-ZIP genes were identified from three Gossypium species (G. arboreum, G. raimondii and G. hirsutum) and clustered into four families (HD-ZIP I, II, III and IV) to separate HD-ZIP IV from the other three families. Systematic analyses of phylogeny, gene structure, conserved domains, and expression profiles in different plant tissues and the expression patterns under osmotic stress in leaves were further conducted in G. arboreum. More importantly, ectopic overexpression of GaHDG11, a representative of the HD-ZIP IV family, confers enhanced osmotic tolerance in transgenic Arabidopsis plants, possibly due to elongated primary root length, lower water loss rates, high osmoprotectant proline levels, significant levels of antioxidants CAT, and/or SOD enzyme activity with reduced levels of MDA. Taken together, these observations may lay the foundation for future functional analysis of cotton HD-ZIP IV genes to unravel their biological roles in cotton.

  7. Production and processing of siRNA precursor transcripts from the highly repetitive maize genome.

    Directory of Open Access Journals (Sweden)

    Christopher J Hale

    2009-08-01

    Full Text Available Mutations affecting the maintenance of heritable epigenetic states in maize identify multiple RNA-directed DNA methylation (RdDM factors including RMR1, a novel member of a plant-specific clade of Snf2-related proteins. Here we show that RMR1 is necessary for the accumulation of a majority of 24 nt small RNAs, including those derived from Long-Terminal Repeat (LTR retrotransposons, the most common repetitive feature in the maize genome. A genetic analysis of DNA transposon repression indicates that RMR1 acts upstream of the RNA-dependent RNA polymerase, RDR2 (MOP1. Surprisingly, we show that non-polyadenylated transcripts from a sampling of LTR retrotransposons are lost in both rmr1 and rdr2 mutants. In contrast, plants deficient for RNA Polymerase IV (Pol IV function show an increase in polyadenylated LTR RNA transcripts. These findings support a model in which Pol IV functions independently of the small RNA accumulation facilitated by RMR1 and RDR2 and support that a loss of Pol IV leads to RNA Polymerase II-based transcription. Additionally, the lack of changes in general genome homeostasis in rmr1 mutants, despite the global loss of 24 nt small RNAs, challenges the perceived roles of siRNAs in maintaining functional heterochromatin in the genomes of outcrossing grass species.

  8. Genome-wide conserved consensus transcription factor binding motifs are hyper-methylated

    Directory of Open Access Journals (Sweden)

    Down Thomas A

    2010-09-01

    Full Text Available Abstract Background DNA methylation can regulate gene expression by modulating the interaction between DNA and proteins or protein complexes. Conserved consensus motifs exist across the human genome ("predicted transcription factor binding sites": "predicted TFBS" but the large majority of these are proven by chromatin immunoprecipitation and high throughput sequencing (ChIP-seq not to be biological transcription factor binding sites ("empirical TFBS". We hypothesize that DNA methylation at conserved consensus motifs prevents promiscuous or disorderly transcription factor binding. Results Using genome-wide methylation maps of the human heart and sperm, we found that all conserved consensus motifs as well as the subset of those that reside outside CpG islands have an aggregate profile of hyper-methylation. In contrast, empirical TFBS with conserved consensus motifs have a profile of hypo-methylation. 40% of empirical TFBS with conserved consensus motifs resided in CpG islands whereas only 7% of all conserved consensus motifs were in CpG islands. Finally we further identified a minority subset of TF whose profiles are either hypo-methylated or neutral at their respective conserved consensus motifs implicating that these TF may be responsible for establishing or maintaining an un-methylated DNA state, or whose binding is not regulated by DNA methylation. Conclusions Our analysis supports the hypothesis that at least for a subset of TF, empirical binding to conserved consensus motifs genome-wide may be controlled by DNA methylation.

  9. The raccoon polyomavirus genome and tumor antigen transcription are stable and abundant in neuroglial tumors.

    Science.gov (United States)

    Brostoff, Terza; Dela Cruz, Florante N; Church, Molly E; Woolard, Kevin D; Pesavento, Patricia A

    2014-11-01

    Raccoon polyomavirus (RacPyV) is associated with 100% of neuroglial tumors in free-ranging raccoons. Other tumor-associated polyomaviruses (PyVs), including simian virus 40 (SV40), murine PyV, and Merkel cell PyV, are found integrated in the host genome in neoplastic cells, where they constitutively express splice variants of the tumor antigen (TAg) gene. We have previously reported that RacPyV exists only as an episome (nonintegrated) in neuroglial tumors. Here, we have investigated TAg transcription in primary tumor tissue by transcriptome analysis, and we identified the alternatively spliced TAg transcripts for RacPyV. We also determined that TAg was highly transcribed relative to host cellular genes. We further colocalized TAg DNA and mRNA by in situ hybridization and found that the majority of tumor cells showed positive staining. Lastly, we examined the stability of the viral genome and TAg transcription by quantitative reverse transcriptase PCR in cultured tumor cells in vitro and in a mouse xenograft model. When tumor cells were cultured in vitro, TAg transcription increased nearly 2 log-fold over that of parental tumor tissue by passage 17. Both episomal viral genome and TAg transcription were faithfully maintained in culture and in tumors arising from xenotransplantation of cultured cells in mice. This study represents a minimal criterion for RacPyV's association with neuroglial tumors and a novel mechanism of stability for a polyomavirus in cancer. The natural cycle of polyomaviruses in mammals is to persist in the host without causing disease, but they can cause cancer in humans or in other animals. Because this is an unpredictable and rare event, the oncogenic potential of polyomavirus is primarily evaluated in laboratory animal models. Recently, raccoon polyomavirus (RacPyV) was identified in neuroglial tumors of free-ranging raccoons. Viral copy number was consistently high in these tumors but was low or undetectable in nontumor tissue or in

  10. The Csr system regulates genome-wide mRNA stability and transcription and thus gene expression in Escherichia coli.

    Science.gov (United States)

    Esquerré, Thomas; Bouvier, Marie; Turlan, Catherine; Carpousis, Agamemnon J; Girbal, Laurence; Cocaign-Bousquet, Muriel

    2016-04-26

    Bacterial adaptation requires large-scale regulation of gene expression. We have performed a genome-wide analysis of the Csr system, which regulates many important cellular functions. The Csr system is involved in post-transcriptional regulation, but a role in transcriptional regulation has also been suggested. Two proteins, an RNA-binding protein CsrA and an atypical signaling protein CsrD, participate in the Csr system. Genome-wide transcript stabilities and levels were compared in wildtype E. coli (MG1655) and isogenic mutant strains deficient in CsrA or CsrD activity demonstrating for the first time that CsrA and CsrD are global negative and positive regulators of transcription, respectively. The role of CsrA in transcription regulation may be indirect due to the 4.6-fold increase in csrD mRNA concentration in the CsrA deficient strain. Transcriptional action of CsrA and CsrD on a few genes was validated by transcriptional fusions. In addition to an effect on transcription, CsrA stabilizes thousands of mRNAs. This is the first demonstration that CsrA is a global positive regulator of mRNA stability. For one hundred genes, we predict that direct control of mRNA stability by CsrA might contribute to metabolic adaptation by regulating expression of genes involved in carbon metabolism and transport independently of transcriptional regulation.

  11. Complete genomic and transcriptional landscape analysis using third-generation sequencing: a case study of Saccharomyces cerevisiae CEN.PK113-7D

    DEFF Research Database (Denmark)

    Jenjaroenpun, Piroon; Wongsurawat, Thidathip; Pereira, Rui

    2018-01-01

    Completion of eukaryal genomes can be difficult task with the highly repetitive sequences along the chromosomes and short read lengths of secondgeneration sequencing. Saccharomyces cerevisiae strain CEN. PK113-7D, widely used as a model organism and a cell factory, was selected for this study...... to demonstrate the superior capability of very long sequence reads for de novo genome assembly. We generated long reads using two common third-generation sequencing technologies (Oxford Nanopore Technology (ONT) and Pacific Biosciences (PacBio)) and used short reads obtained using Illumina sequencing for error...... correction. Assembly of the reads derived from all three technologies resulted in complete sequences for all 16 yeast chromosomes, as well as themitochondrial chromosome, in one step. Further, we identified three types of DNA methylation (5mC, 4mC and 6mA). Comparison between the reference strain S288C...

  12. Genome-Wide Identification of the Target Genes of AP2-O, a Plasmodium AP2-Family Transcription Factor.

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    Izumi Kaneko

    2015-05-01

    Full Text Available Stage-specific transcription is a fundamental biological process in the life cycle of the Plasmodium parasite. Proteins containing the AP2 DNA-binding domain are responsible for stage-specific transcriptional regulation and belong to the only known family of transcription factors in Plasmodium parasites. Comprehensive identification of their target genes will advance our understanding of the molecular basis of stage-specific transcriptional regulation and stage-specific parasite development. AP2-O is an AP2 family transcription factor that is expressed in the mosquito midgut-invading stage, called the ookinete, and is essential for normal morphogenesis of this stage. In this study, we identified the genome-wide target genes of AP2-O by chromatin immunoprecipitation-sequencing and elucidate how this AP2 family transcription factor contributes to the formation of this motile stage. The analysis revealed that AP2-O binds specifically to the upstream genomic regions of more than 500 genes, suggesting that approximately 10% of the parasite genome is directly regulated by AP2-O. These genes are involved in distinct biological processes such as morphogenesis, locomotion, midgut penetration, protection against mosquito immunity and preparation for subsequent oocyst development. This direct and global regulation by AP2-O provides a model for gene regulation in Plasmodium parasites and may explain how these parasites manage to control their complex life cycle using a small number of sequence-specific AP2 transcription factors.

  13. TSSer: an automated method to identify transcription start sites in prokaryotic genomes from differential RNA sequencing data.

    Science.gov (United States)

    Jorjani, Hadi; Zavolan, Mihaela

    2014-04-01

    Accurate identification of transcription start sites (TSSs) is an essential step in the analysis of transcription regulatory networks. In higher eukaryotes, the capped analysis of gene expression technology enabled comprehensive annotation of TSSs in genomes such as those of mice and humans. In bacteria, an equivalent approach, termed differential RNA sequencing (dRNA-seq), has recently been proposed, but the application of this approach to a large number of genomes is hindered by the paucity of computational analysis methods. With few exceptions, when the method has been used, annotation of TSSs has been largely done manually. In this work, we present a computational method called 'TSSer' that enables the automatic inference of TSSs from dRNA-seq data. The method rests on a probabilistic framework for identifying both genomic positions that are preferentially enriched in the dRNA-seq data as well as preferentially captured relative to neighboring genomic regions. Evaluating our approach for TSS calling on several publicly available datasets, we find that TSSer achieves high consistency with the curated lists of annotated TSSs, but identifies many additional TSSs. Therefore, TSSer can accelerate genome-wide identification of TSSs in bacterial genomes and can aid in further characterization of bacterial transcription regulatory networks. TSSer is freely available under GPL license at http://www.clipz.unibas.ch/TSSer/index.php

  14. Bivariate Genomic Footprinting Detects Changes in Transcription Factor Activity

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    Songjoon Baek

    2017-05-01

    Full Text Available In response to activating signals, transcription factors (TFs bind DNA and regulate gene expression. TF binding can be measured by protection of the bound sequence from DNase digestion (i.e., footprint. Here, we report that 80% of TF binding motifs do not show a measurable footprint, partly because of a variable cleavage pattern within the motif sequence. To more faithfully portray the effect of TFs on chromatin, we developed an algorithm that captures two TF-dependent effects on chromatin accessibility: footprinting and motif-flanking accessibility. The algorithm, termed bivariate genomic footprinting (BaGFoot, efficiently detects TF activity. BaGFoot is robust to different accessibility assays (DNase-seq, ATAC-seq, all examined peak-calling programs, and a variety of cut bias correction approaches. BaGFoot reliably predicts TF binding and provides valuable information regarding the TFs affecting chromatin accessibility in various biological systems and following various biological events, including in cases where an absolute footprint cannot be determined.

  15. Genome-wide transcription responses to synchrotron microbeam radiotherapy.

    Science.gov (United States)

    Sprung, Carl N; Yang, Yuqing; Forrester, Helen B; Li, Jason; Zaitseva, Marina; Cann, Leonie; Restall, Tina; Anderson, Robin L; Crosbie, Jeffrey C; Rogers, Peter A W

    2012-10-01

    The majority of cancer patients achieve benefit from radiotherapy. A significant limitation of radiotherapy is its relatively low therapeutic index, defined as the maximum radiation dose that causes acceptable normal tissue damage to the minimum dose required to achieve tumor control. Recently, a new radiotherapy modality using synchrotron-generated X-ray microbeam radiotherapy has been demonstrated in animal models to ablate tumors with concurrent sparing of normal tissue. Very little work has been undertaken into the cellular and molecular mechanisms that differentiate microbeam radiotherapy from broad beam. The purpose of this study was to investigate and compare the whole genome transcriptional response of in vivo microbeam radiotherapy versus broad beam irradiated tumors. We hypothesized that gene expression changes after microbeam radiotherapy are different from those seen after broad beam. We found that in EMT6.5 tumors at 4-48 h postirradiation, microbeam radiotherapy differentially regulates a number of genes, including major histocompatibility complex (MHC) class II antigen gene family members, and other immunity-related genes including Ciita, Ifng, Cxcl1, Cxcl9, Indo and Ubd when compared to broad beam. Our findings demonstrate molecular differences in the tumor response to microbeam versus broad beam irradiation and these differences provide insight into the underlying mechanisms of microbeam radiotherapy and broad beam.

  16. Transcription regulatory networks analysis using CAGE

    KAUST Repository

    Tegnér, Jesper N.

    2009-10-01

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

  17. Genome-wide transcriptional reorganization associated with senescence-to-immortality switch during human hepatocellular carcinogenesis.

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    Gokhan Yildiz

    Full Text Available Senescence is a permanent proliferation arrest in response to cell stress such as DNA damage. It contributes strongly to tissue aging and serves as a major barrier against tumor development. Most tumor cells are believed to bypass the senescence barrier (become "immortal" by inactivating growth control genes such as TP53 and CDKN2A. They also reactivate telomerase reverse transcriptase. Senescence-to-immortality transition is accompanied by major phenotypic and biochemical changes mediated by genome-wide transcriptional modifications. This appears to happen during hepatocellular carcinoma (HCC development in patients with liver cirrhosis, however, the accompanying transcriptional changes are virtually unknown. We investigated genome-wide transcriptional changes related to the senescence-to-immortality switch during hepatocellular carcinogenesis. Initially, we performed transcriptome analysis of senescent and immortal clones of Huh7 HCC cell line, and identified genes with significant differential expression to establish a senescence-related gene list. Through the analysis of senescence-related gene expression in different liver tissues we showed that cirrhosis and HCC display expression patterns compatible with senescent and immortal phenotypes, respectively; dysplasia being a transitional state. Gene set enrichment analysis revealed that cirrhosis/senescence-associated genes were preferentially expressed in non-tumor tissues, less malignant tumors, and differentiated or senescent cells. In contrast, HCC/immortality genes were up-regulated in tumor tissues, or more malignant tumors and progenitor cells. In HCC tumors and immortal cells genes involved in DNA repair, cell cycle, telomere extension and branched chain amino acid metabolism were up-regulated, whereas genes involved in cell signaling, as well as in drug, lipid, retinoid and glycolytic metabolism were down-regulated. Based on these distinctive gene expression features we developed a 15

  18. RegPrecise 3.0--a resource for genome-scale exploration of transcriptional regulation in bacteria.

    Science.gov (United States)

    Novichkov, Pavel S; Kazakov, Alexey E; Ravcheev, Dmitry A; Leyn, Semen A; Kovaleva, Galina Y; Sutormin, Roman A; Kazanov, Marat D; Riehl, William; Arkin, Adam P; Dubchak, Inna; Rodionov, Dmitry A

    2013-11-01

    Genome-scale prediction of gene regulation and reconstruction of transcriptional regulatory networks in prokaryotes is one of the critical tasks of modern genomics. Bacteria from different taxonomic groups, whose lifestyles and natural environments are substantially different, possess highly diverged transcriptional regulatory networks. The comparative genomics approaches are useful for in silico reconstruction of bacterial regulons and networks operated by both transcription factors (TFs) and RNA regulatory elements (riboswitches). RegPrecise (http://regprecise.lbl.gov) is a web resource for collection, visualization and analysis of transcriptional regulons reconstructed by comparative genomics. We significantly expanded a reference collection of manually curated regulons we introduced earlier. RegPrecise 3.0 provides access to inferred regulatory interactions organized by phylogenetic, structural and functional properties. Taxonomy-specific collections include 781 TF regulogs inferred in more than 160 genomes representing 14 taxonomic groups of Bacteria. TF-specific collections include regulogs for a selected subset of 40 TFs reconstructed across more than 30 taxonomic lineages. Novel collections of regulons operated by RNA regulatory elements (riboswitches) include near 400 regulogs inferred in 24 bacterial lineages. RegPrecise 3.0 provides four classifications of the reference regulons implemented as controlled vocabularies: 55 TF protein families; 43 RNA motif families; ~150 biological processes or metabolic pathways; and ~200 effectors or environmental signals. Genome-wide visualization of regulatory networks and metabolic pathways covered by the reference regulons are available for all studied genomes. A separate section of RegPrecise 3.0 contains draft regulatory networks in 640 genomes obtained by an conservative propagation of the reference regulons to closely related genomes. RegPrecise 3.0 gives access to the transcriptional regulons reconstructed in

  19. The Role of Genome Accessibility in Transcription Factor Binding in Bacteria.

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    Antonio L C Gomes

    2016-04-01

    Full Text Available ChIP-seq enables genome-scale identification of regulatory regions that govern gene expression. However, the biological insights generated from ChIP-seq analysis have been limited to predictions of binding sites and cooperative interactions. Furthermore, ChIP-seq data often poorly correlate with in vitro measurements or predicted motifs, highlighting that binding affinity alone is insufficient to explain transcription factor (TF-binding in vivo. One possibility is that binding sites are not equally accessible across the genome. A more comprehensive biophysical representation of TF-binding is required to improve our ability to understand, predict, and alter gene expression. Here, we show that genome accessibility is a key parameter that impacts TF-binding in bacteria. We developed a thermodynamic model that parameterizes ChIP-seq coverage in terms of genome accessibility and binding affinity. The role of genome accessibility is validated using a large-scale ChIP-seq dataset of the M. tuberculosis regulatory network. We find that accounting for genome accessibility led to a model that explains 63% of the ChIP-seq profile variance, while a model based in motif score alone explains only 35% of the variance. Moreover, our framework enables de novo ChIP-seq peak prediction and is useful for inferring TF-binding peaks in new experimental conditions by reducing the need for additional experiments. We observe that the genome is more accessible in intergenic regions, and that increased accessibility is positively correlated with gene expression and anti-correlated with distance to the origin of replication. Our biophysically motivated model provides a more comprehensive description of TF-binding in vivo from first principles towards a better representation of gene regulation in silico, with promising applications in systems biology.

  20. A genomic approach to identify regulatory nodes in the transcriptional network of systemic acquired resistance in plants.

    Directory of Open Access Journals (Sweden)

    Dong Wang

    2006-11-01

    Full Text Available Many biological processes are controlled by intricate networks of transcriptional regulators. With the development of microarray technology, transcriptional changes can be examined at the whole-genome level. However, such analysis often lacks information on the hierarchical relationship between components of a given system. Systemic acquired resistance (SAR is an inducible plant defense response involving a cascade of transcriptional events induced by salicylic acid through the transcription cofactor NPR1. To identify additional regulatory nodes in the SAR network, we performed microarray analysis on Arabidopsis plants expressing the NPR1-GR (glucocorticoid receptor fusion protein. Since nuclear translocation of NPR1-GR requires dexamethasone, we were able to control NPR1-dependent transcription and identify direct transcriptional targets of NPR1. We show that NPR1 directly upregulates the expression of eight WRKY transcription factor genes. This large family of 74 transcription factors has been implicated in various defense responses, but no specific WRKY factor has been placed in the SAR network. Identification of NPR1-regulated WRKY factors allowed us to perform in-depth genetic analysis on a small number of WRKY factors and test well-defined phenotypes of single and double mutants associated with NPR1. Among these WRKY factors we found both positive and negative regulators of SAR. This genomics-directed approach unambiguously positioned five WRKY factors in the complex transcriptional regulatory network of SAR. Our work not only discovered new transcription regulatory components in the signaling network of SAR but also demonstrated that functional studies of large gene families have to take into consideration sequence similarity as well as the expression patterns of the candidates.

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

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    Christopher A Lavender

    2016-08-01

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

  2. Aberrant methylation and associated transcriptional mobilization of Alu elements contributes to genomic instability in hypoxia.

    Science.gov (United States)

    Pal, Arnab; Srivastava, Tapasya; Sharma, Manish K; Mehndiratta, Mohit; Das, Prerna; Sinha, Subrata; Chattopadhyay, Parthaprasad

    2010-11-01

    Hypoxia is an integral part of tumorigenesis and contributes extensively to the neoplastic phenotype including drug resistance and genomic instability. It has also been reported that hypoxia results in global demethylation. Because a majority of the cytosine-phosphate-guanine (CpG) islands are found within the repeat elements of DNA, and are usually methylated under normoxic conditions, we suggested that retrotransposable Alu or short interspersed nuclear elements (SINEs) which show altered methylation and associated changes of gene expression during hypoxia, could be associated with genomic instability. U87MG glioblastoma cells were cultured in 0.1% O₂ for 6 weeks and compared with cells cultured in 21% O₂ for the same duration. Real-time PCR analysis showed a significant increase in SINE and reverse transcriptase coding long interspersed nuclear element (LINE) transcripts during hypoxia. Sequencing of bisulphite treated DNA as well as the Combined Bisulfite Restriction Analysis (COBRA) assay showed that the SINE loci studied underwent significant hypomethylation though there was patchy hypermethylation at a few sites. The inter-alu PCR profile of DNA from cells cultured under 6-week hypoxia, its 4-week revert back to normoxia and 6-week normoxia showed several changes in the band pattern indicating increased alu mediated genomic alteration. Our results show that aberrant methylation leading to increased transcription of SINE and reverse transcriptase associated LINE elements could lead to increased genomic instability in hypoxia. This might be a cause of genetic heterogeneity in tumours especially in variegated hypoxic environment and lead to a development of foci of more aggressive tumour cells. © 2009 The Authors Journal compilation © 2010 Foundation for Cellular and Molecular Medicine/Blackwell Publishing Ltd.

  3. Does selection against transcriptional interference shape retroelement-free regions in mammalian genomes?

    DEFF Research Database (Denmark)

    Mourier, Tobias; Willerslev, Eske

    2008-01-01

    in generating and maintaining retroelement-free regions in the human genome. METHODOLOGY/PRINCIPAL FINDINGS: Based on the known transcriptional properties of retroelements, we expect long interspersed elements (LINEs) to be able to display a high degree of transcriptional interference. In contrast, we expect......BACKGROUND: Eukaryotic genomes are scattered with retroelements that proliferate through retrotransposition. Although retroelements make up around 40 percent of the human genome, large regions are found to be completely devoid of retroelements. This has been hypothesised to be a result of genomic...... activity of LINEs has been identified previously. CONCLUSIONS/SIGNIFICANCE: Our observations are consistent with the notion that selection against transcriptional interference has contributed to the maintenance and/or generation of retroelement-free regions in the human genome....

  4. Transcriptional profiling in response to terminal drought stress reveals differential responses along the wheat genome

    Directory of Open Access Journals (Sweden)

    Ferrari Francesco

    2009-06-01

    Full Text Available Abstract Background Water stress during grain filling has a marked effect on grain yield, leading to a reduced endosperm cell number and thus sink capacity to accumulate dry matter. The bread wheat cultivar Chinese Spring (CS, a Chinese Spring terminal deletion line (CS_5AL-10 and the durum wheat cultivar Creso were subjected to transcriptional profiling after exposure to mild and severe drought stress at the grain filling stage to find evidences of differential stress responses associated to different wheat genome regions. Results The transcriptome analysis of Creso, CS and its deletion line revealed 8,552 non redundant probe sets with different expression levels, mainly due to the comparisons between the two species. The drought treatments modified the expression of 3,056 probe sets. Besides a set of genes showing a similar drought response in Creso and CS, cluster analysis revealed several drought response features that can be associated to the different genomic structure of Creso, CS and CS_5AL-10. Some drought-related genes were expressed at lower level (or not expressed in Creso (which lacks the D genome or in the CS_5AL-10 deletion line compared to CS. The chromosome location of a set of these genes was confirmed by PCR-based mapping on the D genome (or the 5AL-10 region. Many clusters were characterized by different level of expression in Creso, CS and CS_AL-10, suggesting that the different genome organization of the three genotypes may affect plant adaptation to stress. Clusters with similar expression trend were grouped and functional classified to mine the biological mean of their activation or repression. Genes involved in ABA, proline, glycine-betaine and sorbitol pathways were found up-regulated by drought stress. Furthermore, the enhanced expression of a set of transposons and retrotransposons was detected in CS_5AL-10. Conclusion Bread and durum wheat genotypes were characterized by a different physiological reaction to water

  5. eRNAs promote transcription by establishing chromatin accessibility at defined genomic loci

    DEFF Research Database (Denmark)

    Mousavi, Kambiz; Zare, Hossein; Dell'orso, Stefania

    2013-01-01

    )RNA acted to activate the downstream myogenic genes. The deployment of transcriptional machinery to appropriate loci is contingent on chromatin accessibility, a rate-limiting step preceding Pol II assembly. By nuclease sensitivity assay, we found that eRNAs regulate genomic access of the transcriptional...... complex to defined regulatory regions. In conclusion, our data suggest that eRNAs contribute to establishing a cell-type-specific transcriptional circuitry by directing chromatin-remodeling events....

  6. Systematic analysis of transcription start sites in avian development.

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    Marina Lizio

    2017-09-01

    Full Text Available Cap Analysis of Gene Expression (CAGE in combination with single-molecule sequencing technology allows precision mapping of transcription start sites (TSSs and genome-wide capture of promoter activities in differentiated and steady state cell populations. Much less is known about whether TSS profiling can characterize diverse and non-steady state cell populations, such as the approximately 400 transitory and heterogeneous cell types that arise during ontogeny of vertebrate animals. To gain such insight, we used the chick model and performed CAGE-based TSS analysis on embryonic samples covering the full 3-week developmental period. In total, 31,863 robust TSS peaks (>1 tag per million [TPM] were mapped to the latest chicken genome assembly, of which 34% to 46% were active in any given developmental stage. ZENBU, a web-based, open-source platform, was used for interactive data exploration. TSSs of genes critical for lineage differentiation could be precisely mapped and their activities tracked throughout development, suggesting that non-steady state and heterogeneous cell populations are amenable to CAGE-based transcriptional analysis. Our study also uncovered a large set of extremely stable housekeeping TSSs and many novel stage-specific ones. We furthermore demonstrated that TSS mapping could expedite motif-based promoter analysis for regulatory modules associated with stage-specific and housekeeping genes. Finally, using Brachyury as an example, we provide evidence that precise TSS mapping in combination with Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR-on technology enables us, for the first time, to efficiently target endogenous avian genes for transcriptional activation. Taken together, our results represent the first report of genome-wide TSS mapping in birds and the first systematic developmental TSS analysis in any amniote species (birds and mammals. By facilitating promoter-based molecular analysis and genetic

  7. From the Beauty of Genomic Landscapes to the Strength of Transcriptional Mechanisms.

    Science.gov (United States)

    Natoli, Gioacchino

    2016-03-24

    Genomic analyses are commonly used to infer trends and broad rules underlying transcriptional control. The innovative approach by Tong et al. to interrogate genomic datasets allows extracting mechanistic information on the specific regulation of individual genes. Copyright © 2016 Elsevier Inc. All rights reserved.

  8. Does selection against transcriptional interference shape retroelement-free regions in mammalian genomes?

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    Tobias Mourier

    Full Text Available BACKGROUND: Eukaryotic genomes are scattered with retroelements that proliferate through retrotransposition. Although retroelements make up around 40 percent of the human genome, large regions are found to be completely devoid of retroelements. This has been hypothesised to be a result of genomic regions being intolerant to insertions of retroelements. The inadvertent transcriptional activity of retroelements may affect neighbouring genes, which in turn could be detrimental to an organism. We speculate that such retroelement transcription, or transcriptional interference, is a contributing factor in generating and maintaining retroelement-free regions in the human genome. METHODOLOGY/PRINCIPAL FINDINGS: Based on the known transcriptional properties of retroelements, we expect long interspersed elements (LINEs to be able to display a high degree of transcriptional interference. In contrast, we expect short interspersed elements (SINEs to display very low levels of transcriptional interference. We find that genomic regions devoid of long interspersed elements (LINEs are enriched for protein-coding genes, but that this is not the case for regions devoid of short interspersed elements (SINEs. This is expected if genes are subject to selection against transcriptional interference. We do not find microRNAs to be associated with genomic regions devoid of either SINEs or LINEs. We further observe an increased relative activity of genes overlapping LINE-free regions during early embryogenesis, where activity of LINEs has been identified previously. CONCLUSIONS/SIGNIFICANCE: Our observations are consistent with the notion that selection against transcriptional interference has contributed to the maintenance and/or generation of retroelement-free regions in the human genome.

  9. Novel transcripts discovered by mining genomic DNA from defined regions of bovine chromosome 6

    Directory of Open Access Journals (Sweden)

    Eberlein Annett

    2009-04-01

    Full Text Available Abstract Background Linkage analyses strongly suggest a number of QTL for production, health and conformation traits in the middle part of bovine chromosome 6 (BTA6. The identification of the molecular background underlying the genetic variation at the QTL and subsequent functional studies require a well-annotated gene sequence map of the critical QTL intervals. To complete the sequence map of the defined subchromosomal regions on BTA6 poorly covered with comparative gene information, we focused on targeted isolation of transcribed sequences from bovine bacterial artificial chromosome (BAC clones mapped to the QTL intervals. Results Using the method of exon trapping, 92 unique exon trapping sequences (ETS were discovered in a chromosomal region of poor gene coverage. Sequence identity to the current NCBI sequence assembly for BTA6 was detected for 91% of unique ETS. Comparative sequence similarity search revealed that 11% of the isolated ETS displayed high similarity to genomic sequences located on the syntenic chromosomes of the human and mouse reference genome assemblies. Nearly a third of the ETS identified similar equivalent sequences in genomic sequence scaffolds from the alternative Celera-based sequence assembly of the human genome. Screening gene, EST, and protein databases detected 17% of ETS with identity to known transcribed sequences. Expression analysis of a subset of the ETS showed that most ETS (84% displayed a distinctive expression pattern in a multi-tissue panel of a lactating cow verifying their existence in the bovine transcriptome. Conclusion The results of our study demonstrate that the exon trapping method based on region-specific BAC clones is very useful for targeted screening for novel transcripts located within a defined chromosomal region being deficiently endowed with annotated gene information. The majority of identified ETS represents unknown noncoding sequences in intergenic regions on BTA6 displaying a

  10. Comparative analysis of the full genome sequence of European bat lyssavirus type 1 and type 2 with other lyssaviruses and evidence for a conserved transcription termination and polyadenylation motif in the G-L 3' non-translated region.

    Science.gov (United States)

    Marston, D A; McElhinney, L M; Johnson, N; Müller, T; Conzelmann, K K; Tordo, N; Fooks, A R

    2007-04-01

    We report the first full-length genomic sequences for European bat lyssavirus type-1 (EBLV-1) and type-2 (EBLV-2). The EBLV-1 genomic sequence was derived from a virus isolated from a serotine bat in Hamburg, Germany, in 1968 and the EBLV-2 sequence was derived from a virus isolate from a human case of rabies that occurred in Scotland in 2002. A long-distance PCR strategy was used to amplify the open reading frames (ORFs), followed by standard and modified RACE (rapid amplification of cDNA ends) techniques to amplify the 3' and 5' ends. The lengths of each complete viral genome for EBLV-1 and EBLV-2 were 11 966 and 11 930 base pairs, respectively, and follow the standard rhabdovirus genome organization of five viral proteins. Comparison with other lyssavirus sequences demonstrates variation in degrees of homology, with the genomic termini showing a high degree of complementarity. The nucleoprotein was the most conserved, both intra- and intergenotypically, followed by the polymerase (L), matrix and glyco- proteins, with the phosphoprotein being the most variable. In addition, we have shown that the two EBLVs utilize a conserved transcription termination and polyadenylation (TTP) motif, approximately 50 nt upstream of the L gene start codon. All available lyssavirus sequences to date, with the exception of Pasteur virus (PV) and PV-derived isolates, use the second TTP site. This observation may explain differences in pathogenicity between lyssavirus strains, dependent on the length of the untranslated region, which might affect transcriptional activity and RNA stability.

  11. Hierarchical role for transcription factors and chromatin structure in genome organization along adipogenesis

    DEFF Research Database (Denmark)

    Sarusi Portuguez, Avital; Schwartz, Michal; Siersbaek, Rasmus

    2017-01-01

    The three dimensional folding of mammalian genomes is cell type specific and difficult to alter suggesting that it is an important component of gene regulation. However, given the multitude of chromatin-associating factors, the mechanisms driving the colocalization of active chromosomal domains...... by PPARγ and Lpin1, undergoes orchestrated reorganization during adipogenesis. Coupling the dynamics of genome architecture with multiple chromatin datasets indicated that among all the transcription factors (TFs) tested, RXR is central to genome reorganization at the beginning of adipogenesis...

  12. Genome-wide in silico identification of GPI proteins in Mycosphaerella fijiensis and transcriptional analysis of two GPI-anchored β-1,3-glucanosyltransferases.

    Science.gov (United States)

    Kantún-Moreno, Nuvia; Vázquez-Euán, Roberto; Tzec-Simá, Miguel; Peraza-Echeverría, Leticia; Grijalva-Arango, Rosa; Rodríguez-García, Cecilia; James, Andrew C; Ramírez-Prado, Jorge; Islas-Flores, Ignacio; Canto-Canché, Blondy

    2013-01-01

    The hemibiotrophic fungus Mycosphaerella fijiensis is the causal agent of black Sigatoka (BS), the most devastating foliar disease in banana (Musa spp.) worldwide. Little is known about genes that are important during M. fijiensis-Musa sp. interaction. The fungal cell wall is an attractive area of study because it is essential for maintenance of cellular homeostasis and it is the most external structure in the fungal cell and therefore mediates the interaction of the pathogen with the host. In this manuscript we describe the in silico identification of glycosyl phosphatidylinositol-protein (GPI) family in M. fijiensis, and the analysis of two β-1,3-glucanosyltrans-ferases (Gas), selected by homology with fungal pathogenicity factors. Potential roles in pathogenesis were evaluated through analyzing expression during different stages of black Sigatoka disease, comparing expression data with BS symptoms and fungal biomass inside leaves. Real-time quantitative RT-PCR showed nearly constant expression of MfGAS1 with slightly increases (about threefold) in conidia and at speck-necrotrophic stage during banana-pathogen interaction. Conversely, MfGAS2 expression was increased during biotrophy (about seven times) and reached a maximum at speck (about 23 times) followed by a progressive decrease in next stages, suggesting an active role in M. fijiensis pathogenesis.

  13. Distinct Contributions of Replication and Transcription to Mutation Rate Variation of Human Genomes

    KAUST Repository

    Cui, Peng; Ding, Feng; Lin, Qiang; Zhang, Lingfang; Li, Ang; Zhang, Zhang; Hu, Songnian; Yu, Jun

    2012-01-01

    Here, we evaluate the contribution of two major biological processes—DNA replication and transcription—to mutation rate variation in human genomes. Based on analysis of the public human tissue transcriptomics data, high-resolution replicating map of Hela cells and dbSNP data, we present significant correlations between expression breadth, replication time in local regions and SNP density. SNP density of tissue-specific (TS) genes is significantly higher than that of housekeeping (HK) genes. TS genes tend to locate in late-replicating genomic regions and genes in such regions have a higher SNP density compared to those in early-replication regions. In addition, SNP density is found to be positively correlated with expression level among HK genes. We conclude that the process of DNA replication generates stronger mutational pressure than transcription-associated biological processes do, resulting in an increase of mutation rate in TS genes while having weaker effects on HK genes. In contrast, transcription-associated processes are mainly responsible for the accumulation of mutations in highly-expressed HK genes.

  14. Distinct Contributions of Replication and Transcription to Mutation Rate Variation of Human Genomes

    KAUST Repository

    Cui, Peng

    2012-03-23

    Here, we evaluate the contribution of two major biological processes—DNA replication and transcription—to mutation rate variation in human genomes. Based on analysis of the public human tissue transcriptomics data, high-resolution replicating map of Hela cells and dbSNP data, we present significant correlations between expression breadth, replication time in local regions and SNP density. SNP density of tissue-specific (TS) genes is significantly higher than that of housekeeping (HK) genes. TS genes tend to locate in late-replicating genomic regions and genes in such regions have a higher SNP density compared to those in early-replication regions. In addition, SNP density is found to be positively correlated with expression level among HK genes. We conclude that the process of DNA replication generates stronger mutational pressure than transcription-associated biological processes do, resulting in an increase of mutation rate in TS genes while having weaker effects on HK genes. In contrast, transcription-associated processes are mainly responsible for the accumulation of mutations in highly-expressed HK genes.

  15. A code for transcription initiation in mammalian genomes

    DEFF Research Database (Denmark)

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

    2007-01-01

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

  16. Integrated analysis of whole genome and transcriptome sequencing reveals diverse transcriptomic aberrations driven by somatic genomic changes in liver cancers.

    Directory of Open Access Journals (Sweden)

    Yuichi Shiraishi

    Full Text Available Recent studies applying high-throughput sequencing technologies have identified several recurrently mutated genes and pathways in multiple cancer genomes. However, transcriptional consequences from these genomic alterations in cancer genome remain unclear. In this study, we performed integrated and comparative analyses of whole genomes and transcriptomes of 22 hepatitis B virus (HBV-related hepatocellular carcinomas (HCCs and their matched controls. Comparison of whole genome sequence (WGS and RNA-Seq revealed much evidence that various types of genomic mutations triggered diverse transcriptional changes. Not only splice-site mutations, but also silent mutations in coding regions, deep intronic mutations and structural changes caused splicing aberrations. HBV integrations generated diverse patterns of virus-human fusion transcripts depending on affected gene, such as TERT, CDK15, FN1 and MLL4. Structural variations could drive over-expression of genes such as WNT ligands, with/without creating gene fusions. Furthermore, by taking account of genomic mutations causing transcriptional aberrations, we could improve the sensitivity of deleterious mutation detection in known cancer driver genes (TP53, AXIN1, ARID2, RPS6KA3, and identified recurrent disruptions in putative cancer driver genes such as HNF4A, CPS1, TSC1 and THRAP3 in HCCs. These findings indicate genomic alterations in cancer genome have diverse transcriptomic effects, and integrated analysis of WGS and RNA-Seq can facilitate the interpretation of a large number of genomic alterations detected in cancer genome.

  17. Improved methods and resources for paramecium genomics: transcription units, gene annotation and gene expression.

    Science.gov (United States)

    Arnaiz, Olivier; Van Dijk, Erwin; Bétermier, Mireille; Lhuillier-Akakpo, Maoussi; de Vanssay, Augustin; Duharcourt, Sandra; Sallet, Erika; Gouzy, Jérôme; Sperling, Linda

    2017-06-26

    The 15 sibling species of the Paramecium aurelia cryptic species complex emerged after a whole genome duplication that occurred tens of millions of years ago. Given extensive knowledge of the genetics and epigenetics of Paramecium acquired over the last century, this species complex offers a uniquely powerful system to investigate the consequences of whole genome duplication in a unicellular eukaryote as well as the genetic and epigenetic mechanisms that drive speciation. High quality Paramecium gene models are important for research using this system. The major aim of the work reported here was to build an improved gene annotation pipeline for the Paramecium lineage. We generated oriented RNA-Seq transcriptome data across the sexual process of autogamy for the model species Paramecium tetraurelia. We determined, for the first time in a ciliate, candidate P. tetraurelia transcription start sites using an adapted Cap-Seq protocol. We developed TrUC, multi-threaded Perl software that in conjunction with TopHat mapping of RNA-Seq data to a reference genome, predicts transcription units for the annotation pipeline. We used EuGene software to combine annotation evidence. The high quality gene structural annotations obtained for P. tetraurelia were used as evidence to improve published annotations for 3 other Paramecium species. The RNA-Seq data were also used for differential gene expression analysis, providing a gene expression atlas that is more sensitive than the previously established microarray resource. We have developed a gene annotation pipeline tailored for the compact genomes and tiny introns of Paramecium species. A novel component of this pipeline, TrUC, predicts transcription units using Cap-Seq and oriented RNA-Seq data. TrUC could prove useful beyond Paramecium, especially in the case of high gene density. Accurate predictions of 3' and 5' UTR will be particularly valuable for studies of gene expression (e.g. nucleosome positioning, identification of cis

  18. Genome-wide identification of soybean WRKY transcription factors in response to salt stress.

    Science.gov (United States)

    Yu, Yanchong; Wang, Nan; Hu, Ruibo; Xiang, Fengning

    2016-01-01

    Members of the large family of WRKY transcription factors are involved in a wide range of developmental and physiological processes, most particularly in the plant response to biotic and abiotic stress. Here, an analysis of the soybean genome sequence allowed the identification of the full complement of 188 soybean WRKY genes. Phylogenetic analysis revealed that soybean WRKY genes were classified into three major groups (I, II, III), with the second group further categorized into five subgroups (IIa-IIe). The soybean WRKYs from each group shared similar gene structures and motif compositions. The location of the GmWRKYs was dispersed over all 20 soybean chromosomes. The whole genome duplication appeared to have contributed significantly to the expansion of the family. Expression analysis by RNA-seq indicated that in soybean root, 66 of the genes responded rapidly and transiently to the imposition of salt stress, all but one being up-regulated. While in aerial part, 49 GmWRKYs responded, all but two being down-regulated. RT-qPCR analysis showed that in the whole soybean plant, 66 GmWRKYs exhibited distinct expression patterns in response to salt stress, of which 12 showed no significant change, 35 were decreased, while 19 were induced. The data present here provide critical clues for further functional studies of WRKY gene in soybean salt tolerance.

  19. Dynamic analysis of stochastic transcription cycles.

    Directory of Open Access Journals (Sweden)

    Claire V Harper

    2011-04-01

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

  20. Transcription facilitated genome-wide recruitment of topoisomerase I and DNA gyrase.

    Science.gov (United States)

    Ahmed, Wareed; Sala, Claudia; Hegde, Shubhada R; Jha, Rajiv Kumar; Cole, Stewart T; Nagaraja, Valakunja

    2017-05-01

    Movement of the transcription machinery along a template alters DNA topology resulting in the accumulation of supercoils in DNA. The positive supercoils generated ahead of transcribing RNA polymerase (RNAP) and the negative supercoils accumulating behind impose severe topological constraints impeding transcription process. Previous studies have implied the role of topoisomerases in the removal of torsional stress and the maintenance of template topology but the in vivo interaction of functionally distinct topoisomerases with heterogeneous chromosomal territories is not deciphered. Moreover, how the transcription-induced supercoils influence the genome-wide recruitment of DNA topoisomerases remains to be explored in bacteria. Using ChIP-Seq, we show the genome-wide occupancy profile of both topoisomerase I and DNA gyrase in conjunction with RNAP in Mycobacterium tuberculosis taking advantage of minimal topoisomerase representation in the organism. The study unveils the first in vivo genome-wide interaction of both the topoisomerases with the genomic regions and establishes that transcription-induced supercoils govern their recruitment at genomic sites. Distribution profiles revealed co-localization of RNAP and the two topoisomerases on the active transcriptional units (TUs). At a given locus, topoisomerase I and DNA gyrase were localized behind and ahead of RNAP, respectively, correlating with the twin-supercoiled domains generated. The recruitment of topoisomerases was higher at the genomic loci with higher transcriptional activity and/or at regions under high torsional stress compared to silent genomic loci. Importantly, the occupancy of DNA gyrase, sole type II topoisomerase in Mtb, near the Ter domain of the Mtb chromosome validates its function as a decatenase.

  1. Genome Mutational and Transcriptional Hotspots Are Traps for Duplicated Genes and Sources of Adaptations.

    Science.gov (United States)

    Fares, Mario A; Sabater-Muñoz, Beatriz; Toft, Christina

    2017-05-01

    Gene duplication generates new genetic material, which has been shown to lead to major innovations in unicellular and multicellular organisms. A whole-genome duplication occurred in the ancestor of Saccharomyces yeast species but 92% of duplicates returned to single-copy genes shortly after duplication. The persisting duplicated genes in Saccharomyces led to the origin of major metabolic innovations, which have been the source of the unique biotechnological capabilities in the Baker's yeast Saccharomyces cerevisiae. What factors have determined the fate of duplicated genes remains unknown. Here, we report the first demonstration that the local genome mutation and transcription rates determine the fate of duplicates. We show, for the first time, a preferential location of duplicated genes in the mutational and transcriptional hotspots of S. cerevisiae genome. The mechanism of duplication matters, with whole-genome duplicates exhibiting different preservation trends compared to small-scale duplicates. Genome mutational and transcriptional hotspots are rich in duplicates with large repetitive promoter elements. Saccharomyces cerevisiae shows more tolerance to deleterious mutations in duplicates with repetitive promoter elements, which in turn exhibit higher transcriptional plasticity against environmental perturbations. Our data demonstrate that the genome traps duplicates through the accelerated regulatory and functional divergence of their gene copies providing a source of novel adaptations in yeast. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  2. Genomic survey of bZIP transcription factor genes related to tanshinone biosynthesis in Salvia miltiorrhiza

    Directory of Open Access Journals (Sweden)

    Yu Zhang

    2018-03-01

    Full Text Available Tanshinones are a class of bioactive components in the traditional Chinese medicine Salvia miltiorrhiza, and their biosynthesis and regulation have been widely studied. Current studies show that basic leucine zipper (bZIP proteins regulate plant secondary metabolism, growth and developmental processes. However, the bZIP transcription factors involved in tanshinone biosynthesis are unknown. Here, we conducted the first genome-wide survey of the bZIP gene family and analyzed the phylogeny, gene structure, additional conserved motifs and alternative splicing events in S. miltiorrhiza. A total of 70 SmbZIP transcription factors were identified and categorized into 11 subgroups based on their phylogenetic relationships with those in Arabidopsis. Moreover, seventeen SmbZIP genes underwent alternative splicing events. According to the transcriptomic data, the SmbZIP genes that were highly expressed in the Danshen root and periderm were selected. Based on the prediction of bZIP binding sites in the promoters and the co-expression analysis and co-induction patterns in response to Ag+ treatment via quantitative real-time polymerase chain reaction (qRT-PCR, we concluded that SmbZIP7 and SmbZIP20 potentially participate in the regulation of tanshinone biosynthesis. These results provide a foundation for further functional characterization of the candidate SmbZIP genes, which have the potential to increase tanshinone production. KEY WORDS: bZIP genes, Salvia miltiorrhiza, Phylogenetic analysis, Expression pattern analysis, Tanshinone biosynthesis

  3. Targeted genome regulation via synthetic programmable transcriptional regulators

    KAUST Repository

    Piatek, Agnieszka Anna; Mahfouz, Magdy M.

    2016-01-01

    genes in linear and interacting pathways in a native context. Modular DNA-binding domains from zinc fingers (ZFs) and transcriptional activator-like proteins (TALE) are amenable to bioengineering to bind DNA target sequences of interest. As a result, ZF

  4. Comparative Genome Analysis and Genome Evolution

    NARCIS (Netherlands)

    Snel, Berend

    2002-01-01

    This thesis described a collection of bioinformatic analyses on complete genome sequence data. We have studied the evolution of gene content and find that vertical inheritance dominates over horizontal gene trasnfer, even to the extent that we can use the gene content to make genome phylogenies.

  5. A pilot study of transcription unit analysis in rice using oligonucleotide tiling-path microarray

    DEFF Research Database (Denmark)

    Stolc, Viktor; Li, Lei; Wang, Xiangfeng

    2005-01-01

    As the international efforts to sequence the rice genome are completed, an immediate challenge and opportunity is to comprehensively and accurately define all transcription units in the rice genome. Here we describe a strategy of using high-density oligonucleotide tiling-path microarrays to map...... transcription of the japonica rice genome. In a pilot experiment to test this approach, one array representing the reverse strand of the last 11.2 Mb sequence of chromosome 10 was analyzed in detail based on a mathematical model developed in this study. Analysis of the array data detected 77% of the reference...... gene models in a mixture of four RNA populations. Moreover, significant transcriptional activities were found in many of the previously annotated intergenic regions. These preliminary results demonstrate the utility of genome tiling microarrays in evaluating annotated rice gene models...

  6. DOT1L and H3K79 Methylation in Transcription and Genomic Stability.

    Science.gov (United States)

    Wood, Katherine; Tellier, Michael; Murphy, Shona

    2018-02-27

    The organization of eukaryotic genomes into chromatin provides challenges for the cell to accomplish basic cellular functions, such as transcription, DNA replication and repair of DNA damage. Accordingly, a range of proteins modify and/or read chromatin states to regulate access to chromosomal DNA. Yeast Dot1 and the mammalian homologue DOT1L are methyltransferases that can add up to three methyl groups to histone H3 lysine 79 (H3K79). H3K79 methylation is implicated in several processes, including transcription elongation by RNA polymerase II, the DNA damage response and cell cycle checkpoint activation. DOT1L is also an important drug target for treatment of mixed lineage leukemia (MLL)-rearranged leukemia where aberrant transcriptional activation is promoted by DOT1L mislocalisation. This review summarizes what is currently known about the role of Dot1/DOT1L and H3K79 methylation in transcription and genomic stability.

  7. DOT1L and H3K79 Methylation in Transcription and Genomic Stability

    Directory of Open Access Journals (Sweden)

    Katherine Wood

    2018-02-01

    Full Text Available The organization of eukaryotic genomes into chromatin provides challenges for the cell to accomplish basic cellular functions, such as transcription, DNA replication and repair of DNA damage. Accordingly, a range of proteins modify and/or read chromatin states to regulate access to chromosomal DNA. Yeast Dot1 and the mammalian homologue DOT1L are methyltransferases that can add up to three methyl groups to histone H3 lysine 79 (H3K79. H3K79 methylation is implicated in several processes, including transcription elongation by RNA polymerase II, the DNA damage response and cell cycle checkpoint activation. DOT1L is also an important drug target for treatment of mixed lineage leukemia (MLL-rearranged leukemia where aberrant transcriptional activation is promoted by DOT1L mislocalisation. This review summarizes what is currently known about the role of Dot1/DOT1L and H3K79 methylation in transcription and genomic stability.

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

    Directory of Open Access Journals (Sweden)

    Qianli Huang

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

  9. Genome-Wide Chromosomal Targets of Oncogenic Transcription Factors

    Science.gov (United States)

    2008-04-01

    Altman, W.E., Attiya, S., Bader, J.S., Bemben, L.A., Berka , J., Braverman, M.S., Chen, Y.J., Chen, Z., et al. 2005. Genome sequencing in microfabricated...software after filtering to exclude bad spots. qPCR validation. Primer pairs used in Figure 1 were designed to cover three peaks and three troughs in

  10. A hyperactive transcriptional state marks genome reactivation at the mitosis–G1 transition

    Science.gov (United States)

    Hsiung, Chris C.-S.; Bartman, Caroline R.; Huang, Peng; Ginart, Paul; Stonestrom, Aaron J.; Keller, Cheryl A.; Face, Carolyne; Jahn, Kristen S.; Evans, Perry; Sankaranarayanan, Laavanya; Giardine, Belinda; Hardison, Ross C.; Raj, Arjun; Blobel, Gerd A.

    2016-01-01

    During mitosis, RNA polymerase II (Pol II) and many transcription factors dissociate from chromatin, and transcription ceases globally. Transcription is known to restart in bulk by telophase, but whether de novo transcription at the mitosis–G1 transition is in any way distinct from later in interphase remains unknown. We tracked Pol II occupancy genome-wide in mammalian cells progressing from mitosis through late G1. Unexpectedly, during the earliest rounds of transcription at the mitosis–G1 transition, ∼50% of active genes and distal enhancers exhibit a spike in transcription, exceeding levels observed later in G1 phase. Enhancer–promoter chromatin contacts are depleted during mitosis and restored rapidly upon G1 entry but do not spike. Of the chromatin-associated features examined, histone H3 Lys27 acetylation levels at individual loci in mitosis best predict the mitosis–G1 transcriptional spike. Single-molecule RNA imaging supports that the mitosis–G1 transcriptional spike can constitute the maximum transcriptional activity per DNA copy throughout the cell division cycle. The transcriptional spike occurs heterogeneously and propagates to cell-to-cell differences in mature mRNA expression. Our results raise the possibility that passage through the mitosis–G1 transition might predispose cells to diverge in gene expression states. PMID:27340175

  11. A hyperactive transcriptional state marks genome reactivation at the mitosis-G1 transition.

    Science.gov (United States)

    Hsiung, Chris C-S; Bartman, Caroline R; Huang, Peng; Ginart, Paul; Stonestrom, Aaron J; Keller, Cheryl A; Face, Carolyne; Jahn, Kristen S; Evans, Perry; Sankaranarayanan, Laavanya; Giardine, Belinda; Hardison, Ross C; Raj, Arjun; Blobel, Gerd A

    2016-06-15

    During mitosis, RNA polymerase II (Pol II) and many transcription factors dissociate from chromatin, and transcription ceases globally. Transcription is known to restart in bulk by telophase, but whether de novo transcription at the mitosis-G1 transition is in any way distinct from later in interphase remains unknown. We tracked Pol II occupancy genome-wide in mammalian cells progressing from mitosis through late G1. Unexpectedly, during the earliest rounds of transcription at the mitosis-G1 transition, ∼50% of active genes and distal enhancers exhibit a spike in transcription, exceeding levels observed later in G1 phase. Enhancer-promoter chromatin contacts are depleted during mitosis and restored rapidly upon G1 entry but do not spike. Of the chromatin-associated features examined, histone H3 Lys27 acetylation levels at individual loci in mitosis best predict the mitosis-G1 transcriptional spike. Single-molecule RNA imaging supports that the mitosis-G1 transcriptional spike can constitute the maximum transcriptional activity per DNA copy throughout the cell division cycle. The transcriptional spike occurs heterogeneously and propagates to cell-to-cell differences in mature mRNA expression. Our results raise the possibility that passage through the mitosis-G1 transition might predispose cells to diverge in gene expression states. © 2016 Hsiung et al.; Published by Cold Spring Harbor Laboratory Press.

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

    Directory of Open Access Journals (Sweden)

    Simona Cugusi

    2016-01-01

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

  13. Transcription-associated mutational pressure in the Parvovirus B19 genome: Reactivated genomes contribute to the variability of viral populations.

    Science.gov (United States)

    Khrustalev, Vladislav Victorovich; Ermalovich, Marina Anatolyevna; Hübschen, Judith M; Khrustaleva, Tatyana Aleksandrovna

    2017-12-21

    In this study we used non-overlapping parts of the two long open reading frames coding for nonstructural (NS) and capsid (VP) proteins of all available sequences of the Parvovirus B19 subgenotype 1a genome and found out that the rates of A to G, C to T and A to T mutations are higher in the first long reading frame (NS) of the virus than in the second one (VP). This difference in mutational pressure directions for two parts of the same viral genome can be explained by the fact of transcription of just the first long reading frame during the lifelong latency in nonerythroid cells. Adenine deamination (producing A to G and A to T mutations) and cytosine deamination (producing C to T mutations) occur more frequently in transcriptional bubbles formed by DNA "plus" strand of the first open reading frame. These mutations can be inherited only in case of reactivation of the infectious virus due to the help of Adenovirus that allows latent Parvovirus B19 to start transcription of the second reading frame and then to replicate its genome by the rolling circle mechanism using the specific origin. Results of this study provide evidence that the genomes reactivated from latency make significant contributions to the variability of Parvovirus B19. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. Gene discovery and transcript analyses in the corn smut pathogen Ustilago maydis: expressed sequence tag and genome sequence comparison

    Directory of Open Access Journals (Sweden)

    Saville Barry J

    2007-09-01

    Full Text Available Abstract Background Ustilago maydis is the basidiomycete fungus responsible for common smut of corn and is a model organism for the study of fungal phytopathogenesis. To aid in the annotation of the genome sequence of this organism, several expressed sequence tag (EST libraries were generated from a variety of U. maydis cell types. In addition to utility in the context of gene identification and structure annotation, the ESTs were analyzed to identify differentially abundant transcripts and to detect evidence of alternative splicing and anti-sense transcription. Results Four cDNA libraries were constructed using RNA isolated from U. maydis diploid teliospores (U. maydis strains 518 × 521 and haploid cells of strain 521 grown under nutrient rich, carbon starved, and nitrogen starved conditions. Using the genome sequence as a scaffold, the 15,901 ESTs were assembled into 6,101 contiguous expressed sequences (contigs; among these, 5,482 corresponded to predicted genes in the MUMDB (MIPS Ustilago maydis database, while 619 aligned to regions of the genome not yet designated as genes in MUMDB. A comparison of EST abundance identified numerous genes that may be regulated in a cell type or starvation-specific manner. The transcriptional response to nitrogen starvation was assessed using RT-qPCR. The results of this suggest that there may be cross-talk between the nitrogen and carbon signalling pathways in U. maydis. Bioinformatic analysis identified numerous examples of alternative splicing and anti-sense transcription. While intron retention was the predominant form of alternative splicing in U. maydis, other varieties were also evident (e.g. exon skipping. Selected instances of both alternative splicing and anti-sense transcription were independently confirmed using RT-PCR. Conclusion Through this work: 1 substantial sequence information has been provided for U. maydis genome annotation; 2 new genes were identified through the discovery of 619

  15. A Genome-Scale Resource for the Functional Characterization of Arabidopsis Transcription Factors

    Directory of Open Access Journals (Sweden)

    Jose L. Pruneda-Paz

    2014-07-01

    Full Text Available Extensive transcriptional networks play major roles in cellular and organismal functions. Transcript levels are in part determined by the combinatorial and overlapping functions of multiple transcription factors (TFs bound to gene promoters. Thus, TF-promoter interactions provide the basic molecular wiring of transcriptional regulatory networks. In plants, discovery of the functional roles of TFs is limited by an increased complexity of network circuitry due to a significant expansion of TF families. Here, we present the construction of a comprehensive collection of Arabidopsis TFs clones created to provide a versatile resource for uncovering TF biological functions. We leveraged this collection by implementing a high-throughput DNA binding assay and identified direct regulators of a key clock gene (CCA1 that provide molecular links between different signaling modules and the circadian clock. The resources introduced in this work will significantly contribute to a better understanding of the transcriptional regulatory landscape of plant genomes.

  16. Genomic and transcriptional landscape of P2RY8-CRLF2-positive childhood acute lymphoblastic leukemia

    Science.gov (United States)

    Vesely, C; Frech, C; Eckert, C; Cario, G; Mecklenbräuker, A; zur Stadt, U; Nebral, K; Kraler, F; Fischer, S; Attarbaschi, A; Schuster, M; Bock, C; Cavé, H; von Stackelberg, A; Schrappe, M; Horstmann, M A; Mann, G; Haas, O A; Panzer-Grümayer, R

    2017-01-01

    Children with P2RY8-CRLF2-positive acute lymphoblastic leukemia have an increased relapse risk. Their mutational and transcriptional landscape, as well as the respective patterns at relapse remain largely elusive. We, therefore, performed an integrated analysis of whole-exome and RNA sequencing in 41 major clone fusion-positive cases including 19 matched diagnosis/relapse pairs. We detected a variety of frequently subclonal and highly instable JAK/STAT but also RTK/Ras pathway-activating mutations in 76% of cases at diagnosis and virtually all relapses. Unlike P2RY8-CRLF2 that was lost in 32% of relapses, all other genomic alterations affecting lymphoid development (58%) and cell cycle (39%) remained stable. Only IKZF1 alterations predominated in relapsing cases (P=0.001) and increased from initially 36 to 58% in matched cases. IKZF1’s critical role is further corroborated by its specific transcriptional signature comprising stem cell features with signs of impaired lymphoid differentiation, enhanced focal adhesion, activated hypoxia pathway, deregulated cell cycle and increased drug resistance. Our findings support the notion that P2RY8-CRLF2 is dispensable for relapse development and instead highlight the prominent rank of IKZF1 for relapse development by mediating self-renewal and homing to the bone marrow niche. Consequently, reverting aberrant IKAROS signaling or its disparate programs emerges as an attractive potential treatment option in these leukemias. PMID:27899802

  17. Genome wide transcriptional response of Saccharomyces cerevisiae to stress-induced perturbations

    Directory of Open Access Journals (Sweden)

    Hilal eTaymaz-Nikerel

    2016-02-01

    Full Text Available Cells respond to environmental and/or genetic perturbations in order to survive and proliferate. Characterization of the changes after various stimuli at different -omics levels is crucial to comprehend the adaptation of cells to changing conditions. Genome wide quantification and analysis of transcript levels, the genes affected by perturbations, extends our understanding of cellular metabolism by pointing out the mechanisms that play role in sensing the stress caused by those perturbations and related signaling pathways, and in this way guides us to achieve endeavors such as rational engineering of cells or interpretation of disease mechanisms. Saccharomyces cerevisiae as a model system has been studied in response to different perturbations and corresponding transcriptional profiles were followed either statically or/and dynamically, short- and long- term. This review focuses on response of yeast cells to diverse stress inducing perturbations including nutritional changes, ionic stress, salt stress, oxidative stress, osmotic shock, as well as to genetic interventions such as deletion and over-expression of genes. It is aimed to conclude on common regulatory phenomena that allow yeast to organize its transcriptomic response after any perturbation under different external conditions.

  18. Transient Genome-Wide Transcriptional Response to Low-Dose Ionizing Radiation In Vivo in Humans

    International Nuclear Information System (INIS)

    Berglund, Susanne R.; Rocke, David M.; Dai Jian; Schwietert, Chad W.; Santana, Alison; Stern, Robin L.; Lehmann, Joerg; Hartmann Siantar, Christine L.; Goldberg, Zelanna

    2008-01-01

    Purpose: The in vivo effects of low-dose low linear energy transfer ionizing radiation on healthy human skin are largely unknown. Using a patient-based tissue acquisition protocol, we have performed a series of genomic analyses on the temporal dynamics over a 24-hour period to determine the radiation response after a single exposure of 10 cGy. Methods and Materials: RNA from each patient tissue sample was hybridized to an Affymetrix Human Genome U133 Plus 2.0 array. Data analysis was performed on selected gene groups and pathways. Results: Nineteen gene groups and seven gene pathways that had been shown to be radiation responsive were analyzed. Of these, nine gene groups showed significant transient transcriptional changes in the human tissue samples, which returned to baseline by 24 hours postexposure. Conclusions: Low doses of ionizing radiation on full-thickness human skin produce a definable temporal response out to 24 hours postexposure. Genes involved in DNA and tissue remodeling, cell cycle transition, and inflammation show statistically significant changes in expression, despite variability between patients. These data serve as a reference for the temporal dynamics of ionizing radiation response following low-dose exposure in healthy full-thickness human skin

  19. Analysis of phage Mu DNA transposition by whole-genome Escherichia coli tiling arrays reveals a complex relationship to distribution of target selection protein B, transcription and chromosome architectural elements.

    Science.gov (United States)

    Ge, Jun; Lou, Zheng; Cui, Hong; Shang, Lei; Harshey, Rasika M

    2011-09-01

    Of all known transposable elements, phage Mu exhibits the highest transposition efficiency and the lowest target specificity. In vitro, MuB protein is responsible for target choice. In this work, we provide a comprehensive assessment of the genome-wide distribution of MuB and its relationship to Mu target selection using high-resolution Escherichia coli tiling DNA arrays. We have also assessed how MuB binding and Mu transposition are influenced by chromosome-organizing elements such as AT-rich DNA signatures, or the binding of the nucleoid-associated protein Fis, or processes such as transcription. The results confirm and extend previous biochemical and lower resolution in vivo data. Despite the generally random nature of Mu transposition and MuB binding, there were hot and cold insertion sites and MuB binding sites in the genome, and differences between the hottest and coldest sites were large. The new data also suggest that MuB distribution and subsequent Mu integration is responsive to DNA sequences that contribute to the structural organization of the chromosome.

  20. CRISPR-Cas9-Mediated Genome Editing and Transcriptional Control in Yarrowia lipolytica.

    Science.gov (United States)

    Schwartz, Cory; Wheeldon, Ian

    2018-01-01

    The discovery and adaptation of RNA-guided nucleases has resulted in the rapid development of efficient, scalable, and easily accessible synthetic biology tools for targeted genome editing and transcriptional control. In these systems, for example CRISPR-Cas9 from Streptococcus pyogenes, a protein with nuclease activity is targeted to a specific nucleotide sequence by a short RNA molecule, whereupon binding it cleaves the targeted nucleotide strand. To extend this genome-editing ability to the industrially important oleaginous yeast Yarrowia lipolytica, we developed a set of easily usable and effective CRISPR-Cas9 episomal vectors. In this protocols chapter, we first present a method by which arbitrary protein-coding genes can be disrupted via indel formation after CRISPR-Cas9 targeting. A second method demonstrates how the same CRISPR-Cas9 system can be used to induce markerless gene cassette integration into the genome by inducing homologous recombination after DNA cleavage by Cas9. Finally, we describe how a catalytically inactive form of Cas9 fused to a transcriptional repressor can be used to control transcription of native genes in Y. lipolytica. The CRISPR-Cas9 tools and strategies described here greatly increase the types of genome editing and transcriptional control that can be achieved in Y. lipolytica, and promise to facilitate more advanced engineering of this important oleaginous host.

  1. Mapping 3 ' transcript ends in the bank vole (Clethrionomys glareolus) mitochondrial genome with RNA-Seq

    Czech Academy of Sciences Publication Activity Database

    Marková, Silvia; Filipi, Karolína; Searle, J. B.; Kotlík, Petr

    2015-01-01

    Roč. 16, č. 870 (2015) ISSN 1471-2164 R&D Projects: GA ČR GAP506/11/1872 Institutional support: RVO:67985904 Keywords : bicistronic transcript * mitochondrial genome * Myodes glareolus * transcriptome * polyadenylation * stop codon Subject RIV: EG - Zoology Impact factor: 3.867, year: 2015

  2. Transcription Restores DNA Repair to Heterochromatin, Determining Regional Mutation Rates in Cancer Genomes

    Directory of Open Access Journals (Sweden)

    Christina L. Zheng

    2014-11-01

    Full Text Available Somatic mutations in cancer are more frequent in heterochromatic and late-replicating regions of the genome. We report that regional disparities in mutation density are virtually abolished within transcriptionally silent genomic regions of cutaneous squamous cell carcinomas (cSCCs arising in an XPC−/− background. XPC−/− cells lack global genome nucleotide excision repair (GG-NER, thus establishing differential access of DNA repair machinery within chromatin-rich regions of the genome as the primary cause for the regional disparity. Strikingly, we find that increasing levels of transcription reduce mutation prevalence on both strands of gene bodies embedded within H3K9me3-dense regions, and only to those levels observed in H3K9me3-sparse regions, also in an XPC-dependent manner. Therefore, transcription appears to reduce mutation prevalence specifically by relieving the constraints imposed by chromatin structure on DNA repair. We model this relationship among transcription, chromatin state, and DNA repair, revealing a new, personalized determinant of cancer risk.

  3. TFIIS-Dependent Non-coding Transcription Regulates Developmental Genome Rearrangements.

    Directory of Open Access Journals (Sweden)

    Kamila Maliszewska-Olejniczak

    2015-07-01

    Full Text Available 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

  4. Conflict Resolution in the Genome: How Transcription and Replication Make It Work.

    Science.gov (United States)

    Hamperl, Stephan; Cimprich, Karlene A

    2016-12-01

    The complex machineries involved in replication and transcription translocate along the same DNA template, often in opposing directions and at different rates. These processes routinely interfere with each other in prokaryotes, and mounting evidence now suggests that RNA polymerase complexes also encounter replication forks in higher eukaryotes. Indeed, cells rely on numerous mechanisms to avoid, tolerate, and resolve such transcription-replication conflicts, and the absence of these mechanisms can lead to catastrophic effects on genome stability and cell viability. In this article, we review the cellular responses to transcription-replication conflicts and highlight how these inevitable encounters shape the genome and impact diverse cellular processes. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Revised genomic structure of the human ghrelin gene and identification of novel exons, alternative splice variants and natural antisense transcripts

    Directory of Open Access Journals (Sweden)

    Herington Adrian C

    2007-08-01

    Full Text Available Abstract Background Ghrelin is a multifunctional peptide hormone expressed in a range of normal tissues and pathologies. It has been reported that the human ghrelin gene consists of five exons which span 5 kb of genomic DNA on chromosome 3 and includes a 20 bp non-coding first exon (20 bp exon 0. The availability of bioinformatic tools enabling comparative analysis and the finalisation of the human genome prompted us to re-examine the genomic structure of the ghrelin locus. Results We have demonstrated the presence of an additional novel exon (exon -1 and 5' extensions to exon 0 and 1 using comparative in silico analysis and have demonstrated their existence experimentally using RT-PCR and 5' RACE. A revised exon-intron structure demonstrates that the human ghrelin gene spans 7.2 kb and consists of six rather than five exons. Several ghrelin gene-derived splice forms were detected in a range of human tissues and cell lines. We have demonstrated ghrelin gene-derived mRNA transcripts that do not code for ghrelin, but instead may encode the C-terminal region of full-length preproghrelin (C-ghrelin, which contains the coding region for obestatin and a transcript encoding obestatin-only. Splice variants that differed in their 5' untranslated regions were also found, suggesting a role of these regions in the post-transcriptional regulation of preproghrelin translation. Finally, several natural antisense transcripts, termed ghrelinOS (ghrelin opposite strand transcripts, were demonstrated via orientation-specific RT-PCR, 5' RACE and in silico analysis of ESTs and cloned amplicons. Conclusion The sense and antisense alternative transcripts demonstrated in this study may function as non-coding regulatory RNA, or code for novel protein isoforms. This is the first demonstration of putative obestatin and C-ghrelin specific transcripts and these findings suggest that these ghrelin gene-derived peptides may also be produced independently of preproghrelin

  6. The integrated microbial genome resource of analysis.

    Science.gov (United States)

    Checcucci, Alice; Mengoni, Alessio

    2015-01-01

    Integrated Microbial Genomes and Metagenomes (IMG) is a biocomputational system that allows to provide information and support for annotation and comparative analysis of microbial genomes and metagenomes. IMG has been developed by the US Department of Energy (DOE)-Joint Genome Institute (JGI). IMG platform contains both draft and complete genomes, sequenced by Joint Genome Institute and other public and available genomes. Genomes of strains belonging to Archaea, Bacteria, and Eukarya domains are present as well as those of viruses and plasmids. Here, we provide some essential features of IMG system and case study for pangenome analysis.

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

    Science.gov (United States)

    Boldogköi, Zsolt

    2012-01-01

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

  8. RNA-Seq-Based Transcript Structure Analysis with TrBorderExt.

    Science.gov (United States)

    Wang, Yejun; Sun, Ming-An; White, Aaron P

    2018-01-01

    RNA-Seq has become a routine strategy for genome-wide gene expression comparisons in bacteria. Despite lower resolution in transcript border parsing compared with dRNA-Seq, TSS-EMOTE, Cappable-seq, Term-seq, and others, directional RNA-Seq still illustrates its advantages: low cost, quantification and transcript border analysis with a medium resolution (±10-20 nt). To facilitate mining of directional RNA-Seq datasets especially with respect to transcript structure analysis, we developed a tool, TrBorderExt, which can parse transcript start sites and termination sites accurately in bacteria. A detailed protocol is described in this chapter for how to use the software package step by step to identify bacterial transcript borders from raw RNA-Seq data. The package was developed with Perl and R programming languages, and is accessible freely through the website: http://www.szu-bioinf.org/TrBorderExt .

  9. Induced Genome-Wide Binding of Three Arabidopsis WRKY Transcription Factors during Early MAMP-Triggered Immunity.

    Science.gov (United States)

    Birkenbihl, Rainer P; Kracher, Barbara; Somssich, Imre E

    2017-01-01

    During microbial-associated molecular pattern-triggered immunity (MTI), molecules derived from microbes are perceived by cell surface receptors and upon signaling to the nucleus initiate a massive transcriptional reprogramming critical to mount an appropriate host defense response. WRKY transcription factors play an important role in regulating these transcriptional processes. Here, we determined on a genome-wide scale the flg22-induced in vivo DNA binding dynamics of three of the most prominent WRKY factors, WRKY18, WRKY40, and WRKY33. The three WRKY factors each bound to more than 1000 gene loci predominantly at W-box elements, the known WRKY binding motif. Binding occurred mainly in the 500-bp promoter regions of these genes. Many of the targeted genes are involved in signal perception and transduction not only during MTI but also upon damage-associated molecular pattern-triggered immunity, providing a mechanistic link between these functionally interconnected basal defense pathways. Among the additional targets were genes involved in the production of indolic secondary metabolites and in modulating distinct plant hormone pathways. Importantly, among the targeted genes were numerous transcription factors, encoding predominantly ethylene response factors, active during early MTI, and WRKY factors, supporting the previously hypothesized existence of a WRKY subregulatory network. Transcriptional analysis revealed that WRKY18 and WRKY40 function redundantly as negative regulators of flg22-induced genes often to prevent exaggerated defense responses. © 2016 American Society of Plant Biologists. All rights reserved.

  10. Modifiers of notch transcriptional activity identified by genome-wide RNAi

    Directory of Open Access Journals (Sweden)

    Firnhaber Christopher B

    2010-10-01

    Full Text Available Abstract Background The Notch signaling pathway regulates a diverse array of developmental processes, and aberrant Notch signaling can lead to diseases, including cancer. To obtain a more comprehensive understanding of the genetic network that integrates into Notch signaling, we performed a genome-wide RNAi screen in Drosophila cell culture to identify genes that modify Notch-dependent transcription. Results Employing complementary data analyses, we found 399 putative modifiers: 189 promoting and 210 antagonizing Notch activated transcription. These modifiers included several known Notch interactors, validating the robustness of the assay. Many novel modifiers were also identified, covering a range of cellular localizations from the extracellular matrix to the nucleus, as well as a large number of proteins with unknown function. Chromatin-modifying proteins represent a major class of genes identified, including histone deacetylase and demethylase complex components and other chromatin modifying, remodeling and replacement factors. A protein-protein interaction map of the Notch-dependent transcription modifiers revealed that a large number of the identified proteins interact physically with these core chromatin components. Conclusions The genome-wide RNAi screen identified many genes that can modulate Notch transcriptional output. A protein interaction map of the identified genes highlighted a network of chromatin-modifying enzymes and remodelers that regulate Notch transcription. Our results open new avenues to explore the mechanisms of Notch signal regulation and the integration of this pathway into diverse cellular processes.

  11. Transcriptional Regulation During Zygotic Genome Activation in Zebrafish and Other Anamniote Embryos.

    Science.gov (United States)

    Wragg, J; Müller, F

    2016-01-01

    Embryo development commences with the fusion of two terminally differentiated haploid gametes into the totipotent fertilized egg, which through a series of major cellular and molecular transitions generate a pluripotent cell mass. The activation of the zygotic genome occurs during the so-called maternal to zygotic transition and prepares the embryo for zygotic takeover from maternal factors, in the control of the development of cellular lineages during differentiation. Recent advances in next generation sequencing technologies have allowed the dissection of the genomic and epigenomic processes mediating this transition. These processes include reorganization of the chromatin structure to a transcriptionally permissive state, changes in composition and function of structural and regulatory DNA-binding proteins, and changeover of the transcriptome as it is overhauled from that deposited by the mother in the oocyte to a zygotically transcribed complement. Zygotic genome activation in zebrafish occurs 10 cell cycles after fertilization and provides an ideal experimental platform for elucidating the temporal sequence and dynamics of establishment of a transcriptionally active chromatin state and helps in identifying the determinants of transcription activation at polymerase II transcribed gene promoters. The relatively large number of pluripotent cells generated by the fast cell divisions before zygotic transcription provides sufficient biomass for next generation sequencing technology approaches to establish the temporal dynamics of events and suggest causative relationship between them. However, genomic and genetic technologies need to be improved further to capture the earliest events in development, where cell number is a limiting factor. These technologies need to be complemented with precise, inducible genetic interference studies using the latest genome editing tools to reveal the function of candidate determinants and to confirm the predictions made by classic

  12. Transcriptional analysis of phloem-associated cells of potato.

    Science.gov (United States)

    Lin, Tian; Lashbrook, Coralie C; Cho, Sung Ki; Butler, Nathaniel M; Sharma, Pooja; Muppirala, Usha; Severin, Andrew J; Hannapel, David J

    2015-09-03

    Numerous signal molecules, including proteins and mRNAs, are transported through the architecture of plants via the vascular system. As the connection between leaves and other organs, the petiole and stem are especially important in their transport function, which is carried out by the phloem and xylem, especially by the sieve elements in the phloem system. The phloem is an important conduit for transporting photosynthate and signal molecules like metabolites, proteins, small RNAs, and full-length mRNAs. Phloem sap has been used as an unadulterated source to profile phloem proteins and RNAs, but unfortunately, pure phloem sap cannot be obtained in most plant species. Here we make use of laser capture microdissection (LCM) and RNA-seq for an in-depth transcriptional profile of phloem-associated cells of both petioles and stems of potato. To expedite our analysis, we have taken advantage of the potato genome that has recently been fully sequenced and annotated. Out of the 27 k transcripts assembled that we identified, approximately 15 k were present in phloem-associated cells of petiole and stem with greater than ten reads. Among these genes, roughly 10 k are affected by photoperiod. Several RNAs from this day length-regulated group are also abundant in phloem cells of petioles and encode for proteins involved in signaling or transcriptional control. Approximately 22 % of the transcripts in phloem cells contained at least one binding motif for Pumilio, Nova, or polypyrimidine tract-binding proteins in their downstream sequences. Highlighting the predominance of binding processes identified in the gene ontology analysis of active genes from phloem cells, 78 % of the 464 RNA-binding proteins present in the potato genome were detected in our phloem transcriptome. As a reasonable alternative when phloem sap collection is not possible, LCM can be used to isolate RNA from specific cell types, and along with RNA-seq, provides practical access to expression profiles of

  13. Pan-Cancer Mutational and Transcriptional Analysis of the Integrator Complex

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    Antonio Federico

    2017-04-01

    Full Text Available The integrator complex has been recently identified as a key regulator of RNA Polymerase II-mediated transcription, with many functions including the processing of small nuclear RNAs, the pause-release and elongation of polymerase during the transcription of protein coding genes, and the biogenesis of enhancer derived transcripts. Moreover, some of its components also play a role in genome maintenance. Thus, it is reasonable to hypothesize that their functional impairment or altered expression can contribute to malignancies. Indeed, several studies have described the mutations or transcriptional alteration of some Integrator genes in different cancers. Here, to draw a comprehensive pan-cancer picture of the genomic and transcriptomic alterations for the members of the complex, we reanalyzed public data from The Cancer Genome Atlas. Somatic mutations affecting Integrator subunit genes and their transcriptional profiles have been investigated in about 11,000 patients and 31 tumor types. A general heterogeneity in the mutation frequencies was observed, mostly depending on tumor type. Despite the fact that we could not establish them as cancer drivers, INTS7 and INTS8 genes were highly mutated in specific cancers. A transcriptome analysis of paired (normal and tumor samples revealed that the transcription of INTS7, INTS8, and INTS13 is significantly altered in several cancers. Experimental validation performed on primary tumors confirmed these findings.

  14. Analysis tools for the interplay between genome layout and regulation.

    Science.gov (United States)

    Bouyioukos, Costas; Elati, Mohamed; Képès, François

    2016-06-06

    Genome layout and gene regulation appear to be interdependent. Understanding this interdependence is key to exploring the dynamic nature of chromosome conformation and to engineering functional genomes. Evidence for non-random genome layout, defined as the relative positioning of either co-functional or co-regulated genes, stems from two main approaches. Firstly, the analysis of contiguous genome segments across species, has highlighted the conservation of gene arrangement (synteny) along chromosomal regions. Secondly, the study of long-range interactions along a chromosome has emphasised regularities in the positioning of microbial genes that are co-regulated, co-expressed or evolutionarily correlated. While one-dimensional pattern analysis is a mature field, it is often powerless on biological datasets which tend to be incomplete, and partly incorrect. Moreover, there is a lack of comprehensive, user-friendly tools to systematically analyse, visualise, integrate and exploit regularities along genomes. Here we present the Genome REgulatory and Architecture Tools SCAN (GREAT:SCAN) software for the systematic study of the interplay between genome layout and gene expression regulation. SCAN is a collection of related and interconnected applications currently able to perform systematic analyses of genome regularities as well as to improve transcription factor binding sites (TFBS) and gene regulatory network predictions based on gene positional information. We demonstrate the capabilities of these tools by studying on one hand the regular patterns of genome layout in the major regulons of the bacterium Escherichia coli. On the other hand, we demonstrate the capabilities to improve TFBS prediction in microbes. Finally, we highlight, by visualisation of multivariate techniques, the interplay between position and sequence information for effective transcription regulation.

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

    Science.gov (United States)

    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 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.

  16. Next-Generation Sequencing of Genomic DNA Fragments Bound to a Transcription Factor in Vitro Reveals Its Regulatory Potential

    Directory of Open Access Journals (Sweden)

    Yukio Kurihara

    2014-12-01

    Full Text Available Several transcription factors (TFs coordinate to regulate expression of specific genes at the transcriptional level. In Arabidopsis thaliana it is estimated that approximately 10% of all genes encode TFs or TF-like proteins. It is important to identify target genes that are directly regulated by TFs in order to understand the complete picture of a plant’s transcriptome profile. Here, we investigate the role of the LONG HYPOCOTYL5 (HY5 transcription factor that acts as a regulator of photomorphogenesis. We used an in vitro genomic DNA binding assay coupled with immunoprecipitation and next-generation sequencing (gDB-seq instead of the in vivo chromatin immunoprecipitation (ChIP-based methods. The results demonstrate that the HY5-binding motif predicted here was similar to the motif reported previously and that in vitro HY5-binding loci largely overlapped with the HY5-targeted candidate genes identified in previous ChIP-chip analysis. By combining these results with microarray analysis, we identified hundreds of HY5-binding genes that were differentially expressed in hy5. We also observed delayed induction of some transcripts of HY5-binding genes in hy5 mutants in response to blue-light exposure after dark treatment. Thus, an in vitro gDNA-binding assay coupled with sequencing is a convenient and powerful method to bridge the gap between identifying TF binding potential and establishing function.

  17. Two transcription products of the vesicular stomatitis virus genome may control L-cell protein synthesis

    International Nuclear Information System (INIS)

    Dunigan, D.D.; Lucas-Lenard, J.M.

    1983-01-01

    When mouse L-cells are infected with vesicular stomatitis virus, there is a decrease in the rate of protein synthesis ranging from 20 to 85% of that in mock-infected cells. Vesicular stomatitis virus, irradiated with increasing doses of UV light, eventually loses this capacity to inhibit protein synthesis. The UV inactivation curve was biphasic, suggesting that transcription of two regions of the viral genome is necessary for the virus to become inactivated in this capacity. The first transcription produced corresponded to about 373 nucleotides, and the second corresponded to about 42 nucleotides. Inhibition of transcription of the larger product by irradiating the virus with low doses of UV light left a residual inhibition of protein synthesis consisting of approximately 60 to 65% of the total inhibition. This residual inhibition could be obviated by irradiating the virus with a UV dose of greater than 20,000 ergs/mm 2 and was thus considered to represent the effect of the smaller transcription product. In the R1 mutant of another author, the inhibition of transcription of the larger product sufficed to restore protein synthesis to the mock-infected level, suggesting that the smaller transcription product is nonfunctional with respect to protein synthesis inhibition. Extracts from cells infected with virus irradiated with low doses of UV light showed a protein synthesis capacity quite similar to that of their in vivo counterparts, indicating that these extracts closely reflect the in vivo effects of virus infection

  18. Strawberry: Fast and accurate genome-guided transcript reconstruction and quantification from RNA-Seq.

    Science.gov (United States)

    Liu, Ruolin; Dickerson, Julie

    2017-11-01

    We propose a novel method and software tool, Strawberry, for transcript reconstruction and quantification from RNA-Seq data under the guidance of genome alignment and independent of gene annotation. Strawberry consists of two modules: assembly and quantification. The novelty of Strawberry is that the two modules use different optimization frameworks but utilize the same data graph structure, which allows a highly efficient, expandable and accurate algorithm for dealing large data. The assembly module parses aligned reads into splicing graphs, and uses network flow algorithms to select the most likely transcripts. The quantification module uses a latent class model to assign read counts from the nodes of splicing graphs to transcripts. Strawberry simultaneously estimates the transcript abundances and corrects for sequencing bias through an EM algorithm. Based on simulations, Strawberry outperforms Cufflinks and StringTie in terms of both assembly and quantification accuracies. Under the evaluation of a real data set, the estimated transcript expression by Strawberry has the highest correlation with Nanostring probe counts, an independent experiment measure for transcript expression. Strawberry is written in C++14, and is available as open source software at https://github.com/ruolin/strawberry under the MIT license.

  19. Comparative genomics of CytR, an unusual member of the LacI family of transcription factors.

    Directory of Open Access Journals (Sweden)

    Natalia V Sernova

    Full Text Available CytR is a transcription regulator from the LacI family, present in some gamma-proteobacteria including Escherichia coli and known not only for its cellular role, control of transport and utilization of nucleosides, but for a number of unusual structural properties. The present study addressed three related problems: structure of CytR-binding sites and motifs, their evolutionary conservation, and identification of new members of the CytR regulon. While the majority of CytR-binding sites are imperfect inverted repeats situated between binding sites for another transcription factor, CRP, other architectures were observed, in particular, direct repeats. While the similarity between sites for different genes in one genome is rather low, and hence the consensus motif is weak, there is high conservation of orthologous sites in different genomes (mainly in the Enterobacteriales arguing for the presence of specific CytR-DNA contacts. On larger evolutionary distances candidate CytR sites may migrate but the approximate distance between flanking CRP sites tends to be conserved, which demonstrates that the overall structure of the CRP-CytR-DNA complex is gene-specific. The analysis yielded candidate CytR-binding sites for orthologs of known regulon members in less studied genomes of the Enterobacteriales and Vibrionales and identified a new candidate member of the CytR regulon, encoding a transporter named NupT (YcdZ.

  20. Mining whole genomes and transcriptomes of Jatropha (Jatropha curcas) and Castor bean (Ricinus communis) for NBS-LRR genes and defense response associated transcription factors.

    Science.gov (United States)

    Sood, Archit; Jaiswal, Varun; Chanumolu, Sree Krishna; Malhotra, Nikhil; Pal, Tarun; Chauhan, Rajinder Singh

    2014-11-01

    Jatropha (Jatropha curcas L.) and Castor bean (Ricinus communis) are oilseed crops of family Euphorbiaceae with the potential of producing high quality biodiesel and having industrial value. Both the bioenergy plants are becoming susceptible to various biotic stresses directly affecting the oil quality and content. No report exists as of today on analysis of Nucleotide Binding Site-Leucine Rich Repeat (NBS-LRR) gene repertoire and defense response transcription factors in both the plant species. In silico analysis of whole genomes and transcriptomes identified 47 new NBS-LRR genes in both the species and 122 and 318 defense response related transcription factors in Jatropha and Castor bean, respectively. The identified NBS-LRR genes and defense response transcription factors were mapped onto the respective genomes. Common and unique NBS-LRR genes and defense related transcription factors were identified in both the plant species. All NBS-LRR genes in both the species were characterized into Toll/interleukin-1 receptor NBS-LRRs (TNLs) and coiled-coil NBS-LRRs (CNLs), position on contigs, gene clusters and motifs and domains distribution. Transcript abundance or expression values were measured for all NBS-LRR genes and defense response transcription factors, suggesting their functional role. The current study provides a repertoire of NBS-LRR genes and transcription factors which can be used in not only dissecting the molecular basis of disease resistance phenotype but also in developing disease resistant genotypes in Jatropha and Castor bean through transgenic or molecular breeding approaches.

  1. Coordination of genomic structure and transcription by the main bacterial nucleoid-associated protein HU

    Science.gov (United States)

    Berger, Michael; Farcas, Anca; Geertz, Marcel; Zhelyazkova, Petya; Brix, Klaudia; Travers, Andrew; Muskhelishvili, Georgi

    2010-01-01

    The histone-like protein HU is a highly abundant DNA architectural protein that is involved in compacting the DNA of the bacterial nucleoid and in regulating the main DNA transactions, including gene transcription. However, the coordination of the genomic structure and function by HU is poorly understood. Here, we address this question by comparing transcript patterns and spatial distributions of RNA polymerase in Escherichia coli wild-type and hupA/B mutant cells. We demonstrate that, in mutant cells, upregulated genes are preferentially clustered in a large chromosomal domain comprising the ribosomal RNA operons organized on both sides of OriC. Furthermore, we show that, in parallel to this transcription asymmetry, mutant cells are also impaired in forming the transcription foci—spatially confined aggregations of RNA polymerase molecules transcribing strong ribosomal RNA operons. Our data thus implicate HU in coordinating the global genomic structure and function by regulating the spatial distribution of RNA polymerase in the nucleoid. PMID:20010798

  2. A cysteine protease (cathepsin Z) from disk abalone, Haliotis discus discus: Genomic characterization and transcriptional profiling during bacterial infections.

    Science.gov (United States)

    Godahewa, G I; Perera, N C N; Lee, Sukkyoung; Kim, Myoung-Jin; Lee, Jehee

    2017-09-05

    Cathepsin Z (CTSZ) is lysosomal cysteine protease of the papain superfamily. It participates in the host immune defense via phagocytosis, signal transduction, cell-cell communication, proliferation, and migration of immune cells such as monocytes, macrophages, and dendritic cells. Hence, CTSZ is also acknowledged as an acute-phase protein in host immunity. In this study, we sought to identify the CTSZ homolog from disk abalone (AbCTSZ) and characterize it at the molecular, genomic, and transcriptional levels. AbCTSZ encodes a protein with 318 amino acids and a molecular mass of 36kDa. The structure of AbCTSZ reveals amino acid sequences that are characteristic of the signal sequence, pro-peptide, peptidase-C1 papain family cysteine protease domain, mini-loop, HIP motif, N-linked glycosylation sites, active sites, and conserved Cys residues. A pairwise comparison revealed that AbCTSZ shared the highest amino acid homology with its molluscan counterpart from Crassostrea gigas. A multiple alignment analysis revealed the conservation of functionally crucial elements of AbCTSZ, and a phylogenetic study further confirmed a proximal evolutionary relationship with its invertebrate counterparts. Further, an analysis of AbCTSZ genomic structure revealed seven exons separated by six introns, which differs from that of its vertebrate counterparts. Quantitative real time PCR (qPCR) detected the transcripts of AbCTSZ in early developmental stages and in eight different tissues. Higher levels of AbCTSZ transcripts were found in trochophore, gill, and hemocytes, highlighting its importance in the early development and immunity of disk abalone. In addition, we found that viable bacteria (Vibrio parahaemolyticus and Listeria monocytogenes) and bacterial lipopolysaccharides significantly modulated AbCTSZ transcription. Collectively, these lines of evidences suggest that AbCTSZ plays an indispensable role in the innate immunity of disk abalone. Copyright © 2017. Published by Elsevier

  3. Enhancement of single guide RNA transcription for efficient CRISPR/Cas-based genomic engineering.

    Science.gov (United States)

    Ui-Tei, Kumiko; Maruyama, Shohei; Nakano, Yuko

    2017-06-01

    Genomic engineering using clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) protein is a promising approach for targeting the genomic DNA of virtually any organism in a sequence-specific manner. Recent remarkable advances in CRISPR/Cas technology have made it a feasible system for use in therapeutic applications and biotechnology. In the CRISPR/Cas system, a guide RNA (gRNA), interacting with the Cas protein, recognizes a genomic region with sequence complementarity, and the double-stranded DNA at the target site is cleaved by the Cas protein. A widely used gRNA is an RNA polymerase III (pol III)-driven single gRNA (sgRNA), which is produced by artificial fusion of CRISPR RNA (crRNA) and trans-activation crRNA (tracrRNA). However, we identified a TTTT stretch, known as a termination signal of RNA pol III, in the scaffold region of the sgRNA. Here, we revealed that sgRNA carrying a TTTT stretch reduces the efficiency of sgRNA transcription due to premature transcriptional termination, and decreases the efficiency of genome editing. Unexpectedly, it was also shown that the premature terminated sgRNA may have an adverse effect of inducing RNA interference. Such disadvantageous effects were avoided by substituting one base in the TTTT stretch.

  4. Integrative Genomics Reveals Mechanisms of Copy Number Alterations Responsible for Transcriptional Deregulation in Colorectal Cancer

    Science.gov (United States)

    Camps, Jordi; Nguyen, Quang Tri; Padilla-Nash, Hesed M.; Knutsen, Turid; McNeil, Nicole E.; Wangsa, Danny; Hummon, Amanda B.; Grade, Marian; Ried, Thomas; Difilippantonio, Michael J.

    2016-01-01

    To evaluate the mechanisms and consequences of chromosomal aberrations in colorectal cancer (CRC), we used a combination of spectral karyotyping, array comparative genomic hybridization (aCGH), and array-based global gene expression profiling on 31 primary carcinomas and 15 established cell lines. Importantly, aCGH showed that the genomic profiles of primary tumors are recapitulated in the cell lines. We revealed a preponderance of chromosome breakpoints at sites of copy number variants (CNVs) in the CRC cell lines, a novel mechanism of DNA breakage in cancer. The integration of gene expression and aCGH led to the identification of 157 genes localized within high-level copy number changes whose transcriptional deregulation was significantly affected across all of the samples, thereby suggesting that these genes play a functional role in CRC. Genomic amplification at 8q24 was the most recurrent event and led to the overexpression of MYC and FAM84B. Copy number dependent gene expression resulted in deregulation of known cancer genes such as APC, FGFR2, and ERBB2. The identification of only 36 genes whose localization near a breakpoint could account for their observed deregulated expression demonstrates that the major mechanism for transcriptional deregulation in CRC is genomic copy number changes resulting from chromosomal aberrations. PMID:19691111

  5. RSEM: accurate transcript quantification from RNA-Seq data with or without a reference genome

    Directory of Open Access Journals (Sweden)

    Dewey Colin N

    2011-08-01

    Full Text Available Abstract Background RNA-Seq is revolutionizing the way transcript abundances are measured. A key challenge in transcript quantification from RNA-Seq data is the handling of reads that map to multiple genes or isoforms. This issue is particularly important for quantification with de novo transcriptome assemblies in the absence of sequenced genomes, as it is difficult to determine which transcripts are isoforms of the same gene. A second significant issue is the design of RNA-Seq experiments, in terms of the number of reads, read length, and whether reads come from one or both ends of cDNA fragments. Results We present RSEM, an user-friendly software package for quantifying gene and isoform abundances from single-end or paired-end RNA-Seq data. RSEM outputs abundance estimates, 95% credibility intervals, and visualization files and can also simulate RNA-Seq data. In contrast to other existing tools, the software does not require a reference genome. Thus, in combination with a de novo transcriptome assembler, RSEM enables accurate transcript quantification for species without sequenced genomes. On simulated and real data sets, RSEM has superior or comparable performance to quantification methods that rely on a reference genome. Taking advantage of RSEM's ability to effectively use ambiguously-mapping reads, we show that accurate gene-level abundance estimates are best obtained with large numbers of short single-end reads. On the other hand, estimates of the relative frequencies of isoforms within single genes may be improved through the use of paired-end reads, depending on the number of possible splice forms for each gene. Conclusions RSEM is an accurate and user-friendly software tool for quantifying transcript abundances from RNA-Seq data. As it does not rely on the existence of a reference genome, it is particularly useful for quantification with de novo transcriptome assemblies. In addition, RSEM has enabled valuable guidance for cost

  6. Global Analysis of Photosynthesis Transcriptional Regulatory Networks

    Science.gov (United States)

    Imam, Saheed; Noguera, Daniel R.; Donohue, Timothy J.

    2014-01-01

    Photosynthesis is a crucial biological process that depends on the interplay of many components. This work analyzed the gene targets for 4 transcription factors: FnrL, PrrA, CrpK and MppG (RSP_2888), which are known or predicted to control photosynthesis in Rhodobacter sphaeroides. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) identified 52 operons under direct control of FnrL, illustrating its regulatory role in photosynthesis, iron homeostasis, nitrogen metabolism and regulation of sRNA synthesis. Using global gene expression analysis combined with ChIP-seq, we mapped the regulons of PrrA, CrpK and MppG. PrrA regulates ∼34 operons encoding mainly photosynthesis and electron transport functions, while CrpK, a previously uncharacterized Crp-family protein, regulates genes involved in photosynthesis and maintenance of iron homeostasis. Furthermore, CrpK and FnrL share similar DNA binding determinants, possibly explaining our observation of the ability of CrpK to partially compensate for the growth defects of a ΔFnrL mutant. We show that the Rrf2 family protein, MppG, plays an important role in photopigment biosynthesis, as part of an incoherent feed-forward loop with PrrA. Our results reveal a previously unrealized, high degree of combinatorial regulation of photosynthetic genes and significant cross-talk between their transcriptional regulators, while illustrating previously unidentified links between photosynthesis and the maintenance of iron homeostasis. PMID:25503406

  7. Global analysis of photosynthesis transcriptional regulatory networks.

    Directory of Open Access Journals (Sweden)

    Saheed Imam

    2014-12-01

    Full Text Available Photosynthesis is a crucial biological process that depends on the interplay of many components. This work analyzed the gene targets for 4 transcription factors: FnrL, PrrA, CrpK and MppG (RSP_2888, which are known or predicted to control photosynthesis in Rhodobacter sphaeroides. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq identified 52 operons under direct control of FnrL, illustrating its regulatory role in photosynthesis, iron homeostasis, nitrogen metabolism and regulation of sRNA synthesis. Using global gene expression analysis combined with ChIP-seq, we mapped the regulons of PrrA, CrpK and MppG. PrrA regulates ∼34 operons encoding mainly photosynthesis and electron transport functions, while CrpK, a previously uncharacterized Crp-family protein, regulates genes involved in photosynthesis and maintenance of iron homeostasis. Furthermore, CrpK and FnrL share similar DNA binding determinants, possibly explaining our observation of the ability of CrpK to partially compensate for the growth defects of a ΔFnrL mutant. We show that the Rrf2 family protein, MppG, plays an important role in photopigment biosynthesis, as part of an incoherent feed-forward loop with PrrA. Our results reveal a previously unrealized, high degree of combinatorial regulation of photosynthetic genes and significant cross-talk between their transcriptional regulators, while illustrating previously unidentified links between photosynthesis and the maintenance of iron homeostasis.

  8. Inter-replicon Gene Flow Contributes to Transcriptional Integration in the Sinorhizobium meliloti Multipartite Genome

    Directory of Open Access Journals (Sweden)

    George C. diCenzo

    2018-05-01

    Full Text Available Integration of newly acquired genes into existing regulatory networks is necessary for successful horizontal gene transfer (HGT. Ten percent of bacterial species contain at least two DNA replicons over 300 kilobases in size, with the secondary replicons derived predominately through HGT. The Sinorhizobium meliloti genome is split between a 3.7 Mb chromosome, a 1.7 Mb chromid consisting largely of genes acquired through ancient HGT, and a 1.4 Mb megaplasmid consisting primarily of recently acquired genes. Here, RNA-sequencing is used to examine the transcriptional consequences of massive, synthetic genome reduction produced through the removal of the megaplasmid and/or the chromid. Removal of the pSymA megaplasmid influenced the transcription of only six genes. In contrast, removal of the chromid influenced expression of ∼8% of chromosomal genes and ∼4% of megaplasmid genes. This was mediated in part by the loss of the ETR DNA region whose presence on pSymB is due to a translocation from the chromosome. No obvious functional bias among the up-regulated genes was detected, although genes with putative homologs on the chromid were enriched. Down-regulated genes were enriched in motility and sensory transduction pathways. Four transcripts were examined further, and in each case the transcriptional change could be traced to loss of specific pSymB regions. In particularly, a chromosomal transporter was induced due to deletion of bdhA likely mediated through 3-hydroxybutyrate accumulation. These data provide new insights into the evolution of the multipartite bacterial genome, and more generally into the integration of horizontally acquired genes into the transcriptome.

  9. Inter-replicon Gene Flow Contributes to Transcriptional Integration in the Sinorhizobium meliloti Multipartite Genome.

    Science.gov (United States)

    diCenzo, George C; Wellappili, Deelaka; Golding, G Brian; Finan, Turlough M

    2018-05-04

    Integration of newly acquired genes into existing regulatory networks is necessary for successful horizontal gene transfer (HGT). Ten percent of bacterial species contain at least two DNA replicons over 300 kilobases in size, with the secondary replicons derived predominately through HGT. The Sinorhizobium meliloti genome is split between a 3.7 Mb chromosome, a 1.7 Mb chromid consisting largely of genes acquired through ancient HGT, and a 1.4 Mb megaplasmid consisting primarily of recently acquired genes. Here, RNA-sequencing is used to examine the transcriptional consequences of massive, synthetic genome reduction produced through the removal of the megaplasmid and/or the chromid. Removal of the pSymA megaplasmid influenced the transcription of only six genes. In contrast, removal of the chromid influenced expression of ∼8% of chromosomal genes and ∼4% of megaplasmid genes. This was mediated in part by the loss of the ETR DNA region whose presence on pSymB is due to a translocation from the chromosome. No obvious functional bias among the up-regulated genes was detected, although genes with putative homologs on the chromid were enriched. Down-regulated genes were enriched in motility and sensory transduction pathways. Four transcripts were examined further, and in each case the transcriptional change could be traced to loss of specific pSymB regions. In particularly, a chromosomal transporter was induced due to deletion of bdhA likely mediated through 3-hydroxybutyrate accumulation. These data provide new insights into the evolution of the multipartite bacterial genome, and more generally into the integration of horizontally acquired genes into the transcriptome. Copyright © 2018 diCenzo, et al.

  10. FY 1999 Industrial science and technology research and development project. Report on the results of research and development of the technologies for genome informatics (Acceleration of analysis of green mold transcription control information); 1999 nendo genome infomatics gijutsu kenkyu kaihatsu seika hokokusho. Koji kabi no tensha seigyo joho no kaiseki kasokuka nado

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2001-03-01

    A total of 49 budding yeast transcription factor disruptants and one conditional transcription over expression strain are produced, to elucidate the gene regulation networks using the gene expression profile data, and to measure the systematic and high-quality gene expression profiles using the Affymetrix's GeneChip system. The program is also developed for accurately predicting the base sequences which regulate expression of given gene groups, based on the uniqueness of the upstream sequences. The analysis with the aid of the program predicts 8 gene expression regulation sequences, which are considered to be novel, from the gene groups of retarded expression by the transcription factor disruptants. The time course gene expression data are produced from the transcription factor SW14 conditional over expression strain. The analysis of the data indicates that the analysis of the subtracted genes using the gene expression profiles from the wild type strain is useful for clarifying the effects of the derived transcription factor over expression. (NEDO)

  11. Transcriptional analysis of apple fruit proanthocyanidin biosynthesis

    Science.gov (United States)

    Henry-Kirk, Rebecca A.

    2012-01-01

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

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

    DEFF Research Database (Denmark)

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

    2014-01-01

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

  13. Assessing quality and completeness of human transcriptional regulatory pathways on a genome-wide scale

    Directory of Open Access Journals (Sweden)

    Aifantis Iannis

    2011-02-01

    Full Text Available Abstract Background Pathway databases are becoming increasingly important and almost omnipresent in most types of biological and translational research. However, little is known about the quality and completeness of pathways stored in these databases. The present study conducts a comprehensive assessment of transcriptional regulatory pathways in humans for seven well-studied transcription factors: MYC, NOTCH1, BCL6, TP53, AR, STAT1, and RELA. The employed benchmarking methodology first involves integrating genome-wide binding with functional gene expression data to derive direct targets of transcription factors. Then the lists of experimentally obtained direct targets are compared with relevant lists of transcriptional targets from 10 commonly used pathway databases. Results The results of this study show that for the majority of pathway databases, the overlap between experimentally obtained target genes and targets reported in transcriptional regulatory pathway databases is surprisingly small and often is not statistically significant. The only exception is MetaCore pathway database which yields statistically significant intersection with experimental results in 84% cases. Additionally, we suggest that the lists of experimentally derived direct targets obtained in this study can be used to reveal new biological insight in transcriptional regulation and suggest novel putative therapeutic targets in cancer. Conclusions Our study opens a debate on validity of using many popular pathway databases to obtain transcriptional regulatory targets. We conclude that the choice of pathway databases should be informed by solid scientific evidence and rigorous empirical evaluation. Reviewers This article was reviewed by Prof. Wing Hung Wong, Dr. Thiago Motta Venancio (nominated by Dr. L Aravind, and Prof. Geoff J McLachlan.

  14. Genome-wide transcriptional responses of Alteromonas naphthalenivorans SN2 to contaminated seawater and marine tidal flat sediment.

    Science.gov (United States)

    Jin, Hyun Mi; Jeong, Hye Im; Kim, Kyung Hyun; Hahn, Yoonsoo; Madsen, Eugene L; Jeon, Che Ok

    2016-02-18

    A genome-wide transcriptional analysis of Alteromonas naphthalenivorans SN2 was performed to investigate its ecophysiological behavior in contaminated tidal flats and seawater. The experimental design mimicked these habitats that either added naphthalene or pyruvate; tidal flat-naphthalene (TF-N), tidal flat-pyruvate (TF-P), seawater-naphthalene (SW-N), and seawater-pyruvate (SW-P). The transcriptional profiles clustered by habitat (TF-N/TF-P and SW-N/SW-P), rather than carbon source, suggesting that the former may exert a greater influence on genome-wide expression in strain SN2 than the latter. Metabolic mapping of cDNA reads from strain SN2 based on KEGG pathway showed that metabolic and regulatory genes associated with energy metabolism, translation, and cell motility were highly expressed in all four test conditions, probably highlighting the copiotrophic properties of strain SN2 as an opportunistic marine r-strategist. Differential gene expression analysis revealed that strain SN2 displayed specific cellular responses to environmental variables (tidal flat, seawater, naphthalene, and pyruvate) and exhibited certain ecological fitness traits -- its notable PAH degradation capability in seasonally cold tidal flat might be reflected in elevated expression of stress response and chaperone proteins, while fast growth in nitrogen-deficient and aerobic seawater probably correlated with high expression of glutamine synthetase, enzymes utilizing nitrite/nitrate, and those involved in the removal of reactive oxygen species.

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

    Directory of Open Access Journals (Sweden)

    Reini F Luco

    2008-05-01

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

  16. Genome-wide dynamic transcriptional profiling in clostridium beijerinckii NCIMB 8052 using single-nucleotide resolution RNA-Seq

    Directory of Open Access Journals (Sweden)

    Wang Yi

    2012-03-01

    Full Text Available Abstract Background Clostridium beijerinckii is a prominent solvent-producing microbe that has great potential for biofuel and chemical industries. Although transcriptional analysis is essential to understand gene functions and regulation and thus elucidate proper strategies for further strain improvement, limited information is available on the genome-wide transcriptional analysis for C. beijerinckii. Results The genome-wide transcriptional dynamics of C. beijerinckii NCIMB 8052 over a batch fermentation process was investigated using high-throughput RNA-Seq technology. The gene expression profiles indicated that the glycolysis genes were highly expressed throughout the fermentation, with comparatively more active expression during acidogenesis phase. The expression of acid formation genes was down-regulated at the onset of solvent formation, in accordance with the metabolic pathway shift from acidogenesis to solventogenesis. The acetone formation gene (adc, as a part of the sol operon, exhibited highly-coordinated expression with the other sol genes. Out of the > 20 genes encoding alcohol dehydrogenase in C. beijerinckii, Cbei_1722 and Cbei_2181 were highly up-regulated at the onset of solventogenesis, corresponding to their key roles in primary alcohol production. Most sporulation genes in C. beijerinckii 8052 demonstrated similar temporal expression patterns to those observed in B. subtilis and C. acetobutylicum, while sporulation sigma factor genes sigE and sigG exhibited accelerated and stronger expression in C. beijerinckii 8052, which is consistent with the more rapid forespore and endspore development in this strain. Global expression patterns for specific gene functional classes were examined using self-organizing map analysis. The genes associated with specific functional classes demonstrated global expression profiles corresponding to the cell physiological variation and metabolic pathway switch. Conclusions The results from this

  17. A Transcription Activator-Like Effector (TALE) Toolbox for Genome Engineering

    Science.gov (United States)

    Sanjana, Neville E.; Cong, Le; Zhou, Yang; Cunniff, Margaret M.; Feng, Guoping; Zhang, Feng

    2013-01-01

    Transcription activator-like effectors (TALEs) are a class of naturally occurring DNA binding proteins found in the plant pathogen Xanthomonas sp. The DNA binding domain of each TALE consists of tandem 34-amino acid repeat modules that can be rearranged according to a simple cipher to target new DNA sequences. Customized TALEs can be used for a wide variety of genome engineering applications, including transcriptional modulation and genome editing. Here we describe a toolbox for rapid construction of custom TALE transcription factors (TALE-TFs) and nucleases (TALENs) using a hierarchical ligation procedure. This toolbox facilitates affordable and rapid construction of custom TALE-TFs and TALENs within one week and can be easily scaled up to construct TALEs for multiple targets in parallel. We also provide details for testing the activity in mammalian cells of custom TALE-TFs and TALENs using, respectively, qRT-PCR and Surveyor nuclease. The TALE toolbox described here will enable a broad range of biological applications. PMID:22222791

  18. Transcriptional and Genomic Targets of Neural Stem Cells for Functional Recovery after Hemorrhagic Stroke

    Directory of Open Access Journals (Sweden)

    Le Zhang

    2017-01-01

    Full Text Available Hemorrhagic stroke is a life-threatening disease characterized by a sudden rupture of cerebral blood vessels, and it is widely believed that neural cell death occurs after exposure to blood metabolites or subsequently damaged cells. Neural stem cells (NSCs, which maintain neurogenesis and are found in subgranular zone and subventricular zone, are thought to be an endogenous neuroprotective mechanism for these brain injuries. However, due to the complexity of NSCs and their microenvironment, current strategies cannot satisfactorily enhance functional recovery after hemorrhagic stroke. It is well known that transcriptional and genomic pathways play important roles in ensuring the normal functions of NSCs, including proliferation, migration, differentiation, and neural reconnection. Recently, emerging evidence from the use of new technologies such as next-generation sequencing and transcriptome profiling has provided insight into our understanding of genomic function and regulation of NSCs. In the present article, we summarize and present the current data on the control of NSCs at both the transcriptional and genomic levels. Using bioinformatics methods, we sought to predict novel therapeutic targets of endogenous neurogenesis and exogenous NSC transplantation for functional recovery after hemorrhagic stroke, which could also advance our understanding of its pathophysiology.

  19. Multimode drug inducible CRISPR/Cas9 devices for transcriptional activation and genome editing

    Science.gov (United States)

    Lu, Jia; Zhao, Chen; Zhao, Yingze; Zhang, Jingfang; Zhang, Yue; Chen, Li; Han, Qiyuan; Ying, Yue; Peng, Shuai; Ai, Runna; Wang, Yu

    2018-01-01

    Abstract Precise investigation and manipulation of dynamic biological processes often requires molecular modulation in a controlled inducible manner. The clustered, regularly interspaced, short palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9) has emerged as a versatile tool for targeted gene editing and transcriptional programming. Here, we designed and vigorously optimized a series of Hybrid drug Inducible CRISPR/Cas9 Technologies (HIT) for transcriptional activation by grafting a mutated human estrogen receptor (ERT2) to multiple CRISPR/Cas9 systems, which renders them 4-hydroxytamoxifen (4-OHT) inducible for the access of genome. Further, extra functionality of simultaneous genome editing was achieved with one device we named HIT2. Optimized terminal devices herein delivered advantageous performances in comparison with several existing designs. They exerted selective, titratable, rapid and reversible response to drug induction. In addition, these designs were successfully adapted to an orthogonal Cas9. HIT systems developed in this study can be applied for controlled modulation of potentially any genomic loci in multiple modes. PMID:29237052

  20. Enriching Genomic Resources and Marker Development from Transcript Sequences of Jatropha curcas for Microgravity Studies

    Science.gov (United States)

    Tian, Wenlan; Paudel, Dev

    2017-01-01

    Jatropha (Jatropha curcas L.) is an economically important species with a great potential for biodiesel production. To enrich the jatropha genomic databases and resources for microgravity studies, we sequenced and annotated the transcriptome of jatropha and developed SSR and SNP markers from the transcriptome sequences. In total 1,714,433 raw reads with an average length of 441.2 nucleotides were generated. De novo assembling and clustering resulted in 115,611 uniquely assembled sequences (UASs) including 21,418 full-length cDNAs and 23,264 new jatropha transcript sequences. The whole set of UASs were fully annotated, out of which 59,903 (51.81%) were assigned with gene ontology (GO) term, 12,584 (10.88%) had orthologs in Eukaryotic Orthologous Groups (KOG), and 8,822 (7.63%) were mapped to 317 pathways in six different categories in Kyoto Encyclopedia of Genes and Genome (KEGG) database, and it contained 3,588 putative transcription factors. From the UASs, 9,798 SSRs were discovered with AG/CT as the most frequent (45.8%) SSR motif type. Further 38,693 SNPs were detected and 7,584 remained after filtering. This UAS set has enriched the current jatropha genomic databases and provided a large number of genetic markers, which can facilitate jatropha genetic improvement and many other genetic and biological studies. PMID:28154822

  1. Versatile Gene-Specific Sequence Tags for Arabidopsis Functional Genomics: Transcript Profiling and Reverse Genetics Applications

    Science.gov (United States)

    Hilson, Pierre; Allemeersch, Joke; Altmann, Thomas; Aubourg, Sébastien; Avon, Alexandra; Beynon, Jim; Bhalerao, Rishikesh P.; Bitton, Frédérique; Caboche, Michel; Cannoot, Bernard; Chardakov, Vasil; Cognet-Holliger, Cécile; Colot, Vincent; Crowe, Mark; Darimont, Caroline; Durinck, Steffen; Eickhoff, Holger; de Longevialle, Andéol Falcon; Farmer, Edward E.; Grant, Murray; Kuiper, Martin T.R.; Lehrach, Hans; Léon, Céline; Leyva, Antonio; Lundeberg, Joakim; Lurin, Claire; Moreau, Yves; Nietfeld, Wilfried; Paz-Ares, Javier; Reymond, Philippe; Rouzé, Pierre; Sandberg, Goran; Segura, Maria Dolores; Serizet, Carine; Tabrett, Alexandra; Taconnat, Ludivine; Thareau, Vincent; Van Hummelen, Paul; Vercruysse, Steven; Vuylsteke, Marnik; Weingartner, Magdalena; Weisbeek, Peter J.; Wirta, Valtteri; Wittink, Floyd R.A.; Zabeau, Marc; Small, Ian

    2004-01-01

    Microarray transcript profiling and RNA interference are two new technologies crucial for large-scale gene function studies in multicellular eukaryotes. Both rely on sequence-specific hybridization between complementary nucleic acid strands, inciting us to create a collection of gene-specific sequence tags (GSTs) representing at least 21,500 Arabidopsis genes and which are compatible with both approaches. The GSTs were carefully selected to ensure that each of them shared no significant similarity with any other region in the Arabidopsis genome. They were synthesized by PCR amplification from genomic DNA. Spotted microarrays fabricated from the GSTs show good dynamic range, specificity, and sensitivity in transcript profiling experiments. The GSTs have also been transferred to bacterial plasmid vectors via recombinational cloning protocols. These cloned GSTs constitute the ideal starting point for a variety of functional approaches, including reverse genetics. We have subcloned GSTs on a large scale into vectors designed for gene silencing in plant cells. We show that in planta expression of GST hairpin RNA results in the expected phenotypes in silenced Arabidopsis lines. These versatile GST resources provide novel and powerful tools for functional genomics. PMID:15489341

  2. Comparative analysis of prophages in Streptococcus mutans genomes

    Science.gov (United States)

    Fu, Tiwei; Fan, Xiangyu; Long, Quanxin; Deng, Wanyan; Song, Jinlin

    2017-01-01

    Prophages have been considered genetic units that have an intimate association with novel phenotypic properties of bacterial hosts, such as pathogenicity and genomic variation. Little is known about the genetic information of prophages in the genome of Streptococcus mutans, a major pathogen of human dental caries. In this study, we identified 35 prophage-like elements in S. mutans genomes and performed a comparative genomic analysis. Comparative genomic and phylogenetic analyses of prophage sequences revealed that the prophages could be classified into three main large clusters: Cluster A, Cluster B, and Cluster C. The S. mutans prophages in each cluster were compared. The genomic sequences of phismuN66-1, phismuNLML9-1, and phismu24-1 all shared similarities with the previously reported S. mutans phages M102, M102AD, and ϕAPCM01. The genomes were organized into seven major gene clusters according to the putative functions of the predicted open reading frames: packaging and structural modules, integrase, host lysis modules, DNA replication/recombination modules, transcriptional regulatory modules, other protein modules, and hypothetical protein modules. Moreover, an integrase gene was only identified in phismuNLML9-1 prophages. PMID:29158986

  3. Genome-wide mRNA processing in methanogenic archaea reveals post-transcriptional regulation of ribosomal protein synthesis.

    Science.gov (United States)

    Qi, Lei; Yue, Lei; Feng, Deqin; Qi, Fengxia; Li, Jie; Dong, Xiuzhu

    2017-07-07

    Unlike stable RNAs that require processing for maturation, prokaryotic cellular mRNAs generally follow an 'all-or-none' pattern. Herein, we used a 5΄ monophosphate transcript sequencing (5΄P-seq) that specifically captured the 5΄-end of processed transcripts and mapped the genome-wide RNA processing sites (PSSs) in a methanogenic archaeon. Following statistical analysis and stringent filtration, we identified 1429 PSSs, among which 23.5% and 5.4% were located in 5΄ untranslated region (uPSS) and intergenic region (iPSS), respectively. A predominant uridine downstream PSSs served as a processing signature. Remarkably, 5΄P-seq detected overrepresented uPSS and iPSS in the polycistronic operons encoding ribosomal proteins, and the majority upstream and proximal ribosome binding sites, suggesting a regulatory role of processing on translation initiation. The processed transcripts showed increased stability and translation efficiency. Particularly, processing within the tricistronic transcript of rplA-rplJ-rplL enhanced the translation of rplL, which can provide a driving force for the 1:4 stoichiometry of L10 to L12 in the ribosome. Growth-associated mRNA processing intensities were also correlated with the cellular ribosomal protein levels, thereby suggesting that mRNA processing is involved in tuning growth-dependent ribosome synthesis. In conclusion, our findings suggest that mRNA processing-mediated post-transcriptional regulation is a potential mechanism of ribosomal protein synthesis and stoichiometry. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

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

    Science.gov (United States)

    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.

  5. Genomic analysis of Xenopus organizer function

    Directory of Open Access Journals (Sweden)

    Suhai Sándor

    2006-06-01

    Full Text Available Abstract Background Studies of the Xenopus organizer have laid the foundation for our understanding of the conserved signaling pathways that pattern vertebrate embryos during gastrulation. The two primary activities of the organizer, BMP and Wnt inhibition, can regulate a spectrum of genes that pattern essentially all aspects of the embryo during gastrulation. As our knowledge of organizer signaling grows, it is imperative that we begin knitting together our gene-level knowledge into genome-level signaling models. The goal of this paper was to identify complete lists of genes regulated by different aspects of organizer signaling, thereby providing a deeper understanding of the genomic mechanisms that underlie these complex and fundamental signaling events. Results To this end, we ectopically overexpress Noggin and Dkk-1, inhibitors of the BMP and Wnt pathways, respectively, within ventral tissues. After isolating embryonic ventral halves at early and late gastrulation, we analyze the transcriptional response to these molecules within the generated ectopic organizers using oligonucleotide microarrays. An efficient statistical analysis scheme, combined with a new Gene Ontology biological process annotation of the Xenopus genome, allows reliable and faithful clustering of molecules based upon their roles during gastrulation. From this data, we identify new organizer-related expression patterns for 19 genes. Moreover, our data sub-divides organizer genes into separate head and trunk organizing groups, which each show distinct responses to Noggin and Dkk-1 activity during gastrulation. Conclusion Our data provides a genomic view of the cohorts of genes that respond to Noggin and Dkk-1 activity, allowing us to separate the role of each in organizer function. These patterns demonstrate a model where BMP inhibition plays a largely inductive role during early developmental stages, thereby initiating the suites of genes needed to pattern dorsal tissues

  6. p53 Maintains Genomic Stability by Preventing Interference between Transcription and Replication

    Directory of Open Access Journals (Sweden)

    Constance Qiao Xin Yeo

    2016-04-01

    Full Text Available p53 tumor suppressor maintains genomic stability, typically acting through cell-cycle arrest, senescence, and apoptosis. We discovered a function of p53 in preventing conflicts between transcription and replication, independent of its canonical roles. p53 deficiency sensitizes cells to Topoisomerase (Topo II inhibitors, resulting in DNA damage arising spontaneously during replication. Topoisomerase IIα (TOP2A-DNA complexes preferentially accumulate in isogenic p53 mutant or knockout cells, reflecting an increased recruitment of TOP2A to regulate DNA topology. We propose that p53 acts to prevent DNA topological stress originating from transcription during the S phase and, therefore, promotes normal replication fork progression. Consequently, replication fork progression is impaired in the absence of p53, which is reversed by transcription inhibition. Pharmacologic inhibition of transcription also attenuates DNA damage and decreases Topo-II-DNA complexes, restoring cell viability in p53-deficient cells. Together, our results demonstrate a function of p53 that may underlie its role in tumor suppression.

  7. Genome-wide identification and function analyses of heat shock transcription factors in potato

    Directory of Open Access Journals (Sweden)

    Ruimin eTang

    2016-04-01

    Full Text Available Heat shock transcription factors (Hsfs play vital roles in the regulation of tolerance to various stresses in living organisms. To dissect the mechanisms of the Hsfs in potato adaptation to abiotic stresses, genome and transcriptome analyses of Hsf gene family were investigated in Solanum tuberosum L. Twenty-seven StHsf members were identified by bioinformatics and phylogenetic analyses and were classified into A, B and C groups according to their structural and phylogenetic features. StHsfs in the same class shared similar gene structures and conserved motifs. The chromosomal location analysis showed that 27 Hsfs were located in 10 of 12 chromosomes (except chromosome 1 and chromosome 5 and that 18 of these genes formed 9 paralogous pairs. Expression profiles of StHsfs in 12 different organs and tissues uncovered distinct spatial expression patterns of these genes and their potential roles in the process of growth and development. Promoter and quantitative real-time polymerase chain reaction (qRT-PCR detections of StHsfs were conducted and demonstrated that these genes were all responsive to various stresses. StHsf004, StHsf007, StHsf009, StHsf014 and StHsf019 were constitutively expressed under non-stress conditions, and some specific Hsfs became the predominant Hsfs in response to different abiotic stresses, indicating their important and diverse regulatory roles in adverse conditions. A co-expression network between StHsfs and StHsf-co-expressed genes was generated based on the publicly-available potato transcriptomic databases and identified key candidate StHsfs for further functional studies.

  8. Functional analysis of limb transcriptional enhancers in the mouse.

    Science.gov (United States)

    Nolte, Mark J; Wang, Ying; Deng, Jian Min; Swinton, Paul G; Wei, Caimiao; Guindani, Michele; Schwartz, Robert J; Behringer, Richard R

    2014-01-01

    Transcriptional enhancers are genomic sequences bound by transcription factors that act together with basal transcriptional machinery to regulate gene transcription. Several high-throughput methods have generated large datasets of tissue-specific enhancer sequences with putative roles in developmental processes. However, few enhancers have been deleted from the genome to determine their roles in development. To understand the roles of two enhancers active in the mouse embryonic limb bud we deleted them from the genome. Although the genes regulated by these enhancers are unknown, they were selected because they were identified in a screen for putative limb bud-specific enhancers associated with p300, an acetyltransferase that participates in protein complexes that promote active transcription, and because the orthologous human enhancers (H1442 and H280) drive distinct lacZ expression patterns in limb buds of embryonic day (E) 11.5 transgenic mice. We show that the orthologous mouse sequences, M1442 and M280, regulate dynamic expression in the developing limb. Although significant transcriptional differences in enhancer-proximal genes in embryonic limb buds accompany the deletion of M1442 and M280 no gross limb malformations during embryonic development were observed, demonstrating that M1442 and M280 are not required for mouse limb development. However, M280 is required for the development and/or maintenance of body size; M280 mice are significantly smaller than controls. M280 also harbors an "ultraconserved" sequence that is identical between human, rat, and mouse. This is the first report of a phenotype resulting from the deletion of an ultraconserved element. These studies highlight the importance of determining enhancer regulatory function by experiments that manipulate them in situ and suggest that some of an enhancer's regulatory capacities may be developmentally tolerated rather than developmentally required. © 2014 Wiley Periodicals, Inc.

  9. Rapid and highly efficient construction of TALE-based transcriptional regulators and nucleases for genome modification

    KAUST Repository

    Li, Lixin

    2012-01-22

    Transcription activator-like effectors (TALEs) can be used as DNA-targeting modules by engineering their repeat domains to dictate user-selected sequence specificity. TALEs have been shown to function as site-specific transcriptional activators in a variety of cell types and organisms. TALE nucleases (TALENs), generated by fusing the FokI cleavage domain to TALE, have been used to create genomic double-strand breaks. The identity of the TALE repeat variable di-residues, their number, and their order dictate the DNA sequence specificity. Because TALE repeats are nearly identical, their assembly by cloning or even by synthesis is challenging and time consuming. Here, we report the development and use of a rapid and straightforward approach for the construction of designer TALE (dTALE) activators and nucleases with user-selected DNA target specificity. Using our plasmid set of 100 repeat modules, researchers can assemble repeat domains for any 14-nucleotide target sequence in one sequential restriction-ligation cloning step and in only 24 h. We generated several custom dTALEs and dTALENs with new target sequence specificities and validated their function by transient expression in tobacco leaves and in vitro DNA cleavage assays, respectively. Moreover, we developed a web tool, called idTALE, to facilitate the design of dTALENs and the identification of their genomic targets and potential off-targets in the genomes of several model species. Our dTALE repeat assembly approach along with the web tool idTALE will expedite genome-engineering applications in a variety of cell types and organisms including plants. © 2012 Springer Science+Business Media B.V.

  10. Rapid and highly efficient construction of TALE-based transcriptional regulators and nucleases for genome modification.

    Science.gov (United States)

    Li, Lixin; Piatek, Marek J; Atef, Ahmed; Piatek, Agnieszka; Wibowo, Anjar; Fang, Xiaoyun; Sabir, J S M; Zhu, Jian-Kang; Mahfouz, Magdy M

    2012-03-01

    Transcription activator-like effectors (TALEs) can be used as DNA-targeting modules by engineering their repeat domains to dictate user-selected sequence specificity. TALEs have been shown to function as site-specific transcriptional activators in a variety of cell types and organisms. TALE nucleases (TALENs), generated by fusing the FokI cleavage domain to TALE, have been used to create genomic double-strand breaks. The identity of the TALE repeat variable di-residues, their number, and their order dictate the DNA sequence specificity. Because TALE repeats are nearly identical, their assembly by cloning or even by synthesis is challenging and time consuming. Here, we report the development and use of a rapid and straightforward approach for the construction of designer TALE (dTALE) activators and nucleases with user-selected DNA target specificity. Using our plasmid set of 100 repeat modules, researchers can assemble repeat domains for any 14-nucleotide target sequence in one sequential restriction-ligation cloning step and in only 24 h. We generated several custom dTALEs and dTALENs with new target sequence specificities and validated their function by transient expression in tobacco leaves and in vitro DNA cleavage assays, respectively. Moreover, we developed a web tool, called idTALE, to facilitate the design of dTALENs and the identification of their genomic targets and potential off-targets in the genomes of several model species. Our dTALE repeat assembly approach along with the web tool idTALE will expedite genome-engineering applications in a variety of cell types and organisms including plants.

  11. Prediction of transcriptional regulatory sites in the complete genome sequence of Escherichia coli K-12.

    Science.gov (United States)

    Thieffry, D; Salgado, H; Huerta, A M; Collado-Vides, J

    1998-06-01

    As one of the best-characterized free-living organisms, Escherichia coli and its recently completed genomic sequence offer a special opportunity to exploit systematically the variety of regulatory data available in the literature in order to make a comprehensive set of regulatory predictions in the whole genome. The complete genome sequence of E.coli was analyzed for the binding of transcriptional regulators upstream of coding sequences. The biological information contained in RegulonDB (Huerta, A.M. et al., Nucleic Acids Res.,26,55-60, 1998) for 56 different transcriptional proteins was the support to implement a stringent strategy combining string search and weight matrices. We estimate that our search included representatives of 15-25% of the total number of regulatory binding proteins in E.coli. This search was performed on the set of 4288 putative regulatory regions, each 450 bp long. Within the regions with predicted sites, 89% are regulated by one protein and 81% involve only one site. These numbers are reasonably consistent with the distribution of experimental regulatory sites. Regulatory sites are found in 603 regions corresponding to 16% of operon regions and 10% of intra-operonic regions. Additional evidence gives stronger support to some of these predictions, including the position of the site, biological consistency with the function of the downstream gene, as well as genetic evidence for the regulatory interaction. The predictions described here were incorporated into the map presented in the paper describing the complete E.coli genome (Blattner,F.R. et al., Science, 277, 1453-1461, 1997). The complete set of predictions in GenBank format is available at the url: http://www. cifn.unam.mx/Computational_Biology/E.coli-predictions ecoli-reg@cifn.unam.mx, collado@cifn.unam.mx

  12. Genome-Wide Mapping of Collier In Vivo Binding Sites Highlights Its Hierarchical Position in Different Transcription Regulatory Networks.

    Directory of Open Access Journals (Sweden)

    Mathilde de Taffin

    Full Text Available Collier, the single Drosophila COE (Collier/EBF/Olf-1 transcription factor, is required in several developmental processes, including head patterning and specification of muscle and neuron identity during embryogenesis. To identify direct Collier (Col targets in different cell types, we used ChIP-seq to map Col binding sites throughout the genome, at mid-embryogenesis. In vivo Col binding peaks were associated to 415 potential direct target genes. Gene Ontology analysis revealed a strong enrichment in proteins with DNA binding and/or transcription-regulatory properties. Characterization of a selection of candidates, using transgenic CRM-reporter assays, identified direct Col targets in dorso-lateral somatic muscles and specific neuron types in the central nervous system. These data brought new evidence that Col direct control of the expression of the transcription regulators apterous and eyes-absent (eya is critical to specifying neuronal identities. They also showed that cross-regulation between col and eya in muscle progenitor cells is required for specification of muscle identity, revealing a new parallel between the myogenic regulatory networks operating in Drosophila and vertebrates. Col regulation of eya, both in specific muscle and neuronal lineages, may illustrate one mechanism behind the evolutionary diversification of Col biological roles.

  13. Genome-Wide Mapping of Collier In Vivo Binding Sites Highlights Its Hierarchical Position in Different Transcription Regulatory Networks

    Science.gov (United States)

    Dubois, Laurence; Bataillé, Laetitia; Painset, Anaïs; Le Gras, Stéphanie; Jost, Bernard; Crozatier, Michèle; Vincent, Alain

    2015-01-01

    Collier, the single Drosophila COE (Collier/EBF/Olf-1) transcription factor, is required in several developmental processes, including head patterning and specification of muscle and neuron identity during embryogenesis. To identify direct Collier (Col) targets in different cell types, we used ChIP-seq to map Col binding sites throughout the genome, at mid-embryogenesis. In vivo Col binding peaks were associated to 415 potential direct target genes. Gene Ontology analysis revealed a strong enrichment in proteins with DNA binding and/or transcription-regulatory properties. Characterization of a selection of candidates, using transgenic CRM-reporter assays, identified direct Col targets in dorso-lateral somatic muscles and specific neuron types in the central nervous system. These data brought new evidence that Col direct control of the expression of the transcription regulators apterous and eyes-absent (eya) is critical to specifying neuronal identities. They also showed that cross-regulation between col and eya in muscle progenitor cells is required for specification of muscle identity, revealing a new parallel between the myogenic regulatory networks operating in Drosophila and vertebrates. Col regulation of eya, both in specific muscle and neuronal lineages, may illustrate one mechanism behind the evolutionary diversification of Col biological roles. PMID:26204530

  14. MYB Transcription Factors in Chinese Pear (Pyrus bretschneideri Rehd.: Genome-Wide Identification, Classification and Expression Profiling during Fruit Development

    Directory of Open Access Journals (Sweden)

    Yun Peng eCao

    2016-04-01

    Full Text Available The MYB family is one of the largest families of transcription factors in plants. Although some MYBs have been reported to play roles in secondary metabolism, no comprehensive study of the MYB family in Chinese pear (Pyrus bretschneideri Rehd. has been reported. In the present study, we performed genome-wide analysis of MYB genes in Chinese pear, designated as PbMYBs, including analyses of their phylogenic relationships, structures, chromosomal locations, promoter regions, GO annotations and collinearity. A total of 129 PbMYB genes were identified in the pear genome and were divided into 31 subgroups based on phylogenetic analysis. These PbMYBs were unevenly distributed among 16 chromosomes (total of 17 chromosomes. The occurrence of gene duplication events indicated that whole-genome duplication and segmental duplication likely played key roles in expansion of the PbMYB gene family. Ka/Ks analysis suggested that the duplicated PbMYBs mainly experienced purifying selection with restrictive functional divergence after the duplication events. Interspecies microsynteny analysis revealed maximum orthology between pear and peach, followed by plum and strawberry. Subsequently, the expression patterns of 20 PbMYB genes that may be involved in lignin biosynthesis according to their phylogenetic relationships were examined throughout fruit development. Among the twenty genes examined, PbMYB25 and PbMYB52 exhibited expression patterns consistent with the typical variations in the lignin content previously reported. Moreover, sub-cellular localization analysis revealed that two proteins PbMYB25 and PbMYB52 were localized to the nucleus. All together, PbMYB25 and PbMYB52 were inferred to be candidate genes involved in the regulation of lignin biosynthesis during the development of pear fruit. This study provides useful information for further functional analysis of the MYB gene family in pear.

  15. Virtual Northern analysis of the human genome.

    Directory of Open Access Journals (Sweden)

    Evan H Hurowitz

    2007-05-01

    Full Text Available We applied the Virtual Northern technique to human brain mRNA to systematically measure human mRNA transcript lengths on a genome-wide scale.We used separation by gel electrophoresis followed by hybridization to cDNA microarrays to measure 8,774 mRNA transcript lengths representing at least 6,238 genes at high (>90% confidence. By comparing these transcript lengths to the Refseq and H-Invitational full-length cDNA databases, we found that nearly half of our measurements appeared to represent novel transcript variants. Comparison of length measurements determined by hybridization to different cDNAs derived from the same gene identified clones that potentially correspond to alternative transcript variants. We observed a close linear relationship between ORF and mRNA lengths in human mRNAs, identical in form to the relationship we had previously identified in yeast. Some functional classes of protein are encoded by mRNAs whose untranslated regions (UTRs tend to be longer or shorter than average; these functional classes were similar in both human and yeast.Human transcript diversity is extensive and largely unannotated. Our length dataset can be used as a new criterion for judging the completeness of cDNAs and annotating mRNA sequences. Similar relationships between the lengths of the UTRs in human and yeast mRNAs and the functions of the proteins they encode suggest that UTR sequences serve an important regulatory role among eukaryotes.

  16. Virtual Northern analysis of the human genome.

    Science.gov (United States)

    Hurowitz, Evan H; Drori, Iddo; Stodden, Victoria C; Donoho, David L; Brown, Patrick O

    2007-05-23

    We applied the Virtual Northern technique to human brain mRNA to systematically measure human mRNA transcript lengths on a genome-wide scale. We used separation by gel electrophoresis followed by hybridization to cDNA microarrays to measure 8,774 mRNA transcript lengths representing at least 6,238 genes at high (>90%) confidence. By comparing these transcript lengths to the Refseq and H-Invitational full-length cDNA databases, we found that nearly half of our measurements appeared to represent novel transcript variants. Comparison of length measurements determined by hybridization to different cDNAs derived from the same gene identified clones that potentially correspond to alternative transcript variants. We observed a close linear relationship between ORF and mRNA lengths in human mRNAs, identical in form to the relationship we had previously identified in yeast. Some functional classes of protein are encoded by mRNAs whose untranslated regions (UTRs) tend to be longer or shorter than average; these functional classes were similar in both human and yeast. Human transcript diversity is extensive and largely unannotated. Our length dataset can be used as a new criterion for judging the completeness of cDNAs and annotating mRNA sequences. Similar relationships between the lengths of the UTRs in human and yeast mRNAs and the functions of the proteins they encode suggest that UTR sequences serve an important regulatory role among eukaryotes.

  17. DNA replication factor C1 mediates genomic stability and transcriptional gene silencing in Arabidopsis

    KAUST Repository

    Liu, Qian; Wang, Junguo; Miki, Daisuke; Xia, Ran; Yu, Wenxiang; He, Junna; Zheng, Zhimin; Zhu, Jian-Kang; Gonga, Zhizhong

    2010-01-01

    Genetic screening identified a suppressor of ros1-1, a mutant of REPRESSOR OF SILENCING1 (ROS1; encoding a DNA demethylation protein). The suppressor is a mutation in the gene encoding the largest subunit of replication factor C (RFC1). This mutation of RFC1 reactivates the unlinked 35S-NPTII transgene, which is silenced in ros1 and also increases expression of the pericentromeric Athila retrotransposons named transcriptional silent information in a DNA methylationindependent manner. rfc1 is more sensitive than the wild type to the DNA-damaging agent methylmethane sulphonate and to the DNA inter- and intra- cross-linking agent cisplatin. The rfc1 mutant constitutively expresses the G2/M-specific cyclin CycB1;1 and other DNA repair-related genes. Treatment with DNA-damaging agents mimics the rfc1 mutation in releasing the silenced 35S-NPTII, suggesting that spontaneously induced genomic instability caused by the rfc1 mutation might partially contribute to the released transcriptional gene silencing (TGS). The frequency of somatic homologous recombination is significantly increased in the rfc1 mutant. Interestingly, ros1 mutants show increased telomere length, but rfc1 mutants show decreased telomere length and reduced expression of telomerase. Our results suggest that RFC1 helps mediate genomic stability and TGS in Arabidopsis thaliana. © 2010 American Society of Plant Biologists.

  18. DNA replication factor C1 mediates genomic stability and transcriptional gene silencing in Arabidopsis

    KAUST Repository

    Liu, Qian

    2010-07-01

    Genetic screening identified a suppressor of ros1-1, a mutant of REPRESSOR OF SILENCING1 (ROS1; encoding a DNA demethylation protein). The suppressor is a mutation in the gene encoding the largest subunit of replication factor C (RFC1). This mutation of RFC1 reactivates the unlinked 35S-NPTII transgene, which is silenced in ros1 and also increases expression of the pericentromeric Athila retrotransposons named transcriptional silent information in a DNA methylationindependent manner. rfc1 is more sensitive than the wild type to the DNA-damaging agent methylmethane sulphonate and to the DNA inter- and intra- cross-linking agent cisplatin. The rfc1 mutant constitutively expresses the G2/M-specific cyclin CycB1;1 and other DNA repair-related genes. Treatment with DNA-damaging agents mimics the rfc1 mutation in releasing the silenced 35S-NPTII, suggesting that spontaneously induced genomic instability caused by the rfc1 mutation might partially contribute to the released transcriptional gene silencing (TGS). The frequency of somatic homologous recombination is significantly increased in the rfc1 mutant. Interestingly, ros1 mutants show increased telomere length, but rfc1 mutants show decreased telomere length and reduced expression of telomerase. Our results suggest that RFC1 helps mediate genomic stability and TGS in Arabidopsis thaliana. © 2010 American Society of Plant Biologists.

  19. Genome-scale transcriptional activation by an engineered CRISPR-Cas9 complex.

    Science.gov (United States)

    Konermann, Silvana; Brigham, Mark D; Trevino, Alexandro E; Joung, Julia; Abudayyeh, Omar O; Barcena, Clea; Hsu, Patrick D; Habib, Naomi; Gootenberg, Jonathan S; Nishimasu, Hiroshi; Nureki, Osamu; Zhang, Feng

    2015-01-29

    Systematic interrogation of gene function requires the ability to perturb gene expression in a robust and generalizable manner. Here we describe structure-guided engineering of a CRISPR-Cas9 complex to mediate efficient transcriptional activation at endogenous genomic loci. We used these engineered Cas9 activation complexes to investigate single-guide RNA (sgRNA) targeting rules for effective transcriptional activation, to demonstrate multiplexed activation of ten genes simultaneously, and to upregulate long intergenic non-coding RNA (lincRNA) transcripts. We also synthesized a library consisting of 70,290 guides targeting all human RefSeq coding isoforms to screen for genes that, upon activation, confer resistance to a BRAF inhibitor. The top hits included genes previously shown to be able to confer resistance, and novel candidates were validated using individual sgRNA and complementary DNA overexpression. A gene expression signature based on the top screening hits correlated with markers of BRAF inhibitor resistance in cell lines and patient-derived samples. These results collectively demonstrate the potential of Cas9-based activators as a powerful genetic perturbation technology.

  20. Analysis of Single-cell Gene Transcription by RNA Fluorescent In Situ Hybridization (FISH)

    DEFF Research Database (Denmark)

    Ronander, Elena; Bengtsson, Dominique C; Joergensen, Louise

    2012-01-01

    Adhesion of Plasmodium falciparum infected erythrocytes (IE) to human endothelial receptors during malaria infections is mediated by expression of PfEMP1 protein variants encoded by the var genes. The haploid P. falciparum genome harbors approximately 60 different var genes of which only one has...... been believed to be transcribed per cell at a time during the blood stage of the infection. How such mutually exclusive regulation of var gene transcription is achieved is unclear, as is the identification of individual var genes or sub-groups of var genes associated with different receptors...... fluorescent in situ hybridization (FISH) analysis of var gene transcription by the parasite in individual nuclei of P. falciparum IE(1). Here, we present a detailed protocol for carrying out the RNA-FISH methodology for analysis of var gene transcription in single-nuclei of P. falciparum infected human...

  1. Comparative Genomics of NAC Transcriptional Factors in Angiosperms: Implications for the Adaptation and Diversification of Flowering Plants

    Science.gov (United States)

    Pereira-Santana, Alejandro; Alcaraz, Luis David; Castaño, Enrique; Sanchez-Calderon, Lenin; Sanchez-Teyer, Felipe; Rodriguez-Zapata, Luis

    2015-01-01

    NAC proteins constitute one of the largest groups of plant-specific transcription factors and are known to play essential roles in various developmental processes. They are also important in plant responses to stresses such as drought, soil salinity, cold, and heat, which adversely affect growth. The current knowledge regarding the distribution of NAC proteins in plant lineages comes from relatively small samplings from the available data. In the present study, we broadened the number of plant species containing the NAC family origin and evolution to shed new light on the evolutionary history of this family in angiosperms. A comparative genome analysis was performed on 24 land plant species, and NAC ortholog groups were identified by means of bidirectional BLAST hits. Large NAC gene families are found in those species that have experienced more whole-genome duplication events, pointing to an expansion of the NAC family with divergent functions in flowering plants. A total of 3,187 NAC transcription factors that clustered into six major groups were used in the phylogenetic analysis. Many orthologous groups were found in the monocot and eudicot lineages, but only five orthologous groups were found between P. patens and each representative taxa of flowering plants. These groups were called basal orthologous groups and likely expanded into more recent taxa to cope with their environmental needs. This analysis on the angiosperm NAC family represents an effort to grasp the evolutionary and functional diversity within this gene family while providing a basis for further functional research on vascular plant gene families. PMID:26569117

  2. Comparative Genomics of NAC Transcriptional Factors in Angiosperms: Implications for the Adaptation and Diversification of Flowering Plants.

    Directory of Open Access Journals (Sweden)

    Alejandro Pereira-Santana

    Full Text Available NAC proteins constitute one of the largest groups of plant-specific transcription factors and are known to play essential roles in various developmental processes. They are also important in plant responses to stresses such as drought, soil salinity, cold, and heat, which adversely affect growth. The current knowledge regarding the distribution of NAC proteins in plant lineages comes from relatively small samplings from the available data. In the present study, we broadened the number of plant species containing the NAC family origin and evolution to shed new light on the evolutionary history of this family in angiosperms. A comparative genome analysis was performed on 24 land plant species, and NAC ortholog groups were identified by means of bidirectional BLAST hits. Large NAC gene families are found in those species that have experienced more whole-genome duplication events, pointing to an expansion of the NAC family with divergent functions in flowering plants. A total of 3,187 NAC transcription factors that clustered into six major groups were used in the phylogenetic analysis. Many orthologous groups were found in the monocot and eudicot lineages, but only five orthologous groups were found between P. patens and each representative taxa of flowering plants. These groups were called basal orthologous groups and likely expanded into more recent taxa to cope with their environmental needs. This analysis on the angiosperm NAC family represents an effort to grasp the evolutionary and functional diversity within this gene family while providing a basis for further functional research on vascular plant gene families.

  3. Transcription Activator-Like Effectors (TALEs) Hybrid Nucleases for Genome Engineering Application

    KAUST Repository

    Wibowo, Anjar

    2011-06-06

    Gene targeting is a powerful genome engineering tool that can be used for a variety of biotechnological applications. Genomic double-strand DNA breaks generated by engineered site-specific nucleases can stimulate gene targeting. Hybrid nucleases are composed of DNA binding module and DNA cleavage module. Zinc Finger Nucleases were used to generate double-strand DNA breaks but it suffers from failures and lack of reproducibility. The transcription activator–like effectors (TALEs) from plant pathogenic Xanthomonas contain a unique type of DNA-binding domain that bind specific DNA targets. The purpose of this study is to generate novel sequence specific nucleases by fusing a de novo engineered Hax3 TALE-based DNA binding domain to a FokI cleavage domain. Our data show that the de novo engineered TALE nuclease can bind to its target sequence and create double-strand DNA breaks in vitro. We also show that the de novo engineered TALE nuclease is capable of generating double-strand DNA breaks in its target sequence in vivo, when transiently expressed in Nicotiana benthamiana leaves. In conclusion, our data demonstrate that TALE-based hybrid nucleases can be tailored to bind a user-selected DNA sequence and generate site-specific genomic double-strand DNA breaks. TALE-based hybrid nucleases hold much promise as powerful molecular tools for gene targeting applications.

  4. Identification of transcriptional signals in Encephalitozoon cuniculi widespread among Microsporidia phylum: support for accurate structural genome annotation

    Directory of Open Access Journals (Sweden)

    Wincker Patrick

    2009-12-01

    Full Text Available Abstract Background Microsporidia are obligate intracellular eukaryotic parasites with genomes ranging in size from 2.3 Mbp to more than 20 Mbp. The extremely small (2.9 Mbp and highly compact (~1 gene/kb genome of the human parasite Encephalitozoon cuniculi has been fully sequenced. The aim of this study was to characterize noncoding motifs that could be involved in regulation of gene expression in E. cuniculi and to show whether these motifs are conserved among the phylum Microsporidia. Results To identify such signals, 5' and 3'RACE-PCR experiments were performed on different E. cuniculi mRNAs. This analysis confirmed that transcription overrun occurs in E. cuniculi and may result from stochastic recognition of the AAUAAA polyadenylation signal. Such experiments also showed highly reduced 5'UTR's (E. cuniculi genes presented a CCC-like motif immediately upstream from the coding start. To characterize other signals involved in differential transcriptional regulation, we then focused our attention on the gene family coding for ribosomal proteins. An AAATTT-like signal was identified upstream from the CCC-like motif. In rare cases the cytosine triplet was shown to be substituted by a GGG-like motif. Comparative genomic studies confirmed that these different signals are also located upstream from genes encoding ribosomal proteins in other microsporidian species including Antonospora locustae, Enterocytozoon bieneusi, Anncaliia algerae (syn. Brachiola algerae and Nosema ceranae. Based on these results a systematic analysis of the ~2000 E. cuniculi coding DNA sequences was then performed and brings to highlight that 364 translation initiation codons (18.29% of total CDSs had been badly predicted. Conclusion We identified various signals involved in the maturation of E. cuniculi mRNAs. Presence of such signals, in phylogenetically distant microsporidian species, suggests that a common regulatory mechanism exists among the microsporidia. Furthermore

  5. Identification of conserved regulatory elements by comparative genome analysis

    Directory of Open Access Journals (Sweden)

    Jareborg Niclas

    2003-05-01

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

  6. The future of genome-scale modeling of yeast through integration of a transcriptional regulatory network

    DEFF Research Database (Denmark)

    Liu, Guodong; Marras, Antonio; Nielsen, Jens

    2014-01-01

    regulatory information is necessary to improve the accuracy and predictive ability of metabolic models. Here we review the strategies for the reconstruction of a transcriptional regulatory network (TRN) for yeast and the integration of such a reconstruction into a flux balance analysis-based metabolic model......Metabolism is regulated at multiple levels in response to the changes of internal or external conditions. Transcriptional regulation plays an important role in regulating many metabolic reactions by altering the concentrations of metabolic enzymes. Thus, integration of the transcriptional....... While many large-scale TRN reconstructions have been reported for yeast, these reconstructions still need to be improved regarding the functionality and dynamic property of the regulatory interactions. In addition, mathematical modeling approaches need to be further developed to efficiently integrate...

  7. Comprehensive genome-wide survey, genomic constitution and expression profiling of the NAC transcription factor family in foxtail millet (Setaria italica L..

    Directory of Open Access Journals (Sweden)

    Swati Puranik

    Full Text Available The NAC proteins represent a major plant-specific transcription factor family that has established enormously diverse roles in various plant processes. Aided by the availability of complete genomes, several members of this family have been identified in Arabidopsis, rice, soybean and poplar. However, no comprehensive investigation has been presented for the recently sequenced, naturally stress tolerant crop, Setaria italica (foxtail millet that is famed as a model crop for bioenergy research. In this study, we identified 147 putative NAC domain-encoding genes from foxtail millet by systematic sequence analysis and physically mapped them onto nine chromosomes. Genomic organization suggested that inter-chromosomal duplications may have been responsible for expansion of this gene family in foxtail millet. Phylogenetically, they were arranged into 11 distinct sub-families (I-XI, with duplicated genes fitting into one cluster and possessing conserved motif compositions. Comparative mapping with other grass species revealed some orthologous relationships and chromosomal rearrangements including duplication, inversion and deletion of genes. The evolutionary significance as duplication and divergence of NAC genes based on their amino acid substitution rates was understood. Expression profiling against various stresses and phytohormones provides novel insights into specific and/or overlapping expression patterns of SiNAC genes, which may be responsible for functional divergence among individual members in this crop. Further, we performed structure modeling and molecular simulation of a stress-responsive protein, SiNAC128, proffering an initial framework for understanding its molecular function. Taken together, this genome-wide identification and expression profiling unlocks new avenues for systematic functional analysis of novel NAC gene family candidates which may be applied for improvising stress adaption in plants.

  8. Comprehensive genome-wide survey, genomic constitution and expression profiling of the NAC transcription factor family in foxtail millet (Setaria italica L.).

    Science.gov (United States)

    Puranik, Swati; Sahu, Pranav Pankaj; Mandal, Sambhu Nath; B, Venkata Suresh; Parida, Swarup Kumar; Prasad, Manoj

    2013-01-01

    The NAC proteins represent a major plant-specific transcription factor family that has established enormously diverse roles in various plant processes. Aided by the availability of complete genomes, several members of this family have been identified in Arabidopsis, rice, soybean and poplar. However, no comprehensive investigation has been presented for the recently sequenced, naturally stress tolerant crop, Setaria italica (foxtail millet) that is famed as a model crop for bioenergy research. In this study, we identified 147 putative NAC domain-encoding genes from foxtail millet by systematic sequence analysis and physically mapped them onto nine chromosomes. Genomic organization suggested that inter-chromosomal duplications may have been responsible for expansion of this gene family in foxtail millet. Phylogenetically, they were arranged into 11 distinct sub-families (I-XI), with duplicated genes fitting into one cluster and possessing conserved motif compositions. Comparative mapping with other grass species revealed some orthologous relationships and chromosomal rearrangements including duplication, inversion and deletion of genes. The evolutionary significance as duplication and divergence of NAC genes based on their amino acid substitution rates was understood. Expression profiling against various stresses and phytohormones provides novel insights into specific and/or overlapping expression patterns of SiNAC genes, which may be responsible for functional divergence among individual members in this crop. Further, we performed structure modeling and molecular simulation of a stress-responsive protein, SiNAC128, proffering an initial framework for understanding its molecular function. Taken together, this genome-wide identification and expression profiling unlocks new avenues for systematic functional analysis of novel NAC gene family candidates which may be applied for improvising stress adaption in plants.

  9. Genome-wide mapping of transcription start sites yields novel insights into the primary transcriptome of Pseudomonas putida

    DEFF Research Database (Denmark)

    D'Arrigo, Isotta; Bojanovic, Klara; Yang, Xiaochen

    2016-01-01

    was examined using an in vivo assay with GFP-fusion vectors and shown to function via a translational repression mechanism. Furthermore, 56 novel intergenic small RNAs and 8 putative actuaton transcripts were detected, as well as 8 novel open reading frames (ORFs). This study illustrates how global mapping...... of TSSs can yield novel insights into the transcriptional features and RNA output of bacterial genomes....

  10. A community resource for high-throughput quantitative RT-PCR analysis of transcription factor gene expression in Medicago truncatula

    Directory of Open Access Journals (Sweden)

    Redman Julia C

    2008-07-01

    Full Text Available Abstract Background Medicago truncatula is a model legume species that is currently the focus of an international genome sequencing effort. Although several different oligonucleotide and cDNA arrays have been produced for genome-wide transcript analysis of this species, intrinsic limitations in the sensitivity of hybridization-based technologies mean that transcripts of genes expressed at low-levels cannot be measured accurately with these tools. Amongst such genes are many encoding transcription factors (TFs, which are arguably the most important class of regulatory proteins. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR is the most sensitive method currently available for transcript quantification, and one that can be scaled up to analyze transcripts of thousands of genes in parallel. Thus, qRT-PCR is an ideal method to tackle the problem of TF transcript quantification in Medicago and other plants. Results We established a bioinformatics pipeline to identify putative TF genes in Medicago truncatula and to design gene-specific oligonucleotide primers for qRT-PCR analysis of TF transcripts. We validated the efficacy and gene-specificity of over 1000 TF primer pairs and utilized these to identify sets of organ-enhanced TF genes that may play important roles in organ development or differentiation in this species. This community resource will be developed further as more genome sequence becomes available, with the ultimate goal of producing validated, gene-specific primers for all Medicago TF genes. Conclusion High-throughput qRT-PCR using a 384-well plate format enables rapid, flexible, and sensitive quantification of all predicted Medicago transcription factor mRNAs. This resource has been utilized recently by several groups in Europe, Australia, and the USA, and we expect that it will become the 'gold-standard' for TF transcript profiling in Medicago truncatula.

  11. Rewiring the severe acute respiratory syndrome coronavirus (SARS-CoV) transcription circuit: Engineering a recombination-resistant genome

    Science.gov (United States)

    Yount, Boyd; Roberts, Rhonda S.; Lindesmith, Lisa; Baric, Ralph S.

    2006-08-01

    Live virus vaccines provide significant protection against many detrimental human and animal diseases, but reversion to virulence by mutation and recombination has reduced appeal. Using severe acute respiratory syndrome coronavirus as a model, we engineered a different transcription regulatory circuit and isolated recombinant viruses. The transcription network allowed for efficient expression of the viral transcripts and proteins, and the recombinant viruses replicated to WT levels. Recombinant genomes were then constructed that contained mixtures of the WT and mutant regulatory circuits, reflecting recombinant viruses that might occur in nature. Although viable viruses could readily be isolated from WT and recombinant genomes containing homogeneous transcription circuits, chimeras that contained mixed regulatory networks were invariantly lethal, because viable chimeric viruses were not isolated. Mechanistically, mixed regulatory circuits promoted inefficient subgenomic transcription from inappropriate start sites, resulting in truncated ORFs and effectively minimize viral structural protein expression. Engineering regulatory transcription circuits of intercommunicating alleles successfully introduces genetic traps into a viral genome that are lethal in RNA recombinant progeny viruses. regulation | systems biology | vaccine design

  12. Genomic androgen receptor-occupied regions with different functions, defined by histone acetylation, coregulators and transcriptional capacity.

    Directory of Open Access Journals (Sweden)

    Li Jia

    Full Text Available The androgen receptor (AR is a steroid-activated transcription factor that binds at specific DNA locations and plays a key role in the etiology of prostate cancer. While numerous studies have identified a clear connection between AR binding and expression of target genes for a limited number of loci, high-throughput elucidation of these sites allows for a deeper understanding of the complexities of this process.We have mapped 189 AR occupied regions (ARORs and 1,388 histone H3 acetylation (AcH3 loci to a 3% continuous stretch of human genomic DNA using chromatin immunoprecipitation (ChIP microarray analysis. Of 62 highly reproducible ARORs, 32 (52% were also marked by AcH3. While the number of ARORs detected in prostate cancer cells exceeded the number of nearby DHT-responsive genes, the AcH3 mark defined a subclass of ARORs much more highly associated with such genes -- 12% of the genes flanking AcH3+ARORs were DHT-responsive, compared to only 1% of genes flanking AcH3-ARORs. Most ARORs contained enhancer activities as detected in luciferase reporter assays. Analysis of the AROR sequences, followed by site-directed ChIP, identified binding sites for AR transcriptional coregulators FoxA1, CEBPbeta, NFI and GATA2, which had diverse effects on endogenous AR target gene expression levels in siRNA knockout experiments.We suggest that only some ARORs function under the given physiological conditions, utilizing diverse mechanisms. This diversity points to differential regulation of gene expression by the same transcription factor related to the chromatin structure.

  13. Genomic binding profiles of functionally distinct RNA polymerase III transcription complexes in human cells.

    Science.gov (United States)

    Moqtaderi, Zarmik; Wang, Jie; Raha, Debasish; White, Robert J; Snyder, Michael; Weng, Zhiping; Struhl, Kevin

    2010-05-01

    Genome-wide occupancy profiles of five components of the RNA polymerase III (Pol III) machinery in human cells identified the expected tRNA and noncoding RNA targets and revealed many additional Pol III-associated loci, mostly near short interspersed elements (SINEs). Several genes are targets of an alternative transcription factor IIIB (TFIIIB) containing Brf2 instead of Brf1 and have extremely low levels of TFIIIC. Strikingly, expressed Pol III genes, unlike nonexpressed Pol III genes, are situated in regions with a pattern of histone modifications associated with functional Pol II promoters. TFIIIC alone associates with numerous ETC loci, via the B box or a novel motif. ETCs are often near CTCF binding sites, suggesting a potential role in chromosome organization. Our results suggest that human Pol III complexes associate preferentially with regions near functional Pol II promoters and that TFIIIC-mediated recruitment of TFIIIB is regulated in a locus-specific manner.

  14. Transcriptional and Posttranslational Regulation of Nucleotide Excision Repair: The Guardian of the Genome against Ultraviolet Radiation

    Directory of Open Access Journals (Sweden)

    Jeong-Min Park

    2016-11-01

    Full Text Available Ultraviolet (UV radiation from sunlight represents a constant threat to genome stability by generating modified DNA bases such as cyclobutane pyrimidine dimers (CPD and pyrimidine-pyrimidone (6-4 photoproducts (6-4PP. If unrepaired, these lesions can have deleterious effects, including skin cancer. Mammalian cells are able to neutralize UV-induced photolesions through nucleotide excision repair (NER. The NER pathway has multiple components including seven xeroderma pigmentosum (XP proteins (XPA to XPG and numerous auxiliary factors, including ataxia telangiectasia and Rad3-related (ATR protein kinase and RCC1 like domain (RLD and homologous to the E6-AP carboxyl terminus (HECT domain containing E3 ubiquitin protein ligase 2 (HERC2. In this review we highlight recent data on the transcriptional and posttranslational regulation of NER activity.

  15. Bacterial Genome Editing Strategy for Control of Transcription and Protein Stability

    DEFF Research Database (Denmark)

    Lauritsen, Ida; Martinez, Virginia; Ronda, Carlotta

    2018-01-01

    In molecular biology and cell factory engineering, tools that enable control of protein production and stability are highly important. Here, we describe protocols for tagging genes in Escherichia coli allowing for inducible degradation and transcriptional control of any soluble protein of interest....... The underlying molecular biology is based on the two cross-kingdom tools CRISPRi and the N-end rule for protein degradation. Genome editing is performed with the CRMAGE technology and randomization of the translational initiation region minimizes the polar effects of tag insertion. The approach has previously...... been applied for targeting proteins originating from essential operon-located genes and has potential to serve as a universal synthetic biology tool....

  16. Genome Sequencing and Analysis Conference IV

    Energy Technology Data Exchange (ETDEWEB)

    1993-12-31

    J. Craig Venter and C. Thomas Caskey co-chaired Genome Sequencing and Analysis Conference IV held at Hilton Head, South Carolina from September 26--30, 1992. Venter opened the conference by noting that approximately 400 researchers from 16 nations were present four times as many participants as at Genome Sequencing Conference I in 1989. Venter also introduced the Data Fair, a new component of the conference allowing exchange and on-site computer analysis of unpublished sequence data.

  17. Genome-wide organization and expression profiling of the NAC transcription factor family in potato (Solanum tuberosum L.).

    Science.gov (United States)

    Singh, Anil Kumar; Sharma, Vishal; Pal, Awadhesh Kumar; Acharya, Vishal; Ahuja, Paramvir Singh

    2013-08-01

    NAC [no apical meristem (NAM), Arabidopsis thaliana transcription activation factor [ATAF1/2] and cup-shaped cotyledon (CUC2)] proteins belong to one of the largest plant-specific transcription factor (TF) families and play important roles in plant development processes, response to biotic and abiotic cues and hormone signalling. Our genome-wide analysis identified 110 StNAC genes in potato encoding for 136 proteins, including 14 membrane-bound TFs. The physical map positions of StNAC genes on 12 potato chromosomes were non-random, and 40 genes were found to be distributed in 16 clusters. The StNAC proteins were phylogenetically clustered into 12 subgroups. Phylogenetic analysis of StNACs along with their Arabidopsis and rice counterparts divided these proteins into 18 subgroups. Our comparative analysis has also identified 36 putative TNAC proteins, which appear to be restricted to Solanaceae family. In silico expression analysis, using Illumina RNA-seq transcriptome data, revealed tissue-specific, biotic, abiotic stress and hormone-responsive expression profile of StNAC genes. Several StNAC genes, including StNAC072 and StNAC101that are orthologs of known stress-responsive Arabidopsis RESPONSIVE TO DEHYDRATION 26 (RD26) were identified as highly abiotic stress responsive. Quantitative real-time polymerase chain reaction analysis largely corroborated the expression profile of StNAC genes as revealed by the RNA-seq data. Taken together, this analysis indicates towards putative functions of several StNAC TFs, which will provide blue-print for their functional characterization and utilization in potato improvement.

  18. Big Data Analysis of Human Genome Variations

    KAUST Repository

    Gojobori, Takashi

    2016-01-25

    Since the human genome draft sequence was in public for the first time in 2000, genomic analyses have been intensively extended to the population level. The following three international projects are good examples for large-scale studies of human genome variations: 1) HapMap Data (1,417 individuals) (http://hapmap.ncbi.nlm.nih.gov/downloads/genotypes/2010-08_phaseII+III/forward/), 2) HGDP (Human Genome Diversity Project) Data (940 individuals) (http://www.hagsc.org/hgdp/files.html), 3) 1000 genomes Data (2,504 individuals) http://ftp.1000genomes.ebi.ac.uk/vol1/ftp/release/20130502/ If we can integrate all three data into a single volume of data, we should be able to conduct a more detailed analysis of human genome variations for a total number of 4,861 individuals (= 1,417+940+2,504 individuals). In fact, we successfully integrated these three data sets by use of information on the reference human genome sequence, and we conducted the big data analysis. In particular, we constructed a phylogenetic tree of about 5,000 human individuals at the genome level. As a result, we were able to identify clusters of ethnic groups, with detectable admixture, that were not possible by an analysis of each of the three data sets. Here, we report the outcome of this kind of big data analyses and discuss evolutionary significance of human genomic variations. Note that the present study was conducted in collaboration with Katsuhiko Mineta and Kosuke Goto at KAUST.

  19. Comparative Genome Analysis of Enterobacter cloacae

    Science.gov (United States)

    Liu, Wing-Yee; Wong, Chi-Fat; Chung, Karl Ming-Kar; Jiang, Jing-Wei; Leung, Frederick Chi-Ching

    2013-01-01

    The Enterobacter cloacae species includes an extremely diverse group of bacteria that are associated with plants, soil and humans. Publication of the complete genome sequence of the plant growth-promoting endophytic E. cloacae subsp. cloacae ENHKU01 provided an opportunity to perform the first comparative genome analysis between strains of this dynamic species. Examination of the pan-genome of E. cloacae showed that the conserved core genome retains the general physiological and survival genes of the species, while genomic factors in plasmids and variable regions determine the virulence of the human pathogenic E. cloacae strain; additionally, the diversity of fimbriae contributes to variation in colonization and host determination of different E. cloacae strains. Comparative genome analysis further illustrated that E. cloacae strains possess multiple mechanisms for antagonistic action against other microorganisms, which involve the production of siderophores and various antimicrobial compounds, such as bacteriocins, chitinases and antibiotic resistance proteins. The presence of Type VI secretion systems is expected to provide further fitness advantages for E. cloacae in microbial competition, thus allowing it to survive in different environments. Competition assays were performed to support our observations in genomic analysis, where E. cloacae subsp. cloacae ENHKU01 demonstrated antagonistic activities against a wide range of plant pathogenic fungal and bacterial species. PMID:24069314

  20. Microbial genome analysis: the COG approach.

    Science.gov (United States)

    Galperin, Michael Y; Kristensen, David M; Makarova, Kira S; Wolf, Yuri I; Koonin, Eugene V

    2017-09-14

    For the past 20 years, the Clusters of Orthologous Genes (COG) database had been a popular tool for microbial genome annotation and comparative genomics. Initially created for the purpose of evolutionary classification of protein families, the COG have been used, apart from straightforward functional annotation of sequenced genomes, for such tasks as (i) unification of genome annotation in groups of related organisms; (ii) identification of missing and/or undetected genes in complete microbial genomes; (iii) analysis of genomic neighborhoods, in many cases allowing prediction of novel functional systems; (iv) analysis of metabolic pathways and prediction of alternative forms of enzymes; (v) comparison of organisms by COG functional categories; and (vi) prioritization of targets for structural and functional characterization. Here we review the principles of the COG approach and discuss its key advantages and drawbacks in microbial genome analysis. Published by Oxford University Press 2017. This work is written by US Government employees and is in the public domain in the US.

  1. Comparing genomic expression patterns across plant species reveals highly diverged transcriptional dynamics in response to salt stress

    Directory of Open Access Journals (Sweden)

    Close Timothy J

    2009-08-01

    Full Text Available Abstract Background Rice and barley are both members of Poaceae (grass family but have a marked difference in salt tolerance. The molecular mechanism underlying this difference was previously unexplored. This study employs a comparative genomics approach to identify analogous and contrasting gene expression patterns between rice and barley. Results A hierarchical clustering approach identified several interesting expression trajectories among rice and barley genotypes. There were no major conserved expression patterns between the two species in response to salt stress. A wheat salt-stress dataset was queried for comparison with rice and barley. Roughly one-third of the salt-stress responses of barley were conserved with wheat while overlap between wheat and rice was minimal. These results demonstrate that, at transcriptome level, rice is strikingly different compared to the more closely related barley and wheat. This apparent lack of analogous transcriptional programs in response to salt stress is further highlighted through close examination of genes associated with root growth and development. Conclusion The analysis provides support for the hypothesis that conservation of transcriptional signatures in response to environmental cues depends on the genetic similarity among the genotypes within a species, and on the phylogenetic distance between the species.

  2. PRISM offers a comprehensive genomic approach to transcription factor function prediction

    KAUST Repository

    Wenger, A. M.; Clarke, S. L.; Guturu, H.; Chen, J.; Schaar, B. T.; McLean, C. Y.; Bejerano, G.

    2013-01-01

    The human genome encodes 1500-2000 different transcription factors (TFs). ChIP-seq is revealing the global binding profiles of a fraction of TFs in a fraction of their biological contexts. These data show that the majority of TFs bind directly next to a large number of context-relevant target genes, that most binding is distal, and that binding is context specific. Because of the effort and cost involved, ChIP-seq is seldom used in search of novel TF function. Such exploration is instead done using expression perturbation and genetic screens. Here we propose a comprehensive computational framework for transcription factor function prediction. We curate 332 high-quality nonredundant TF binding motifs that represent all major DNA binding domains, and improve cross-species conserved binding site prediction to obtain 3.3 million conserved, mostly distal, binding site predictions. We combine these with 2.4 million facts about all human and mouse gene functions, in a novel statistical framework, in search of enrichments of particular motifs next to groups of target genes of particular functions. Rigorous parameter tuning and a harsh null are used to minimize false positives. Our novel PRISM (predicting regulatory information from single motifs) approach obtains 2543 TF function predictions in a large variety of contexts, at a false discovery rate of 16%. The predictions are highly enriched for validated TF roles, and 45 of 67 (67%) tested binding site regions in five different contexts act as enhancers in functionally matched cells.

  3. PRISM offers a comprehensive genomic approach to transcription factor function prediction

    KAUST Repository

    Wenger, A. M.

    2013-02-04

    The human genome encodes 1500-2000 different transcription factors (TFs). ChIP-seq is revealing the global binding profiles of a fraction of TFs in a fraction of their biological contexts. These data show that the majority of TFs bind directly next to a large number of context-relevant target genes, that most binding is distal, and that binding is context specific. Because of the effort and cost involved, ChIP-seq is seldom used in search of novel TF function. Such exploration is instead done using expression perturbation and genetic screens. Here we propose a comprehensive computational framework for transcription factor function prediction. We curate 332 high-quality nonredundant TF binding motifs that represent all major DNA binding domains, and improve cross-species conserved binding site prediction to obtain 3.3 million conserved, mostly distal, binding site predictions. We combine these with 2.4 million facts about all human and mouse gene functions, in a novel statistical framework, in search of enrichments of particular motifs next to groups of target genes of particular functions. Rigorous parameter tuning and a harsh null are used to minimize false positives. Our novel PRISM (predicting regulatory information from single motifs) approach obtains 2543 TF function predictions in a large variety of contexts, at a false discovery rate of 16%. The predictions are highly enriched for validated TF roles, and 45 of 67 (67%) tested binding site regions in five different contexts act as enhancers in functionally matched cells.

  4. Novel Genomic and Evolutionary Insight of WRKY Transcription Factors in Plant Lineage.

    Science.gov (United States)

    Mohanta, Tapan Kumar; Park, Yong-Hwan; Bae, Hanhong

    2016-11-17

    The evolutionarily conserved WRKY transcription factor (TF) regulates different aspects of gene expression in plants, and modulates growth, development, as well as biotic and abiotic stress responses. Therefore, understanding the details regarding WRKY TFs is very important. In this study, large-scale genomic analyses of the WRKY TF gene family from 43 plant species were conducted. The results of our study revealed that WRKY TFs could be grouped and specifically classified as those belonging to the monocot or dicot plant lineage. In this study, we identified several novel WRKY TFs. To our knowledge, this is the first report on a revised grouping system of the WRKY TF gene family in plants. The different forms of novel chimeric forms of WRKY TFs in the plant genome might play a crucial role in their evolution. Tissue-specific gene expression analyses in Glycine max and Phaseolus vulgaris showed that WRKY11-1, WRKY11-2 and WRKY11-3 were ubiquitously expressed in all tissue types, and WRKY15-2 was highly expressed in the stem, root, nodule and pod tissues in G. max and P. vulgaris.

  5. Comparative Genomic and Transcriptional Analyses of CRISPR Systems Across the Genus Pyrobaculum

    Directory of Open Access Journals (Sweden)

    David L Bernick

    2012-07-01

    Full Text Available Within the domain Archaea, the CRISPR immune system appears to be nearly ubiquitous based on computational genome analyses. Initial studies in bacteria demonstrated that the CRISPR system targets invading plasmid and viral DNA. Recent experiments in the model archaeon Pyrococcus furiosus uncovered a novel RNA-targeting variant of the CRISPR system potentially unique to archaea. Because our understanding of CRISPR system evolution in other archaea is limited, we have taken a comparative genomic and transcriptomic view of the CRISPR arrays across six diverse species within the crenarchaeal genus Pyrobaculum. We present transcriptional data from each of four species in the genus (P. aerophilum, P. islandicum, P. calidifontis, P. arsenaticum, analyzing mature CRISPR-associated small RNA abundance from over 20 arrays. Within the genus, there is remarkable conservation of CRISPR array structure, as well as unique features that are have not been studied in other archaeal systems. These unique features include: a nearly invariant CRISPR promoter, conservation of direct repeat families, the 5' polarity of CRISPR-associated small RNA abundance, and a novel CRISPR-specific association with homologues of nurA and herA. These analyses provide a genus-level evolutionary perspective on archaeal CRISPR systems, broadening our understanding beyond existing non-comparative model systems.

  6. Genome-wide Analysis of Gene Regulation

    DEFF Research Database (Denmark)

    Chen, Yun

    to protein: through epigenetic modifications, transcription regulators or post-transcriptional controls. The following papers concern several layers of gene regulation with questions answered by different HTS approaches. Genome-wide screening of epigenetic changes by ChIP-seq allowed us to study both spatial...... and temporal alterations of histone modifications (Papers I and II). Coupling the data with machine learning approaches, we established a prediction framework to assess the most informative histone marks as well as their most influential nucleosome positions in predicting the promoter usages. (Papers I...... they regulated or if the sites had global elevated usage rates by multiple TFs. Using RNA-seq, 5’end-seq in combination with depletion of 5’exonuclease as well as nonsensemediated decay (NMD) factors, we systematically analyzed NMD substrates as well as their degradation intermediates in human cells (Paper V...

  7. Knowledge-based analysis of microarrays for the discovery of transcriptional regulation relationships.

    Science.gov (United States)

    Seok, Junhee; Kaushal, Amit; Davis, Ronald W; Xiao, Wenzhong

    2010-01-18

    The large amount of high-throughput genomic data has facilitated the discovery of the regulatory relationships between transcription factors and their target genes. While early methods for discovery of transcriptional regulation relationships from microarray data often focused on the high-throughput experimental data alone, more recent approaches have explored the integration of external knowledge bases of gene interactions. In this work, we develop an algorithm that provides improved performance in the prediction of transcriptional regulatory relationships by supplementing the analysis of microarray data with a new method of integrating information from an existing knowledge base. Using a well-known dataset of yeast microarrays and the Yeast Proteome Database, a comprehensive collection of known information of yeast genes, we show that knowledge-based predictions demonstrate better sensitivity and specificity in inferring new transcriptional interactions than predictions from microarray data alone. We also show that comprehensive, direct and high-quality knowledge bases provide better prediction performance. Comparison of our results with ChIP-chip data and growth fitness data suggests that our predicted genome-wide regulatory pairs in yeast are reasonable candidates for follow-up biological verification. High quality, comprehensive, and direct knowledge bases, when combined with appropriate bioinformatic algorithms, can significantly improve the discovery of gene regulatory relationships from high throughput gene expression data.

  8. Genome-wide profiling of transcription factor binding and epigenetic marks in adipocytes by ChIP-seq

    DEFF Research Database (Denmark)

    Nielsen, Ronni; Mandrup, Susanne

    2014-01-01

    of the most widely used of these technologies. Using these methods, association of transcription factors, cofactors, and epigenetic marks can be mapped to DNA in a genome-wide manner. Here, we provide a detailed protocol for performing ChIP-seq analyses in preadipocytes and adipocytes. We have focused mainly...

  9. A high quality Arabidopsis transcriptome for accurate transcript-level analysis of alternative splicing

    KAUST Repository

    Zhang, Runxuan

    2017-04-05

    Alternative splicing generates multiple transcript and protein isoforms from the same gene and thus is important in gene expression regulation. To date, RNA-sequencing (RNA-seq) is the standard method for quantifying changes in alternative splicing on a genome-wide scale. Understanding the current limitations of RNA-seq is crucial for reliable analysis and the lack of high quality, comprehensive transcriptomes for most species, including model organisms such as Arabidopsis, is a major constraint in accurate quantification of transcript isoforms. To address this, we designed a novel pipeline with stringent filters and assembled a comprehensive Reference Transcript Dataset for Arabidopsis (AtRTD2) containing 82,190 non-redundant transcripts from 34 212 genes. Extensive experimental validation showed that AtRTD2 and its modified version, AtRTD2-QUASI, for use in Quantification of Alternatively Spliced Isoforms, outperform other available transcriptomes in RNA-seq analysis. This strategy can be implemented in other species to build a pipeline for transcript-level expression and alternative splicing analyses.

  10. A high quality Arabidopsis transcriptome for accurate transcript-level analysis of alternative splicing

    KAUST Repository

    Zhang, Runxuan; Calixto, Cristiane  P.  G.; Marquez, Yamile; Venhuizen, Peter; Tzioutziou, Nikoleta A.; Guo, Wenbin; Spensley, Mark; Entizne, Juan Carlos; Lewandowska, Dominika; ten  Have, Sara; Frei  dit  Frey, Nicolas; Hirt, Heribert; James, Allan B.; Nimmo, Hugh G.; Barta, Andrea; Kalyna, Maria; Brown, John  W.  S.

    2017-01-01

    Alternative splicing generates multiple transcript and protein isoforms from the same gene and thus is important in gene expression regulation. To date, RNA-sequencing (RNA-seq) is the standard method for quantifying changes in alternative splicing on a genome-wide scale. Understanding the current limitations of RNA-seq is crucial for reliable analysis and the lack of high quality, comprehensive transcriptomes for most species, including model organisms such as Arabidopsis, is a major constraint in accurate quantification of transcript isoforms. To address this, we designed a novel pipeline with stringent filters and assembled a comprehensive Reference Transcript Dataset for Arabidopsis (AtRTD2) containing 82,190 non-redundant transcripts from 34 212 genes. Extensive experimental validation showed that AtRTD2 and its modified version, AtRTD2-QUASI, for use in Quantification of Alternatively Spliced Isoforms, outperform other available transcriptomes in RNA-seq analysis. This strategy can be implemented in other species to build a pipeline for transcript-level expression and alternative splicing analyses.

  11. Genome-wide Reconstruction of OxyR and SoxRS Transcriptional Regulatory Networks under Oxidative Stress in Escherichia coli K-12 MG1655

    Directory of Open Access Journals (Sweden)

    Sang Woo Seo

    2015-08-01

    Full Text Available Three transcription factors (TFs, OxyR, SoxR, and SoxS, play a critical role in transcriptional regulation of the defense system for oxidative stress in bacteria. However, their full genome-wide regulatory potential is unknown. Here, we perform a genome-scale reconstruction of the OxyR, SoxR, and SoxS regulons in Escherichia coli K-12 MG1655. Integrative data analysis reveals that a total of 68 genes in 51 transcription units (TUs belong to these regulons. Among them, 48 genes showed more than 2-fold changes in expression level under single-TF-knockout conditions. This reconstruction expands the genome-wide roles of these factors to include direct activation of genes related to amino acid biosynthesis (methionine and aromatic amino acids, cell wall synthesis (lipid A biosynthesis and peptidoglycan growth, and divalent metal ion transport (Mn2+, Zn2+, and Mg2+. Investigating the co-regulation of these genes with other stress-response TFs reveals that they are independently regulated by stress-specific TFs.

  12. Mathematical Analysis of Genomic Evolution

    Directory of Open Access Journals (Sweden)

    Cedric Green

    2011-01-01

    Full Text Available Changes in nucleotide sequences, or mutations, accumulate from generation to generation in the genomes of all living organisms. The mutations can be advantageous, deleterious, or neutral. The goal of this project is to determine the amount of advantageous mutations it takes to get human (Homo sapiens DNA from the DNA of genetically distinct organisms. We do this by collecting the genomic data of such organisms, and estimating the amount of mutations it takes to transform yeast (Saccharomyces cerevisiae DNA to the DNA of a human. We calculate the typical number of mutations occurring annually through the organism's average life span and the average mutation rate. This allows us to determine the total number of mutations as well as the probability of advantageous mutations. Not surprisingly, this probability proves to be fairly small. A more precise estimate can be determined by accounting for the differences in the chromosomal structure and phenomena like horizontal gene transfer.

  13. Multiple-integrations of HPV16 genome and altered transcription of viral oncogenes and cellular genes are associated with the development of cervical cancer.

    Directory of Open Access Journals (Sweden)

    Xulian Lu

    Full Text Available The constitutive expression of the high-risk HPV E6 and E7 viral oncogenes is the major cause of cervical cancer. To comprehensively explore the composition of HPV16 early transcripts and their genomic annotation, cervical squamous epithelial tissues from 40 HPV16-infected patients were collected for analysis of papillomavirus oncogene transcripts (APOT. We observed different transcription patterns of HPV16 oncogenes in progression of cervical lesions to cervical cancer and identified one novel transcript. Multiple-integration events in the tissues of cervical carcinoma (CxCa are significantly more often than those of low-grade squamous intraepithelial lesions (LSIL and high-grade squamous intraepithelial lesions (HSIL. Moreover, most cellular genes within or near these integration sites are cancer-associated genes. Taken together, this study suggests that the multiple-integrations of HPV genome during persistent viral infection, which thereby alters the expression patterns of viral oncogenes and integration-related cellular genes, play a crucial role in progression of cervical lesions to cervix cancer.

  14. Genome-Wide Identification and Characterization of BrrTCP Transcription Factors in Brassica rapa ssp. rapa

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    Jiancan Du

    2017-09-01

    Full Text Available The teosinte branched1/cycloidea/proliferating cell factor (TCP gene family is a plant-specific transcription factor that participates in the control of plant development by regulating cell proliferation. However, no report is currently available about this gene family in turnips (Brassica rapa ssp. rapa. In this study, a genome-wide analysis of TCP genes was performed in turnips. Thirty-nine TCP genes in turnip genome were identified and distributed on 10 chromosomes. Phylogenetic analysis clearly showed that the family was classified as two clades: class I and class II. Gene structure and conserved motif analysis showed that the same clade genes have similar gene structures and conserved motifs. The expression profiles of 39 TCP genes were determined through quantitative real-time PCR. Most CIN-type BrrTCP genes were highly expressed in leaf. The members of CYC/TB1 subclade are highly expressed in flower bud and weakly expressed in root. By contrast, class I clade showed more widespread but less tissue-specific expression patterns. Yeast two-hybrid data show that BrrTCP proteins preferentially formed heterodimers. The function of BrrTCP2 was confirmed through ectopic expression of BrrTCP2 in wild-type and loss-of-function ortholog mutant of Arabidopsis. Overexpression of BrrTCP2 in wild-type Arabidopsis resulted in the diminished leaf size. Overexpression of BrrTCP2 in triple mutants of tcp2/4/10 restored the leaf phenotype of tcp2/4/10 to the phenotype of wild type. The comprehensive analysis of turnip TCP gene family provided the foundation to further study the roles of TCP genes in turnips.

  15. Deep Sequencing Reveals the Complete Genome and Evidence for Transcriptional Activity of the First Virus-Like Sequences Identified in Aristotelia chilensis (Maqui Berry

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    Javier Villacreses

    2015-04-01

    Full Text Available Here, we report the genome sequence and evidence for transcriptional activity of a virus-like element in the native Chilean berry tree Aristotelia chilensis. We propose to name the endogenous sequence as Aristotelia chilensis Virus 1 (AcV1. High-throughput sequencing of the genome of this tree uncovered an endogenous viral element, with a size of 7122 bp, corresponding to the complete genome of AcV1. Its sequence contains three open reading frames (ORFs: ORFs 1 and 2 shares 66%–73% amino acid similarity with members of the Caulimoviridae virus family, especially the Petunia vein clearing virus (PVCV, Petuvirus genus. ORF1 encodes a movement protein (MP; ORF2 a Reverse Transcriptase (RT and a Ribonuclease H (RNase H domain; and ORF3 showed no amino acid sequence similarity with any other known virus proteins. Analogous to other known endogenous pararetrovirus sequences (EPRVs, AcV1 is integrated in the genome of Maqui Berry and showed low viral transcriptional activity, which was detected by deep sequencing technology (DNA and RNA-seq. Phylogenetic analysis of AcV1 and other pararetroviruses revealed a closer resemblance with Petuvirus. Overall, our data suggests that AcV1 could be a new member of Caulimoviridae family, genus Petuvirus, and the first evidence of this kind of virus in a fruit plant.

  16. A Distance Measure for Genome Phylogenetic Analysis

    Science.gov (United States)

    Cao, Minh Duc; Allison, Lloyd; Dix, Trevor

    Phylogenetic analyses of species based on single genes or parts of the genomes are often inconsistent because of factors such as variable rates of evolution and horizontal gene transfer. The availability of more and more sequenced genomes allows phylogeny construction from complete genomes that is less sensitive to such inconsistency. For such long sequences, construction methods like maximum parsimony and maximum likelihood are often not possible due to their intensive computational requirement. Another class of tree construction methods, namely distance-based methods, require a measure of distances between any two genomes. Some measures such as evolutionary edit distance of gene order and gene content are computational expensive or do not perform well when the gene content of the organisms are similar. This study presents an information theoretic measure of genetic distances between genomes based on the biological compression algorithm expert model. We demonstrate that our distance measure can be applied to reconstruct the consensus phylogenetic tree of a number of Plasmodium parasites from their genomes, the statistical bias of which would mislead conventional analysis methods. Our approach is also used to successfully construct a plausible evolutionary tree for the γ-Proteobacteria group whose genomes are known to contain many horizontally transferred genes.

  17. Analysis artefacts of the INS-IGF2 fusion transcript

    DEFF Research Database (Denmark)

    Wernersson, Rasmus; Frogne, Thomas; Rescan, Claude

    2015-01-01

    Background: In gene expression analysis, overlapping genes, splice variants, and fusion transcripts are potential sources of data analysis artefacts, depending on how the observed intensity is assigned to one, or more genes. We here exemplify this by an in-depth analysis of the INS-IGF2 fusion...... transcript, which has recently been reported to be among the highest expressed transcripts in human pancreatic beta cells and its protein indicated as a novel autoantigen in Type 1 Diabetes. Results: Through RNA sequencing and variant specific qPCR analyses we demonstrate that the true abundance of INS-IGF2...... is >20,000 fold lower than INS in human beta cells, and we suggest an explanation to the nature of the artefacts which have previously led to overestimation of the gene expression level in selected studies. We reinvestigated the previous reported findings of detection of INS-IGF2 using antibodies both...

  18. Raalin, a transcript enriched in the honey bee brain, is a remnant of genomic rearrangement in Hymenoptera.

    Science.gov (United States)

    Tirosh, Y; Morpurgo, N; Cohen, M; Linial, M; Bloch, G

    2012-06-01

    We identified a predicted compact cysteine-rich sequence in the honey bee genome that we called 'Raalin'. Raalin transcripts are enriched in the brain of adult honey bee workers and drones, with only minimum expression in other tissues or in pre-adult stages. Open-reading frame (ORF) homologues of Raalin were identified in the transcriptomes of fruit flies, mosquitoes and moths. The Raalin-like gene from Drosophila melanogaster encodes for a short secreted protein that is maximally expressed in the adult brain with negligible expression in other tissues or pre-imaginal stages. Raalin-like sequences have also been found in the recently sequenced genomes of six ant species, but not in the jewel wasp Nasonia vitripennis. As in the honey bee, the Raalin-like sequences of ants do not have an ORF. A comparison of the genome region containing Raalin in the genomes of bees, ants and the wasp provides evolutionary support for an extensive genome rearrangement in this sequence. Our analyses identify a new family of ancient cysteine-rich short sequences in insects in which insertions and genome rearrangements may have disrupted this locus in the branch leading to the Hymenoptera. The regulated expression of this transcript suggests that it has a brain-specific function. © 2012 The Authors. Insect Molecular Biology © 2012 The Royal Entomological Society.

  19. Genomic analysis of Fusarium verticillioides.

    Science.gov (United States)

    Brown, D W; Butchko, R A E; Proctor, R H

    2008-09-01

    Fusarium verticillioides (teleomorph Gibberella moniliformis) can be either an endophyte of maize, causing no visible disease, or a pathogen-causing disease of ears, stalks, roots and seedlings. At any stage, this fungus can synthesize fumonisins, a family of mycotoxins structurally similar to the sphingolipid sphinganine. Ingestion of fumonisin-contaminated maize has been associated with a number of animal diseases, including cancer in rodents, and exposure has been correlated with human oesophageal cancer in some regions of the world, and some evidence suggests that fumonisins are a risk factor for neural tube defects. A primary goal of the authors' laboratory is to eliminate fumonisin contamination of maize and maize products. Understanding how and why these toxins are made and the F. verticillioides-maize disease process will allow one to develop novel strategies to limit tissue destruction (rot) and fumonisin production. To meet this goal, genomic sequence data, expressed sequence tags (ESTs) and microarrays are being used to identify F. verticillioides genes involved in the biosynthesis of toxins and plant pathogenesis. This paper describes the current status of F. verticillioides genomic resources and three approaches being used to mine microarray data from a wild-type strain cultured in liquid fumonisin production medium for 12, 24, 48, 72, 96 and 120h. Taken together, these approaches demonstrate the power of microarray technology to provide information on different biological processes.

  20. Identification of concomitant infection with Chlamydia trachomatis IncA-negative mutant and wild-type strains by genomic, transcriptional, and biological characterizations.

    Science.gov (United States)

    Suchland, Robert J; Jeffrey, Brendan M; Xia, Minsheng; Bhatia, Ajay; Chu, Hencelyn G; Rockey, Daniel D; Stamm, Walter E

    2008-12-01

    Clinical isolates of Chlamydia trachomatis that lack IncA on their inclusion membrane form nonfusogenic inclusions and have been associated with milder, subclinical infections in patients. The molecular events associated with the generation of IncA-negative strains and their roles in chlamydial sexually transmitted infections are not clear. We explored the biology of the IncA-negative strains by analyzing their genomic structure, transcription, and growth characteristics in vitro and in vivo in comparison with IncA-positive C. trachomatis strains. Three clinical samples were identified that contained a mixture of IncA-positive and -negative same-serovar C. trachomatis populations, and two more such pairs were found in serial isolates from persistently infected individuals. Genomic sequence analysis of individual strains from each of two serovar-matched pairs showed that these pairs were very similar genetically. In contrast, the genome sequence of an unmatched IncA-negative strain contained over 5,000 nucleotide polymorphisms relative to the genome sequence of a serovar-matched but otherwise unlinked strain. Transcriptional analysis, in vitro culture kinetics, and animal modeling demonstrated that IncA-negative strains isolated in the presence of a serovar-matched wild-type strain are phenotypically more similar to the wild-type strain than are IncA-negative strains isolated in the absence of a serovar-matched wild-type strain. These studies support a model suggesting that a change from an IncA-positive strain to the previously described IncA-negative phenotype may involve multiple steps, the first of which involves a translational inactivation of incA, associated with subsequent unidentified steps that lead to the observed decrease in transcript level, differences in growth rate, and differences in mouse infectivity.

  1. Nucleotide Interdependency in Transcription Factor Binding Sites in the Drosophila Genome.

    Science.gov (United States)

    Dresch, Jacqueline M; Zellers, Rowan G; Bork, Daniel K; Drewell, Robert A

    2016-01-01

    A long-standing objective in modern biology is to characterize the molecular components that drive the development of an organism. At the heart of eukaryotic development lies gene regulation. On the molecular level, much of the research in this field has focused on the binding of transcription factors (TFs) to regulatory regions in the genome known as cis-regulatory modules (CRMs). However, relatively little is known about the sequence-specific binding preferences of many TFs, especially with respect to the possible interdependencies between the nucleotides that make up binding sites. A particular limitation of many existing algorithms that aim to predict binding site sequences is that they do not allow for dependencies between nonadjacent nucleotides. In this study, we use a recently developed computational algorithm, MARZ, to compare binding site sequences using 32 distinct models in a systematic and unbiased approach to explore nucleotide dependencies within binding sites for 15 distinct TFs known to be critical to Drosophila development. Our results indicate that many of these proteins have varying levels of nucleotide interdependencies within their DNA recognition sequences, and that, in some cases, models that account for these dependencies greatly outperform traditional models that are used to predict binding sites. We also directly compare the ability of different models to identify the known KRUPPEL TF binding sites in CRMs and demonstrate that a more complex model that accounts for nucleotide interdependencies performs better when compared with simple models. This ability to identify TFs with critical nucleotide interdependencies in their binding sites will lead to a deeper understanding of how these molecular characteristics contribute to the architecture of CRMs and the precise regulation of transcription during organismal development.

  2. Genome-wide transcriptional response of Silurana (Xenopus tropicalis to infection with the deadly chytrid fungus.

    Directory of Open Access Journals (Sweden)

    Erica Bree Rosenblum

    Full Text Available Emerging infectious diseases are of great concern for both wildlife and humans. Several highly virulent fungal pathogens have recently been discovered in natural populations, highlighting the need for a better understanding of fungal-vertebrate host-pathogen interactions. Because most fungal pathogens are not fatal in the absence of other predisposing conditions, host-pathogen dynamics for deadly fungal pathogens are of particular interest. The chytrid fungus Batrachochytrium dendrobatidis (hereafter Bd infects hundreds of species of frogs in the wild. It is found worldwide and is a significant contributor to the current global amphibian decline. However, the mechanism by which Bd causes death in amphibians, and the response of the host to Bd infection, remain largely unknown. Here we use whole-genome microarrays to monitor the transcriptional responses to Bd infection in the model frog species, Silurana (Xenopus tropicalis, which is susceptible to chytridiomycosis. To elucidate the immune response to Bd and evaluate the physiological effects of chytridiomycosis, we measured gene expression changes in several tissues (liver, skin, spleen following exposure to Bd. We detected a strong transcriptional response for genes involved in physiological processes that can help explain some clinical symptoms of chytridiomycosis at the organismal level. However, we detected surprisingly little evidence of an immune response to Bd exposure, suggesting that this susceptible species may not be mounting efficient innate and adaptive immune responses against Bd. The weak immune response may be partially explained by the thermal conditions of the experiment, which were optimal for Bd growth. However, many immune genes exhibited decreased expression in Bd-exposed frogs compared to control frogs, suggesting a more complex effect of Bd on the immune system than simple temperature-mediated immune suppression. This study generates important baseline data for ongoing

  3. Sexual Polyploidization in Medicago sativa L.: Impact on the Phenotype, Gene Transcription, and Genome Methylation.

    Science.gov (United States)

    Rosellini, Daniele; Ferradini, Nicoletta; Allegrucci, Stefano; Capomaccio, Stefano; Zago, Elisa Debora; Leonetti, Paola; Balech, Bachir; Aversano, Riccardo; Carputo, Domenico; Reale, Lara; Veronesi, Fabio

    2016-04-07

    Polyploidization as the consequence of 2n gamete formation is a prominent mechanism in plant evolution. Studying its effects on the genome, and on genome expression, has both basic and applied interest. We crossed two diploid (2n = 2x = 16) Medicago sativa plants, a subsp. falcata seed parent, and a coerulea × falcata pollen parent that form a mixture of n and 2n eggs and pollen, respectively. Such a cross produced full-sib diploid and tetraploid (2n = 4x = 32) hybrids, the latter being the result of bilateral sexual polyploidization (BSP). These unique materials allowed us to investigate the effects of BSP, and to separate the effect of intraspecific hybridization from those of polyploidization by comparing 2x with 4x full sib progeny plants. Simple sequence repeat marker segregation demonstrated tetrasomic inheritance for all chromosomes but one, demonstrating that these neotetraploids are true autotetraploids. BSP brought about increased biomass, earlier flowering, higher seed set and weight, and larger leaves with larger cells. Microarray analyses with M. truncatula gene chips showed that several hundred genes, related to diverse metabolic functions, changed their expression level as a consequence of polyploidization. In addition, cytosine methylation increased in 2x, but not in 4x, hybrids. Our results indicate that sexual polyploidization induces significant transcriptional novelty, possibly mediated in part by DNA methylation, and phenotypic novelty that could underpin improved adaptation and reproductive success of tetraploid M. sativa with respect to its diploid progenitor. These polyploidy-induced changes may have promoted the adoption of tetraploid alfalfa in agriculture. Copyright © 2016 Rosellini et al.

  4. Sexual Polyploidization in Medicago sativa L.: Impact on the Phenotype, Gene Transcription, and Genome Methylation

    Directory of Open Access Journals (Sweden)

    Daniele Rosellini

    2016-04-01

    Full Text Available Polyploidization as the consequence of 2n gamete formation is a prominent mechanism in plant evolution. Studying its effects on the genome, and on genome expression, has both basic and applied interest. We crossed two diploid (2n = 2x = 16 Medicago sativa plants, a subsp. falcata seed parent, and a coerulea × falcata pollen parent that form a mixture of n and 2n eggs and pollen, respectively. Such a cross produced full-sib diploid and tetraploid (2n = 4x = 32 hybrids, the latter being the result of bilateral sexual polyploidization (BSP. These unique materials allowed us to investigate the effects of BSP, and to separate the effect of intraspecific hybridization from those of polyploidization by comparing 2x with 4x full sib progeny plants. Simple sequence repeat marker segregation demonstrated tetrasomic inheritance for all chromosomes but one, demonstrating that these neotetraploids are true autotetraploids. BSP brought about increased biomass, earlier flowering, higher seed set and weight, and larger leaves with larger cells. Microarray analyses with M. truncatula gene chips showed that several hundred genes, related to diverse metabolic functions, changed their expression level as a consequence of polyploidization. In addition, cytosine methylation increased in 2x, but not in 4x, hybrids. Our results indicate that sexual polyploidization induces significant transcriptional novelty, possibly mediated in part by DNA methylation, and phenotypic novelty that could underpin improved adaptation and reproductive success of tetraploid M. sativa with respect to its diploid progenitor. These polyploidy-induced changes may have promoted the adoption of tetraploid alfalfa in agriculture.

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

    Directory of Open Access Journals (Sweden)

    Zsolt eBoldogkoi

    2012-07-01

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

  6. Genome-wide binding of transcription factor ZEB1 in triple-negative breast cancer cells.

    Science.gov (United States)

    Maturi, Varun; Enroth, Stefan; Heldin, Carl-Henrik; Moustakas, Aristidis

    2018-05-10

    Zinc finger E-box binding homeobox 1 (ZEB1) is a transcriptional regulator involved in embryonic development and cancer progression. ZEB1 induces epithelial-mesenchymal transition (EMT). Triple-negative human breast cancers express high ZEB1 mRNA levels and exhibit features of EMT. In the human triple-negative breast cancer cell model Hs578T, ZEB1 associates with almost 2,000 genes, representing many cellular functions, including cell polarity regulation (DLG2 and FAT3). By introducing a CRISPR-Cas9-mediated 30 bp deletion into the ZEB1 second exon, we observed reduced migratory and anchorage-independent growth capacity of these tumor cells. Transcriptomic analysis of control and ZEB1 knockout cells, revealed 1,372 differentially expressed genes. The TIMP metallopeptidase inhibitor 3 and the teneurin transmembrane protein 2 genes showed increased expression upon loss of ZEB1, possibly mediating pro-tumorigenic actions of ZEB1. This work provides a resource for regulators of cancer progression that function under the transcriptional control of ZEB1. The data confirm that removing a single EMT transcription factor, such as ZEB1, is not sufficient for reverting the triple-negative mesenchymal breast cancer cells into more differentiated, epithelial-like clones, but can reduce tumorigenic potential, suggesting that not all pro-tumorigenic actions of ZEB1 are linked to the EMT. © 2018 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc.

  7. Iterative Chat Transcript Analysis: Making Meaning from Existing Data

    Directory of Open Access Journals (Sweden)

    Steven Baumgart

    2016-04-01

    Full Text Available Objective – In order to better contextualize library data about patron satisfaction with reference services, we analyzed an existing corpus of chat transcripts. Having conducted a similar analysis in 2010, we also compared librarian behaviors over time. Methods – Drawing from the library literature, we identified a set of librarian behaviors closely associated with patron satisfaction. These behaviors include listening to and understanding patrons’ needs, inviting patrons to use the service again, and providing instruction or completing a search for patrons. Analysis of the chat transcripts included establishing a coding schema, applying these codes to individual chat transcripts, and analyzing these codes across the corpus of transcripts for frequency and correlation with other codes. The currently presented analysis used chat transcripts from the fall of 2013 and seeks changes in librarian behavior over time in order to gauge the success of establishing best practices and improving training standardization over the last three years. Results – The analysis shows that librarian behaviors have changed over time, pointing to what campus librarians are doing well, and that implementation of best practices at a campus level after the 2010 analysis may have increased these positive behaviors. The analysis also shows opportunities for further standardization and reinforcement of best practices. Conclusion – Qualitative analysis of already-collected data serves as a model for other units and suggests areas for process improvement, including enhanced coder training and code schema design. Further analysis of chat patrons’ questions is also warranted, including investigation of the relationship between subject- and location-specific questions and referrals.

  8. Genome-Wide Spectra of Transcription Insertions and Deletions Reveal That Slippage Depends on RNA:DNA Hybrid Complementarity.

    Science.gov (United States)

    Traverse, Charles C; Ochman, Howard

    2017-08-29

    Advances in sequencing technologies have enabled direct quantification of genome-wide errors that occur during RNA transcription. These errors occur at rates that are orders of magnitude higher than rates during DNA replication, but due to technical difficulties such measurements have been limited to single-base substitutions and have not yet quantified the scope of transcription insertions and deletions. Previous reporter gene assay findings suggested that transcription indels are produced exclusively by elongation complex slippage at homopolymeric runs, so we enumerated indels across the protein-coding transcriptomes of Escherichia coli and Buchnera aphidicola , which differ widely in their genomic base compositions and incidence of repeat regions. As anticipated from prior assays, transcription insertions prevailed in homopolymeric runs of A and T; however, transcription deletions arose in much more complex sequences and were rarely associated with homopolymeric runs. By reconstructing the relocated positions of the elongation complex as inferred from the sequences inserted or deleted during transcription, we show that continuation of transcription after slippage hinges on the degree of nucleotide complementarity within the RNA:DNA hybrid at the new DNA template location. IMPORTANCE The high level of mistakes generated during transcription can result in the accumulation of malfunctioning and misfolded proteins which can alter global gene regulation and in the expenditure of energy to degrade these nonfunctional proteins. The transcriptome-wide occurrence of base substitutions has been elucidated in bacteria, but information on transcription insertions and deletions-errors that potentially have more dire effects on protein function-is limited to reporter gene constructs. Here, we capture the transcriptome-wide spectrum of insertions and deletions in Escherichia coli and Buchnera aphidicola and show that they occur at rates approaching those of base substitutions

  9. Genome-Wide Search for Competing Endogenous RNAs Responsible for the Effects Induced by Ebola Virus Replication and Transcription Using a trVLP System

    Directory of Open Access Journals (Sweden)

    Zhong-Yi Wang

    2017-11-01

    Full Text Available Understanding how infected cells respond to Ebola virus (EBOV and how this response changes during the process of viral replication and transcription are very important for establishing effective antiviral strategies. In this study, we conducted a genome-wide screen to identify long non-coding RNAs (lncRNAs, circular RNAs (circRNAs, micro RNAs (miRNAs, and mRNAs differentially expressed during replication and transcription using a tetracistronic transcription and replication-competent virus-like particle (trVLP system that models the life cycle of EBOV in 293T cells. To characterize the expression patterns of these differentially expressed RNAs, we performed a series cluster analysis, and up- or down-regulated genes were selected to establish a gene co-expression network. Competing endogenous RNA (ceRNA networks based on the RNAs responsible for the effects induced by EBOV replication and transcription in human cells, including circRNAs, lncRNAs, miRNAs, and mRNAs, were constructed for the first time. Based on these networks, the interaction details of circRNA-chr19 were explored. Our results demonstrated that circRNA-chr19 targeting miR-30b-3p regulated CLDN18 expression by functioning as a ceRNA. These findings may have important implications for further studies of the mechanisms of EBOV replication and transcription. These RNAs potentially have important functions and may be promising targets for EBOV therapy.

  10. Genome-wide DNA binding pattern of the homeodomain transcription factor Sine oculis (So in the developing eye of Drosophila melanogaster

    Directory of Open Access Journals (Sweden)

    Barbara Jusiak

    2014-12-01

    Full Text Available The eye of the fruit fly Drosophila melanogaster provides a highly tractable genetic model system for the study of animal development, and many genes that regulate Drosophila eye formation have homologs implicated in human development and disease. Among these is the homeobox gene sine oculis (so, which encodes a homeodomain transcription factor (TF that is both necessary for eye development and sufficient to reprogram a subset of cells outside the normal eye field toward an eye fate. We have performed a genome-wide analysis of So binding to DNA prepared from developing Drosophila eye tissue in order to identify candidate direct targets of So-mediated transcriptional regulation, as described in our recent article [20]. The data are available from NCBI Gene Expression Omnibus (GEO with the accession number GSE52943. Here we describe the methods, data analysis, and quality control of our So ChIP-seq dataset.

  11. Genome-wide mapping of boundary element-associated factor (BEAF) binding sites in Drosophila melanogaster links BEAF to transcription.

    Science.gov (United States)

    Jiang, Nan; Emberly, Eldon; Cuvier, Olivier; Hart, Craig M

    2009-07-01

    Insulator elements play a role in gene regulation that is potentially linked to nuclear organization. Boundary element-associated factors (BEAFs) 32A and 32B associate with hundreds of sites on Drosophila polytene chromosomes. We hybridized DNA isolated by chromatin immunoprecipitation to genome tiling microarrays to construct a genome-wide map of BEAF binding locations. A distinct difference in the association of 32A and 32B with chromatin was noted. We identified 1,820 BEAF peaks and found that more than 85% were less than 300 bp from transcription start sites. Half are between head-to-head gene pairs. BEAF-associated genes are transcriptionally active as judged by the presence of RNA polymerase II, dimethylated histone H3 K4, and the alternative histone H3.3. Forty percent of these genes are also associated with the polymerase negative elongation factor NELF. Like NELF-associated genes, most BEAF-associated genes are highly expressed. Using quantitative reverse transcription-PCR, we found that the expression levels of most BEAF-associated genes decrease in embryos and cultured cells lacking BEAF. These results provide an unexpected link between BEAF and transcription, suggesting that BEAF plays a role in maintaining most associated promoter regions in an environment that facilitates high transcription levels.

  12. Predicting transcription factor binding sites using local over-representation and comparative genomics

    Directory of Open Access Journals (Sweden)

    Touzet Hélène

    2006-08-01

    Full Text Available Abstract Background Identifying cis-regulatory elements is crucial to understanding gene expression, which highlights the importance of the computational detection of overrepresented transcription factor binding sites (TFBSs in coexpressed or coregulated genes. However, this is a challenging problem, especially when considering higher eukaryotic organisms. Results We have developed a method, named TFM-Explorer, that searches for locally overrepresented TFBSs in a set of coregulated genes, which are modeled by profiles provided by a database of position weight matrices. The novelty of the method is that it takes advantage of spatial conservation in the sequence and supports multiple species. The efficiency of the underlying algorithm and its robustness to noise allow weak regulatory signals to be detected in large heterogeneous data sets. Conclusion TFM-Explorer provides an efficient way to predict TFBS overrepresentation in related sequences. Promising results were obtained in a variety of examples in human, mouse, and rat genomes. The software is publicly available at http://bioinfo.lifl.fr/TFM-Explorer.

  13. Whole genome sequence analysis of Mycobacterium suricattae

    KAUST Repository

    Dippenaar, Anzaan; Parsons, Sven David Charles; Sampson, Samantha Leigh; Van Der Merwe, Ruben Gerhard; Drewe, Julian Ashley; Abdallah, Abdallah; Siame, Kabengele Keith; Gey Van Pittius, Nicolaas Claudius; Van Helden, Paul David; Pain, Arnab; Warren, Robin Mark

    2015-01-01

    Tuberculosis occurs in various mammalian hosts and is caused by a range of different lineages of the Mycobacterium tuberculosis complex (MTBC). A recently described member, Mycobacterium suricattae, causes tuberculosis in meerkats (Suricata suricatta) in Southern Africa and preliminary genetic analysis showed this organism to be closely related to an MTBC pathogen of rock hyraxes (Procavia capensis), the dassie bacillus. Here we make use of whole genome sequencing to describe the evolution of the genome of M. suricattae, including known and novel regions of difference, SNPs and IS6110 insertion sites. We used genome-wide phylogenetic analysis to show that M. suricattae clusters with the chimpanzee bacillus, previously isolated from a chimpanzee (Pan troglodytes) in West Africa. We propose an evolutionary scenario for the Mycobacterium africanum lineage 6 complex, showing the evolutionary relationship of M. africanum and chimpanzee bacillus, and the closely related members M. suricattae, dassie bacillus and Mycobacterium mungi.

  14. Whole genome sequence analysis of Mycobacterium suricattae

    KAUST Repository

    Dippenaar, Anzaan

    2015-10-21

    Tuberculosis occurs in various mammalian hosts and is caused by a range of different lineages of the Mycobacterium tuberculosis complex (MTBC). A recently described member, Mycobacterium suricattae, causes tuberculosis in meerkats (Suricata suricatta) in Southern Africa and preliminary genetic analysis showed this organism to be closely related to an MTBC pathogen of rock hyraxes (Procavia capensis), the dassie bacillus. Here we make use of whole genome sequencing to describe the evolution of the genome of M. suricattae, including known and novel regions of difference, SNPs and IS6110 insertion sites. We used genome-wide phylogenetic analysis to show that M. suricattae clusters with the chimpanzee bacillus, previously isolated from a chimpanzee (Pan troglodytes) in West Africa. We propose an evolutionary scenario for the Mycobacterium africanum lineage 6 complex, showing the evolutionary relationship of M. africanum and chimpanzee bacillus, and the closely related members M. suricattae, dassie bacillus and Mycobacterium mungi.

  15. Genomewide analysis of TCP transcription factor gene family in ...

    Indian Academy of Sciences (India)

    Home; Journals; Journal of Genetics; Volume 93; Issue 3. Genomewide ... Teosinte branched1/cycloidea/proliferating cell factor1 (TCP) proteins are a large family of transcriptional regulators in angiosperms. They are ... To the best of our knowledge, this is the first study of a genomewide analysis of apple TCP gene family.

  16. Optimization of oligonucleotide arrays and RNA amplification protocols for analysis of transcript structure and alternative splicing.

    Science.gov (United States)

    Castle, John; Garrett-Engele, Phil; Armour, Christopher D; Duenwald, Sven J; Loerch, Patrick M; Meyer, Michael R; Schadt, Eric E; Stoughton, Roland; Parrish, Mark L; Shoemaker, Daniel D; Johnson, Jason M

    2003-01-01

    Microarrays offer a high-resolution means for monitoring pre-mRNA splicing on a genomic scale. We have developed a novel, unbiased amplification protocol that permits labeling of entire transcripts. Also, hybridization conditions, probe characteristics, and analysis algorithms were optimized for detection of exons, exon-intron edges, and exon junctions. These optimized protocols can be used to detect small variations and isoform mixtures, map the tissue specificity of known human alternative isoforms, and provide a robust, scalable platform for high-throughput discovery of alternative splicing.

  17. Microarray and cDNA sequence analysis of transcription during nerve-dependent limb regeneration

    Directory of Open Access Journals (Sweden)

    Bryant Susan V

    2009-01-01

    Full Text Available Abstract Background Microarray analysis and 454 cDNA sequencing were used to investigate a centuries-old problem in regenerative biology: the basis of nerve-dependent limb regeneration in salamanders. Innervated (NR and denervated (DL forelimbs of Mexican axolotls were amputated and transcripts were sampled after 0, 5, and 14 days of regeneration. Results Considerable similarity was observed between NR and DL transcriptional programs at 5 and 14 days post amputation (dpa. Genes with extracellular functions that are critical to wound healing were upregulated while muscle-specific genes were downregulated. Thus, many processes that are regulated during early limb regeneration do not depend upon nerve-derived factors. The majority of the transcriptional differences between NR and DL limbs were correlated with blastema formation; cell numbers increased in NR limbs after 5 dpa and this yielded distinct transcriptional signatures of cell proliferation in NR limbs at 14 dpa. These transcriptional signatures were not observed in DL limbs. Instead, gene expression changes within DL limbs suggest more diverse and protracted wound-healing responses. 454 cDNA sequencing complemented the microarray analysis by providing deeper sampling of transcriptional programs and associated biological processes. Assembly of new 454 cDNA sequences with existing expressed sequence tag (EST contigs from the Ambystoma EST database more than doubled (3935 to 9411 the number of non-redundant human-A. mexicanum orthologous sequences. Conclusion Many new candidate gene sequences were discovered for the first time and these will greatly enable future studies of wound healing, epigenetics, genome stability, and nerve-dependent blastema formation and outgrowth using the axolotl model.

  18. Analysis of transcript and protein overlap in a human osteosarcoma cell line

    Directory of Open Access Journals (Sweden)

    Emanuelsson Olof

    2010-12-01

    Full Text Available Abstract Background An interesting field of research in genomics and proteomics is to compare the overlap between the transcriptome and the proteome. Recently, the tools to analyse gene and protein expression on a whole-genome scale have been improved, including the availability of the new generation sequencing instruments and high-throughput antibody-based methods to analyze the presence and localization of proteins. In this study, we used massive transcriptome sequencing (RNA-seq to investigate the transcriptome of a human osteosarcoma cell line and compared the expression levels with in situ protein data obtained in-situ from antibody-based immunohistochemistry (IHC and immunofluorescence microscopy (IF. Results A large-scale analysis based on 2749 genes was performed, corresponding to approximately 13% of the protein coding genes in the human genome. We found the presence of both RNA and proteins to a large fraction of the analyzed genes with 60% of the analyzed human genes detected by all three methods. Only 34 genes (1.2% were not detected on the transcriptional or protein level with any method. Our data suggest that the majority of the human genes are expressed at detectable transcript or protein levels in this cell line. Since the reliability of antibodies depends on possible cross-reactivity, we compared the RNA and protein data using antibodies with different reliability scores based on various criteria, including Western blot analysis. Gene products detected in all three platforms generally have good antibody validation scores, while those detected only by antibodies, but not by RNA sequencing, generally consist of more low-scoring antibodies. Conclusion This suggests that some antibodies are staining the cells in an unspecific manner, and that assessment of transcript presence by RNA-seq can provide guidance for validation of the corresponding antibodies.

  19. The transcription elongation factor Bur1-Bur2 interacts with replication protein A and maintains genome stability during replication stress

    DEFF Research Database (Denmark)

    Clausing, Emanuel; Mayer, Andreas; Chanarat, Sittinan

    2010-01-01

    Multiple DNA-associated processes such as DNA repair, replication, and recombination are crucial for the maintenance of genome integrity. Here, we show a novel interaction between the transcription elongation factor Bur1-Bur2 and replication protein A (RPA), the eukaryotic single-stranded DNA......-binding protein with functions in DNA repair, recombination, and replication. Bur1 interacted via its C-terminal domain with RPA, and bur1-¿C mutants showed a deregulated DNA damage response accompanied by increased sensitivity to DNA damage and replication stress as well as increased levels of persisting Rad52...... foci. Interestingly, the DNA damage sensitivity of an rfa1 mutant was suppressed by bur1 mutation, further underscoring a functional link between these two protein complexes. The transcription elongation factor Bur1-Bur2 interacts with RPA and maintains genome integrity during DNA replication stress....

  20. Uncovering transcriptional regulation of glycerol metabolism in Aspergilli through genome-wide gene expression data anlysis

    DEFF Research Database (Denmark)

    Salazar, Margarita Pena; Vongsangnak, Wanwipa; Panagiotou, Gianni

    2009-01-01

    Glycerol is catabolized by a wide range of microorganisms including Aspergillus species. To identify the transcriptional regulation of glycerol metabolism in Aspergillus, we analyzed data from triplicate batch fermentations of three different Aspergilli (Aspergillus nidulans, Aspergillus oryzae...... and Aspergillus niger) with glucose and glycerol as carbon sources. Protein comparisons and cross-analysis with gene expression data of all three species resulted in the identification of 88 genes having a conserved response across the three Aspergilli. A promoter analysis of the up-regulated genes led...... to the identification of a conserved binding site for a putative regulator to be 5′-TGCGGGGA-3′, a binding site that is similar to the binding site for Adr1 in yeast and humans. We show that this Adr1 consensus binding sequence was over-represented on promoter regions of several genes in A. nidulans, A. oryzae and A...

  1. Comparative genome analysis of Basidiomycete fungi

    Energy Technology Data Exchange (ETDEWEB)

    Riley, Robert; Salamov, Asaf; Henrissat, Bernard; Nagy, Laszlo; Brown, Daren; Held, Benjamin; Baker, Scott; Blanchette, Robert; Boussau, Bastien; Doty, Sharon L.; Fagnan, Kirsten; Floudas, Dimitris; Levasseur, Anthony; Manning, Gerard; Martin, Francis; Morin, Emmanuelle; Otillar, Robert; Pisabarro, Antonio; Walton, Jonathan; Wolfe, Ken; Hibbett, David; Grigoriev, Igor

    2013-08-07

    Fungi of the phylum Basidiomycota (basidiomycetes), make up some 37percent of the described fungi, and are important in forestry, agriculture, medicine, and bioenergy. This diverse phylum includes symbionts, pathogens, and saprotrophs including the majority of wood decaying and ectomycorrhizal species. To better understand the genetic diversity of this phylum we compared the genomes of 35 basidiomycetes including 6 newly sequenced genomes. These genomes span extremes of genome size, gene number, and repeat content. Analysis of core genes reveals that some 48percent of basidiomycete proteins are unique to the phylum with nearly half of those (22percent) found in only one organism. Correlations between lifestyle and certain gene families are evident. Phylogenetic patterns of plant biomass-degrading genes in Agaricomycotina suggest a continuum rather than a dichotomy between the white rot and brown rot modes of wood decay. Based on phylogenetically-informed PCA analysis of wood decay genes, we predict that that Botryobasidium botryosum and Jaapia argillacea have properties similar to white rot species, although neither has typical ligninolytic class II fungal peroxidases (PODs). This prediction is supported by growth assays in which both fungi exhibit wood decay with white rot-like characteristics. Based on this, we suggest that the white/brown rot dichotomy may be inadequate to describe the full range of wood decaying fungi. Analysis of the rate of discovery of proteins with no or few homologs suggests the value of continued sequencing of basidiomycete fungi.

  2. Transcriptional analysis of exopolysaccharides biosynthesis gene clusters in Lactobacillus plantarum.

    Science.gov (United States)

    Vastano, Valeria; Perrone, Filomena; Marasco, Rosangela; Sacco, Margherita; Muscariello, Lidia

    2016-04-01

    Exopolysaccharides (EPS) from lactic acid bacteria contribute to specific rheology and texture of fermented milk products and find applications also in non-dairy foods and in therapeutics. Recently, four clusters of genes (cps) associated with surface polysaccharide production have been identified in Lactobacillus plantarum WCFS1, a probiotic and food-associated lactobacillus. These clusters are involved in cell surface architecture and probably in release and/or exposure of immunomodulating bacterial molecules. Here we show a transcriptional analysis of these clusters. Indeed, RT-PCR experiments revealed that the cps loci are organized in five operons. Moreover, by reverse transcription-qPCR analysis performed on L. plantarum WCFS1 (wild type) and WCFS1-2 (ΔccpA), we demonstrated that expression of three cps clusters is under the control of the global regulator CcpA. These results, together with the identification of putative CcpA target sequences (catabolite responsive element CRE) in the regulatory region of four out of five transcriptional units, strongly suggest for the first time a role of the master regulator CcpA in EPS gene transcription among lactobacilli.

  3. Rapid Genome-wide Recruitment of RNA Polymerase II Drives Transcription, Splicing, and Translation Events during T Cell Responses

    Directory of Open Access Journals (Sweden)

    Kathrin Davari

    2017-04-01

    Full Text Available Summary: Activation of immune cells results in rapid functional changes, but how such fast changes are accomplished remains enigmatic. By combining time courses of 4sU-seq, RNA-seq, ribosome profiling (RP, and RNA polymerase II (RNA Pol II ChIP-seq during T cell activation, we illustrate genome-wide temporal dynamics for ∼10,000 genes. This approach reveals not only immediate-early and posttranscriptionally regulated genes but also coupled changes in transcription and translation for >90% of genes. Recruitment, rather than release of paused RNA Pol II, primarily mediates transcriptional changes. This coincides with a genome-wide temporary slowdown in cotranscriptional splicing, even for polyadenylated mRNAs that are localized at the chromatin. Subsequent splicing optimization correlates with increasing Ser-2 phosphorylation of the RNA Pol II carboxy-terminal domain (CTD and activation of the positive transcription elongation factor (pTEFb. Thus, rapid de novo recruitment of RNA Pol II dictates the course of events during T cell activation, particularly transcription, splicing, and consequently translation. : Davari et al. visualize global changes in RNA Pol II binding, transcription, splicing, and translation. T cells change their functional program by rapid de novo recruitment of RNA Pol II and coupled changes in transcription and translation. This coincides with fluctuations in RNA Pol II phosphorylation and a temporary reduction in cotranscriptional splicing. Keywords: RNA Pol II, cotranscriptional splicing, T cell activation, ribosome profiling, 4sU, H3K36, Ser-5 RNA Pol II, Ser-2 RNA Pol II, immune response, immediate-early genes

  4. Genome wide predictions of miRNA regulation by transcription factors.

    Science.gov (United States)

    Ruffalo, Matthew; Bar-Joseph, Ziv

    2016-09-01

    Reconstructing regulatory networks from expression and interaction data is a major goal of systems biology. While much work has focused on trying to experimentally and computationally determine the set of transcription-factors (TFs) and microRNAs (miRNAs) that regulate genes in these networks, relatively little work has focused on inferring the regulation of miRNAs by TFs. Such regulation can play an important role in several biological processes including development and disease. The main challenge for predicting such interactions is the very small positive training set currently available. Another challenge is the fact that a large fraction of miRNAs are encoded within genes making it hard to determine the specific way in which they are regulated. To enable genome wide predictions of TF-miRNA interactions, we extended semi-supervised machine-learning approaches to integrate a large set of different types of data including sequence, expression, ChIP-seq and epigenetic data. As we show, the methods we develop achieve good performance on both a labeled test set, and when analyzing general co-expression networks. We next analyze mRNA and miRNA cancer expression data, demonstrating the advantage of using the predicted set of interactions for identifying more coherent and relevant modules, genes, and miRNAs. The complete set of predictions is available on the supporting website and can be used by any method that combines miRNAs, genes, and TFs. Code and full set of predictions are available from the supporting website: http://cs.cmu.edu/~mruffalo/tf-mirna/ zivbj@cs.cmu.edu Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  5. Genome-wide investigation and expression profiling of AP2/ERF transcription factor superfamily in foxtail millet (Setaria italica L.).

    Science.gov (United States)

    Lata, Charu; Mishra, Awdhesh Kumar; Muthamilarasan, Mehanathan; Bonthala, Venkata Suresh; Khan, Yusuf; Prasad, Manoj

    2014-01-01

    The APETALA2/ethylene-responsive element binding factor (AP2/ERF) family is one of the largest transcription factor (TF) families in plants that includes four major sub-families, namely AP2, DREB (dehydration responsive element binding), ERF (ethylene responsive factors) and RAV (Related to ABI3/VP). AP2/ERFs are known to play significant roles in various plant processes including growth and development and biotic and abiotic stress responses. Considering this, a comprehensive genome-wide study was conducted in foxtail millet (Setaria italica L.). A total of 171 AP2/ERF genes were identified by systematic sequence analysis and were physically mapped onto nine chromosomes. Phylogenetic analysis grouped AP2/ERF genes into six classes (I to VI). Duplication analysis revealed that 12 (∼7%) SiAP2/ERF genes were tandem repeated and 22 (∼13%) were segmentally duplicated. Comparative physical mapping between foxtail millet AP2/ERF genes and its orthologs of sorghum (18 genes), maize (14 genes), rice (9 genes) and Brachypodium (6 genes) showed the evolutionary insights of AP2/ERF gene family and also the decrease in orthology with increase in phylogenetic distance. The evolutionary significance in terms of gene-duplication and divergence was analyzed by estimating synonymous and non-synonymous substitution rates. Expression profiling of candidate AP2/ERF genes against drought, salt and phytohormones revealed insights into their precise and/or overlapping expression patterns which could be responsible for their functional divergence in foxtail millet. The study showed that the genes SiAP2/ERF-069, SiAP2/ERF-103 and SiAP2/ERF-120 may be considered as potential candidate genes for further functional validation as well for utilization in crop improvement programs for stress resistance since these genes were up-regulated under drought and salinity stresses in ABA dependent manner. Altogether the present study provides new insights into evolution, divergence and systematic

  6. Genome-wide investigation and expression profiling of AP2/ERF transcription factor superfamily in foxtail millet (Setaria italica L..

    Directory of Open Access Journals (Sweden)

    Charu Lata

    Full Text Available The APETALA2/ethylene-responsive element binding factor (AP2/ERF family is one of the largest transcription factor (TF families in plants that includes four major sub-families, namely AP2, DREB (dehydration responsive element binding, ERF (ethylene responsive factors and RAV (Related to ABI3/VP. AP2/ERFs are known to play significant roles in various plant processes including growth and development and biotic and abiotic stress responses. Considering this, a comprehensive genome-wide study was conducted in foxtail millet (Setaria italica L.. A total of 171 AP2/ERF genes were identified by systematic sequence analysis and were physically mapped onto nine chromosomes. Phylogenetic analysis grouped AP2/ERF genes into six classes (I to VI. Duplication analysis revealed that 12 (∼7% SiAP2/ERF genes were tandem repeated and 22 (∼13% were segmentally duplicated. Comparative physical mapping between foxtail millet AP2/ERF genes and its orthologs of sorghum (18 genes, maize (14 genes, rice (9 genes and Brachypodium (6 genes showed the evolutionary insights of AP2/ERF gene family and also the decrease in orthology with increase in phylogenetic distance. The evolutionary significance in terms of gene-duplication and divergence was analyzed by estimating synonymous and non-synonymous substitution rates. Expression profiling of candidate AP2/ERF genes against drought, salt and phytohormones revealed insights into their precise and/or overlapping expression patterns which could be responsible for their functional divergence in foxtail millet. The study showed that the genes SiAP2/ERF-069, SiAP2/ERF-103 and SiAP2/ERF-120 may be considered as potential candidate genes for further functional validation as well for utilization in crop improvement programs for stress resistance since these genes were up-regulated under drought and salinity stresses in ABA dependent manner. Altogether the present study provides new insights into evolution, divergence and

  7. Genome-wide profiling of H3K56 acetylation and transcription factor binding sites in human adipocytes.

    Directory of Open Access Journals (Sweden)

    Kinyui Alice Lo

    Full Text Available The growing epidemic of obesity and metabolic diseases calls for a better understanding of adipocyte biology. The regulation of transcription in adipocytes is particularly important, as it is a target for several therapeutic approaches. Transcriptional outcomes are influenced by both histone modifications and transcription factor binding. Although the epigenetic states and binding sites of several important transcription factors have been profiled in the mouse 3T3-L1 cell line, such data are lacking in human adipocytes. In this study, we identified H3K56 acetylation sites in human adipocytes derived from mesenchymal stem cells. H3K56 is acetylated by CBP and p300, and deacetylated by SIRT1, all are proteins with important roles in diabetes and insulin signaling. We found that while almost half of the genome shows signs of H3K56 acetylation, the highest level of H3K56 acetylation is associated with transcription factors and proteins in the adipokine signaling and Type II Diabetes pathways. In order to discover the transcription factors that recruit acetyltransferases and deacetylases to sites of H3K56 acetylation, we analyzed DNA sequences near H3K56 acetylated regions and found that the E2F recognition sequence was enriched. Using chromatin immunoprecipitation followed by high-throughput sequencing, we confirmed that genes bound by E2F4, as well as those by HSF-1 and C/EBPα, have higher than expected levels of H3K56 acetylation, and that the transcription factor binding sites and acetylation sites are often adjacent but rarely overlap. We also discovered a significant difference between bound targets of C/EBPα in 3T3-L1 and human adipocytes, highlighting the need to construct species-specific epigenetic and transcription factor binding site maps. This is the first genome-wide profile of H3K56 acetylation, E2F4, C/EBPα and HSF-1 binding in human adipocytes, and will serve as an important resource for better understanding adipocyte

  8. Genome-Wide Classification and Evolutionary and Expression Analyses of Citrus MYB Transcription Factor Families in Sweet Orange

    Science.gov (United States)

    Hou, Xiao-Jin; Li, Si-Bei; Liu, Sheng-Rui; Hu, Chun-Gen; Zhang, Jin-Zhi

    2014-01-01

    MYB family genes are widely distributed in plants and comprise one of the largest transcription factors involved in various developmental processes and defense responses of plants. To date, few MYB genes and little expression profiling have been reported for citrus. Here, we describe and classify 177 members of the sweet orange MYB gene (CsMYB) family in terms of their genomic gene structures and similarity to their putative Arabidopsis orthologs. According to these analyses, these CsMYBs were categorized into four groups (4R-MYB, 3R-MYB, 2R-MYB and 1R-MYB). Gene structure analysis revealed that 1R-MYB genes possess relatively more introns as compared with 2R-MYB genes. Investigation of their chromosomal localizations revealed that these CsMYBs are distributed across nine chromosomes. Sweet orange includes a relatively small number of MYB genes compared with the 198 members in Arabidopsis, presumably due to a paralog reduction related to repetitive sequence insertion into promoter and non-coding transcribed region of the genes. Comparative studies of CsMYBs and Arabidopsis showed that CsMYBs had fewer gene duplication events. Expression analysis revealed that the MYB gene family has a wide expression profile in sweet orange development and plays important roles in development and stress responses. In addition, 337 new putative microsatellites with flanking sequences sufficient for primer design were also identified from the 177 CsMYBs. These results provide a useful reference for the selection of candidate MYB genes for cloning and further functional analysis forcitrus. PMID:25375352

  9. Analysis of Phonetic Transcriptions for Danish Automatic Speech Recognition

    DEFF Research Database (Denmark)

    Kirkedal, Andreas Søeborg

    2013-01-01

    Automatic speech recognition (ASR) relies on three resources: audio, orthographic transcriptions and a pronunciation dictionary. The dictionary or lexicon maps orthographic words to sequences of phones or phonemes that represent the pronunciation of the corresponding word. The quality of a speech....... The analysis indicates that transcribing e.g. stress or vowel duration has a negative impact on performance. The best performance is obtained with coarse phonetic annotation and improves performance 1% word error rate and 3.8% sentence error rate....

  10. Evolutionary Analysis of DELLA-Associated Transcriptional Networks

    Directory of Open Access Journals (Sweden)

    Miguel A. Blázquez

    2017-04-01

    Full Text Available DELLA proteins are transcriptional regulators present in all land plants which have been shown to modulate the activity of over 100 transcription factors in Arabidopsis, involved in multiple physiological and developmental processes. It has been proposed that DELLAs transduce environmental information to pre-wired transcriptional circuits because their stability is regulated by gibberellins (GAs, whose homeostasis largely depends on environmental signals. The ability of GAs to promote DELLA degradation coincides with the origin of vascular plants, but the presence of DELLAs in other land plants poses at least two questions: what regulatory properties have DELLAs provided to the behavior of transcriptional networks in land plants, and how has the recruitment of DELLAs by GA signaling affected this regulation. To address these issues, we have constructed gene co-expression networks of four different organisms within the green lineage with different properties regarding DELLAs: Arabidopsis thaliana and Solanum lycopersicum (both with GA-regulated DELLA proteins, Physcomitrella patens (with GA-independent DELLA proteins and Chlamydomonas reinhardtii (a green alga without DELLA, and we have examined the relative evolution of the subnetworks containing the potential DELLA-dependent transcriptomes. Network analysis indicates a relative increase in parameters associated with the degree of interconnectivity in the DELLA-associated subnetworks of land plants, with a stronger effect in species with GA-regulated DELLA proteins. These results suggest that DELLAs may have played a role in the coordination of multiple transcriptional programs along evolution, and the function of DELLAs as regulatory ‘hubs’ became further consolidated after their recruitment by GA signaling in higher plants.

  11. RNA Transcriptional Biosignature Analysis for Identifying Febrile Infants With Serious Bacterial Infections in the Emergency Department

    Science.gov (United States)

    Mahajan, Prashant; Kuppermann, Nathan; Suarez, Nicolas; Mejias, Asuncion; Casper, Charlie; Dean, J. Michael; Ramilo, Octavio

    2015-01-01

    Objectives To develop the infrastructure and demonstrate the feasibility of conducting microarray-based RNA transcriptional profile analyses for the diagnosis of serious bacterial infections in febrile infants 60 days and younger in a multicenter pediatric emergency research network. Methods We designed a prospective multicenter cohort study with the aim of enrolling more than 4000 febrile infants 60 days and younger. To ensure success of conducting complex genomic studies in emergency department (ED) settings, we established an infrastructure within the Pediatric Emergency Care Applied Research Network, including 21 sites, to evaluate RNA transcriptional profiles in young febrile infants. We developed a comprehensive manual of operations and trained site investigators to obtain and process blood samples for RNA extraction and genomic analyses. We created standard operating procedures for blood sample collection, processing, storage, shipping, and analyses. We planned to prospectively identify, enroll, and collect 1 mL blood samples for genomic analyses from eligible patients to identify logistical issues with study procedures. Finally, we planned to batch blood samples and determined RNA quantity and quality at the central microarray laboratory and organized data analysis with the Pediatric Emergency Care Applied Research Network data coordinating center. Below we report on establishment of the infrastructure and the feasibility success in the first year based on the enrollment of a limited number of patients. Results We successfully established the infrastructure at 21 EDs. Over the first 5 months we enrolled 79% (74 of 94) of eligible febrile infants. We were able to obtain and ship 1 mL of blood from 74% (55 of 74) of enrolled participants, with at least 1 sample per participating ED. The 55 samples were shipped and evaluated at the microarray laboratory, and 95% (52 of 55) of blood samples were of adequate quality and contained sufficient RNA for expression

  12. FGWAS: Functional genome wide association analysis.

    Science.gov (United States)

    Huang, Chao; Thompson, Paul; Wang, Yalin; Yu, Yang; Zhang, Jingwen; Kong, Dehan; Colen, Rivka R; Knickmeyer, Rebecca C; Zhu, Hongtu

    2017-10-01

    Functional phenotypes (e.g., subcortical surface representation), which commonly arise in imaging genetic studies, have been used to detect putative genes for complexly inherited neuropsychiatric and neurodegenerative disorders. However, existing statistical methods largely ignore the functional features (e.g., functional smoothness and correlation). The aim of this paper is to develop a functional genome-wide association analysis (FGWAS) framework to efficiently carry out whole-genome analyses of functional phenotypes. FGWAS consists of three components: a multivariate varying coefficient model, a global sure independence screening procedure, and a test procedure. Compared with the standard multivariate regression model, the multivariate varying coefficient model explicitly models the functional features of functional phenotypes through the integration of smooth coefficient functions and functional principal component analysis. Statistically, compared with existing methods for genome-wide association studies (GWAS), FGWAS can substantially boost the detection power for discovering important genetic variants influencing brain structure and function. Simulation studies show that FGWAS outperforms existing GWAS methods for searching sparse signals in an extremely large search space, while controlling for the family-wise error rate. We have successfully applied FGWAS to large-scale analysis of data from the Alzheimer's Disease Neuroimaging Initiative for 708 subjects, 30,000 vertices on the left and right hippocampal surfaces, and 501,584 SNPs. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. Comparative Genomic Analysis of Soybean Flowering Genes

    Science.gov (United States)

    Jung, Chol-Hee; Wong, Chui E.; Singh, Mohan B.; Bhalla, Prem L.

    2012-01-01

    Flowering is an important agronomic trait that determines crop yield. Soybean is a major oilseed legume crop used for human and animal feed. Legumes have unique vegetative and floral complexities. Our understanding of the molecular basis of flower initiation and development in legumes is limited. Here, we address this by using a computational approach to examine flowering regulatory genes in the soybean genome in comparison to the most studied model plant, Arabidopsis. For this comparison, a genome-wide analysis of orthologue groups was performed, followed by an in silico gene expression analysis of the identified soybean flowering genes. Phylogenetic analyses of the gene families highlighted the evolutionary relationships among these candidates. Our study identified key flowering genes in soybean and indicates that the vernalisation and the ambient-temperature pathways seem to be the most variant in soybean. A comparison of the orthologue groups containing flowering genes indicated that, on average, each Arabidopsis flowering gene has 2-3 orthologous copies in soybean. Our analysis highlighted that the CDF3, VRN1, SVP, AP3 and PIF3 genes are paralogue-rich genes in soybean. Furthermore, the genome mapping of the soybean flowering genes showed that these genes are scattered randomly across the genome. A paralogue comparison indicated that the soybean genes comprising the largest orthologue group are clustered in a 1.4 Mb region on chromosome 16 of soybean. Furthermore, a comparison with the undomesticated soybean (Glycine soja) revealed that there are hundreds of SNPs that are associated with putative soybean flowering genes and that there are structural variants that may affect the genes of the light-signalling and ambient-temperature pathways in soybean. Our study provides a framework for the soybean flowering pathway and insights into the relationship and evolution of flowering genes between a short-day soybean and the long-day plant, Arabidopsis. PMID:22679494

  14. Transcription Profiling Demonstrates Epigenetic Control of Non-retroviral RNA Virus-Derived Elements in the Human Genome

    Directory of Open Access Journals (Sweden)

    Kozue Sofuku

    2015-09-01

    Full Text Available Endogenous bornavirus-like nucleoprotein elements (EBLNs are DNA sequences in vertebrate genomes formed by the retrotransposon-mediated integration of ancient bornavirus sequence. Thus, EBLNs evidence a mechanism of retrotransposon-mediated RNA-to-DNA information flow from environment to animals. Although EBLNs are non-transposable, they share some features with retrotransposons. Here, to test whether hosts control the expression of EBLNs similarly to retrotransposons, we profiled the transcription of all Homo sapiens EBLNs (hsEBLN-1 to hsEBLN-7. We could detect transcription of all hsEBLNs in at least one tissue. Among them, hsEBLN-1 is transcribed almost exclusively in the testis. In most tissues, expression from the hsEBLN-1 locus is silenced epigenetically. Finally, we showed the possibility that hsEBLN-1 integration at this locus affects the expression of a neighboring gene. Our results suggest that hosts regulate the expression of endogenous non-retroviral virus elements similarly to how they regulate the expression of retrotransposons, possibly contributing to new transcripts and regulatory complexity to the human genome.

  15. PGSB/MIPS Plant Genome Information Resources and Concepts for the Analysis of Complex Grass Genomes.

    Science.gov (United States)

    Spannagl, Manuel; Bader, Kai; Pfeifer, Matthias; Nussbaumer, Thomas; Mayer, Klaus F X

    2016-01-01

    PGSB (Plant Genome and Systems Biology; formerly MIPS-Munich Institute for Protein Sequences) has been involved in developing, implementing and maintaining plant genome databases for more than a decade. Genome databases and analysis resources have focused on individual genomes and aim to provide flexible and maintainable datasets for model plant genomes as a backbone against which experimental data, e.g., from high-throughput functional genomics, can be organized and analyzed. In addition, genomes from both model and crop plants form a scaffold for comparative genomics, assisted by specialized tools such as the CrowsNest viewer to explore conserved gene order (synteny) between related species on macro- and micro-levels.The genomes of many economically important Triticeae plants such as wheat, barley, and rye present a great challenge for sequence assembly and bioinformatic analysis due to their enormous complexity and large genome size. Novel concepts and strategies have been developed to deal with these difficulties and have been applied to the genomes of wheat, barley, rye, and other cereals. This includes the GenomeZipper concept, reference-guided exome assembly, and "chromosome genomics" based on flow cytometry sorted chromosomes.

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

    NARCIS (Netherlands)

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

    2004-01-01

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

  17. Genome-wide identification of the potato WRKY transcription factor family.

    Science.gov (United States)

    Zhang, Chao; Wang, Dongdong; Yang, Chenghui; Kong, Nana; Shi, Zheng; Zhao, Peng; Nan, Yunyou; Nie, Tengkun; Wang, Ruoqiu; Ma, Haoli; Chen, Qin

    2017-01-01

    WRKY transcription factors play pivotal roles in regulation of stress responses. This study identified 79 WRKY genes in potato (Solanum tuberosum). Based on multiple sequence alignment and phylogenetic relationships, WRKY genes were classified into three major groups. The majority of WRKY genes belonged to Group II (52 StWRKYs), Group III had 14 and Group I consisted of 13. The phylogenetic tree further classified Group II into five sub-groups. All StWRKY genes except StWRKY79 were mapped on potato chromosomes, with eight tandem duplication gene pairs and seven segmental duplication gene pairs found from StWRKY family genes. The expression analysis of 22 StWRKYs showed their differential expression levels under various stress conditions. Cis-element prediction showed that a large number of elements related to drought, heat and salicylic acid were present in the promotor regions of StWRKY genes. The expression analysis indicated that seven StWRKYs seemed to respond to stress (heat, drought and salinity) and salicylic acid treatment. These genes are candidates for abiotic stress signaling for further research.

  18. The genome-wide identification and transcriptional levels of DNA methyltransferases and demethylases in globe artichoke.

    Science.gov (United States)

    Gianoglio, Silvia; Moglia, Andrea; Acquadro, Alberto; Comino, Cinzia; Portis, Ezio

    2017-01-01

    Changes to the cytosine methylation status of DNA, driven by the activity of C5 methyltransferases (C5-MTases) and demethylases, exert an important influence over development, transposon movement, gene expression and imprinting. Three groups of C5-MTase enzymes have been identified in plants, namely MET (methyltransferase 1), CMT (chromomethyltransferases) and DRM (domains rearranged methyltransferases). Here the repertoire of genes encoding C5-MTase and demethylase by the globe artichoke (Cynara cardunculus var. scolymus) is described, based on sequence homology, a phylogenetic analysis and a characterization of their functional domains. A total of ten genes encoding C5-MTase (one MET, five CMTs and four DRMs) and five demethylases was identified. An analysis of their predicted product's protein structure suggested an extensive level of conservation has been retained by the C5-MTases. Transcriptional profiling based on quantitative real time PCR revealed a number of differences between the genes encoding maintenance and de novo methyltransferases, sometimes in a tissue- or development-dependent manner, which implied a degree of functional specialization.

  19. The genome-wide identification and transcriptional levels of DNA methyltransferases and demethylases in globe artichoke.

    Directory of Open Access Journals (Sweden)

    Silvia Gianoglio

    Full Text Available Changes to the cytosine methylation status of DNA, driven by the activity of C5 methyltransferases (C5-MTases and demethylases, exert an important influence over development, transposon movement, gene expression and imprinting. Three groups of C5-MTase enzymes have been identified in plants, namely MET (methyltransferase 1, CMT (chromomethyltransferases and DRM (domains rearranged methyltransferases. Here the repertoire of genes encoding C5-MTase and demethylase by the globe artichoke (Cynara cardunculus var. scolymus is described, based on sequence homology, a phylogenetic analysis and a characterization of their functional domains. A total of ten genes encoding C5-MTase (one MET, five CMTs and four DRMs and five demethylases was identified. An analysis of their predicted product's protein structure suggested an extensive level of conservation has been retained by the C5-MTases. Transcriptional profiling based on quantitative real time PCR revealed a number of differences between the genes encoding maintenance and de novo methyltransferases, sometimes in a tissue- or development-dependent manner, which implied a degree of functional specialization.

  20. Whole genome sequencing and bioinformatics analysis of two Egyptian genomes.

    Science.gov (United States)

    ElHefnawi, Mahmoud; Jeon, Sungwon; Bhak, Youngjune; ElFiky, Asmaa; Horaiz, Ahmed; Jun, JeHoon; Kim, Hyunho; Bhak, Jong

    2018-05-15

    We report two Egyptian male genomes (EGP1 and EGP2) sequenced at ~ 30× sequencing depths. EGP1 had 4.7 million variants, where 198,877 were novel variants while EGP2 had 209,109 novel variants out of 4.8 million variants. The mitochondrial haplogroup of the two individuals were identified to be H7b1 and L2a1c, respectively. We also identified the Y haplogroup of EGP1 (R1b) and EGP2 (J1a2a1a2 > P58 > FGC11). EGP1 had a mutation in the NADH gene of the mitochondrial genome ND4 (m.11778 G > A) that causes Leber's hereditary optic neuropathy. Some SNPs shared by the two genomes were associated with an increased level of cholesterol and triglycerides, probably related with Egyptians obesity. Comparison of these genomes with African and Western-Asian genomes can provide insights on Egyptian ancestry and genetic history. This resource can be used to further understand genomic diversity and functional classification of variants as well as human migration and evolution across Africa and Western-Asia. Copyright © 2017. Published by Elsevier B.V.

  1. Genome-wide comparative analysis of four Indian Drosophila species.

    Science.gov (United States)

    Mohanty, Sujata; Khanna, Radhika

    2017-12-01

    Comparative analysis of multiple genomes of closely or distantly related Drosophila species undoubtedly creates excitement among evolutionary biologists in exploring the genomic changes with an ecology and evolutionary perspective. We present herewith the de novo assembled whole genome sequences of four Drosophila species, D. bipectinata, D. takahashii, D. biarmipes and D. nasuta of Indian origin using Next Generation Sequencing technology on an Illumina platform along with their detailed assembly statistics. The comparative genomics analysis, e.g. gene predictions and annotations, functional and orthogroup analysis of coding sequences and genome wide SNP distribution were performed. The whole genome of Zaprionus indianus of Indian origin published earlier by us and the genome sequences of previously sequenced 12 Drosophila species available in the NCBI database were included in the analysis. The present work is a part of our ongoing genomics project of Indian Drosophila species.

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

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    Shuchi eSmita

    2015-12-01

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

  3. Genome-wide expression profiling shows transcriptional reprogramming in Fusarium graminearum by Fusarium graminearum virus 1-DK21 infection

    Directory of Open Access Journals (Sweden)

    Cho Won

    2012-05-01

    Full Text Available Abstract Background Fusarium graminearum virus 1 strain-DK21 (FgV1-DK21 is a mycovirus that confers hypovirulence to F. graminearum, which is the primary phytopathogenic fungus that causes Fusarium head blight (FHB disease in many cereals. Understanding the interaction between mycoviruses and plant pathogenic fungi is necessary for preventing damage caused by F. graminearum. Therefore, we investigated important cellular regulatory processes in a host containing FgV1-DK21 as compared to an uninfected parent using a transcriptional approach. Results Using a 3′-tiling microarray covering all known F. graminearum genes, we carried out genome-wide expression analyses of F. graminearum at two different time points. At the early point of growth of an infected strain as compared to an uninfected strain, genes associated with protein synthesis, including ribosome assembly, nucleolus, and ribosomal RNA processing, were significantly up-regulated. In addition, genes required for transcription and signal transduction, including fungal-specific transcription factors and cAMP signaling, respectively, were actively up-regulated. In contrast, genes involved in various metabolic pathways, particularly in producing carboxylic acids, aromatic amino acids, nitrogen compounds, and polyamines, showed dramatic down-regulation at the early time point. Moreover, genes associated with transport systems localizing to transmembranes were down-regulated at both time points. Conclusion This is the first report of global change in the prominent cellular pathways in the Fusarium host containing FgV1-DK21. The significant increase in transcripts for transcription and translation machinery in fungal host cells seems to be related to virus replication. In addition, significant down-regulation of genes required for metabolism and transporting systems in a fungal host containing the virus appears to be related to the host defense mechanism and fungal virulence. Taken together

  4. Functional genomic analysis of C. elegans molting.

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    Alison R Frand

    2005-10-01

    Full Text Available Although the molting cycle is a hallmark of insects and nematodes, neither the endocrine control of molting via size, stage, and nutritional inputs nor the enzymatic mechanism for synthesis and release of the exoskeleton is well understood. Here, we identify endocrine and enzymatic regulators of molting in C. elegans through a genome-wide RNA-interference screen. Products of the 159 genes discovered include annotated transcription factors, secreted peptides, transmembrane proteins, and extracellular matrix enzymes essential for molting. Fusions between several genes and green fluorescent protein show a pulse of expression before each molt in epithelial cells that synthesize the exoskeleton, indicating that the corresponding proteins are made in the correct time and place to regulate molting. We show further that inactivation of particular genes abrogates expression of the green fluorescent protein reporter genes, revealing regulatory networks that might couple the expression of genes essential for molting to endocrine cues. Many molting genes are conserved in parasitic nematodes responsible for human disease, and thus represent attractive targets for pesticide and pharmaceutical development.

  5. ATXN1L, CIC, and ETS Transcription Factors Modulate Sensitivity to MAPK Pathway Inhibition | Office of Cancer Genomics

    Science.gov (United States)

    Intrinsic resistance and RTK-RAS-MAPK pathway reactivation has limited the effectiveness of MEK and RAF inhibitors (MAPKi) in RAS- and RAF-mutant cancers. To identify genes that modulate sensitivity to MAPKi, we performed genome-scale CRISPR-Cas9 loss-of-function screens in two KRAS mutant pancreatic cancer cell lines treated with the MEK1/2 inhibitor trametinib. Loss of CIC, a transcriptional repressor of ETV1, ETV4, and ETV5, promoted survival in the setting of MAPKi in cancer cells derived from several lineages.

  6. Genome-wide identification and characterization of GRAS transcription factors in tomato (Solanum lycopersicum).

    Science.gov (United States)

    Niu, Yiling; Zhao, Tingting; Xu, Xiangyang; Li, Jingfu

    2017-01-01

    Solanum lycopersicum , belonging to Solanaceae, is one of the commonly used model plants. The GRAS genes are transcriptional regulators, which play a significant role in plant growth and development, and the functions of several GRAS genes have been recognized, such as, axillary shoot meristem formation, radial root patterning, phytohormones (gibberellins) signal transduction, light signaling, and abiotic/biotic stress; however, only a few of these were identified and functionally characterized. In this study, a gene family was analyzed comprehensively with respect to phylogeny, gene structure, chromosomal localization, and expression pattern; the 54 GRAS members were screened from tomato by bioinformatics for the first time. The GRAS genes among tomato, Arabidopsis , rice, and grapevine were rebuilt to form a phylogenomic tree, which was divided into ten groups according to the previous classification of Arabidopsis and rice. A multiple sequence alignment exhibited the typical GRAS domain and conserved motifs similar to other gene families. Both the segmental and tandem duplications contributed significantly to the expansion and evolution of the GRAS gene family in tomato; the expression patterns across a variety of tissues and biotic conditions revealed potentially different functions of GRAS genes in tomato development and stress responses. Altogether, this study provides valuable information and robust candidate genes for future functional analysis for improving the resistance of tomato growth.

  7. Determining physical constraints in transcriptional initiationcomplexes using DNA sequence analysis

    Energy Technology Data Exchange (ETDEWEB)

    Shultzaberger, Ryan K.; Chiang, Derek Y.; Moses, Alan M.; Eisen,Michael B.

    2007-07-01

    Eukaryotic gene expression is often under the control ofcooperatively acting transcription factors whose binding is limited bystructural constraints. By determining these structural constraints, wecan understand the "rules" that define functional cooperativity.Conversely, by understanding the rules of binding, we can inferstructural characteristics. We have developed an information theory basedmethod for approximating the physical limitations of cooperativeinteractions by comparing sequence analysis to microarray expressiondata. When applied to the coordinated binding of the sulfur amino acidregulatory protein Met4 by Cbf1 and Met31, we were able to create acombinatorial model that can correctly identify Met4 regulatedgenes.

  8. Genome-wide analysis of Pax8 binding provides new insights into thyroid functions

    Directory of Open Access Journals (Sweden)

    Ruiz-Llorente Sergio

    2012-04-01

    Full Text Available Abstract Background The transcription factor Pax8 is essential for the differentiation of thyroid cells. However, there are few data on genes transcriptionally regulated by Pax8 other than thyroid-related genes. To better understand the role of Pax8 in the biology of thyroid cells, we obtained transcriptional profiles of Pax8-silenced PCCl3 thyroid cells using whole genome expression arrays and integrated these signals with global cis-regulatory sequencing studies performed by ChIP-Seq analysis Results Exhaustive analysis of Pax8 immunoprecipitated peaks demonstrated preferential binding to intragenic regions and CpG-enriched islands, which suggests a role of Pax8 in transcriptional regulation of orphan CpG regions. In addition, ChIP-Seq allowed us to identify Pax8 partners, including proteins involved in tertiary DNA structure (CTCF and chromatin remodeling (Sp1, and these direct transcriptional interactions were confirmed in vivo. Moreover, both factors modulate Pax8-dependent transcriptional activation of the sodium iodide symporter (Nis gene promoter. We ultimately combined putative and novel Pax8 binding sites with actual target gene expression regulation to define Pax8-dependent genes. Functional classification suggests that Pax8-regulated genes may be directly involved in important processes of thyroid cell function such as cell proliferation and differentiation, apoptosis, cell polarity, motion and adhesion, and a plethora of DNA/protein-related processes. Conclusion Our study provides novel insights into the role of Pax8 in thyroid biology, exerted through transcriptional regulation of important genes involved in critical thyrocyte processes. In addition, we found new transcriptional partners of Pax8, which functionally cooperate with Pax8 in the regulation of thyroid gene transcription. Besides, our data demonstrate preferential location of Pax8 in non-promoter CpG regions. These data point to an orphan CpG island-mediated mechanism

  9. Identification and analysis of functional elements in 1% of the human genome by the ENCODE pilot project.

    Science.gov (United States)

    Birney, Ewan; Stamatoyannopoulos, John A; Dutta, Anindya; Guigó, Roderic; Gingeras, Thomas R; Margulies, Elliott H; Weng, Zhiping; Snyder, Michael; Dermitzakis, Emmanouil T; Thurman, Robert E; Kuehn, Michael S; Taylor, Christopher M; Neph, Shane; Koch, Christoph M; Asthana, Saurabh; Malhotra, Ankit; Adzhubei, Ivan; Greenbaum, Jason A; Andrews, Robert M; Flicek, Paul; Boyle, Patrick J; Cao, Hua; Carter, Nigel P; Clelland, Gayle K; Davis, Sean; Day, Nathan; Dhami, Pawandeep; Dillon, Shane C; Dorschner, Michael O; Fiegler, Heike; Giresi, Paul G; Goldy, Jeff; Hawrylycz, Michael; Haydock, Andrew; Humbert, Richard; James, Keith D; Johnson, Brett E; Johnson, Ericka M; Frum, Tristan T; Rosenzweig, Elizabeth R; Karnani, Neerja; Lee, Kirsten; Lefebvre, Gregory C; Navas, Patrick A; Neri, Fidencio; Parker, Stephen C J; Sabo, Peter J; Sandstrom, Richard; Shafer, Anthony; Vetrie, David; Weaver, Molly; Wilcox, Sarah; Yu, Man; Collins, Francis S; Dekker, Job; Lieb, Jason D; Tullius, Thomas D; Crawford, Gregory E; Sunyaev, Shamil; Noble, William S; Dunham, Ian; Denoeud, France; Reymond, Alexandre; Kapranov, Philipp; Rozowsky, Joel; Zheng, Deyou; Castelo, Robert; Frankish, Adam; Harrow, Jennifer; Ghosh, Srinka; Sandelin, Albin; Hofacker, Ivo L; Baertsch, Robert; Keefe, Damian; Dike, Sujit; Cheng, Jill; Hirsch, Heather A; Sekinger, Edward A; Lagarde, Julien; Abril, Josep F; Shahab, Atif; Flamm, Christoph; Fried, Claudia; Hackermüller, Jörg; Hertel, Jana; Lindemeyer, Manja; Missal, Kristin; Tanzer, Andrea; Washietl, Stefan; Korbel, Jan; Emanuelsson, Olof; Pedersen, Jakob S; Holroyd, Nancy; Taylor, Ruth; Swarbreck, David; Matthews, Nicholas; Dickson, Mark C; Thomas, Daryl J; Weirauch, Matthew T; Gilbert, James; Drenkow, Jorg; Bell, Ian; Zhao, XiaoDong; Srinivasan, K G; Sung, Wing-Kin; Ooi, Hong Sain; Chiu, Kuo Ping; Foissac, Sylvain; Alioto, Tyler; Brent, Michael; Pachter, Lior; Tress, Michael L; Valencia, Alfonso; Choo, Siew Woh; Choo, Chiou Yu; Ucla, Catherine; Manzano, Caroline; Wyss, Carine; Cheung, Evelyn; Clark, Taane G; Brown, James B; Ganesh, Madhavan; Patel, Sandeep; Tammana, Hari; Chrast, Jacqueline; Henrichsen, Charlotte N; Kai, Chikatoshi; Kawai, Jun; Nagalakshmi, Ugrappa; Wu, Jiaqian; Lian, Zheng; Lian, Jin; Newburger, Peter; Zhang, Xueqing; Bickel, Peter; Mattick, John S; Carninci, Piero; Hayashizaki, Yoshihide; Weissman, Sherman; Hubbard, Tim; Myers, Richard M; Rogers, Jane; Stadler, Peter F; Lowe, Todd M; Wei, Chia-Lin; Ruan, Yijun; Struhl, Kevin; Gerstein, Mark; Antonarakis, Stylianos E; Fu, Yutao; Green, Eric D; Karaöz, Ulaş; Siepel, Adam; Taylor, James; Liefer, Laura A; Wetterstrand, Kris A; Good, Peter J; Feingold, Elise A; Guyer, Mark S; Cooper, Gregory M; Asimenos, George; Dewey, Colin N; Hou, Minmei; Nikolaev, Sergey; Montoya-Burgos, Juan I; Löytynoja, Ari; Whelan, Simon; Pardi, Fabio; Massingham, Tim; Huang, Haiyan; Zhang, Nancy R; Holmes, Ian; Mullikin, James C; Ureta-Vidal, Abel; Paten, Benedict; Seringhaus, Michael; Church, Deanna; Rosenbloom, Kate; Kent, W James; Stone, Eric A; Batzoglou, Serafim; Goldman, Nick; Hardison, Ross C; Haussler, David; Miller, Webb; Sidow, Arend; Trinklein, Nathan D; Zhang, Zhengdong D; Barrera, Leah; Stuart, Rhona; King, David C; Ameur, Adam; Enroth, Stefan; Bieda, Mark C; Kim, Jonghwan; Bhinge, Akshay A; Jiang, Nan; Liu, Jun; Yao, Fei; Vega, Vinsensius B; Lee, Charlie W H; Ng, Patrick; Shahab, Atif; Yang, Annie; Moqtaderi, Zarmik; Zhu, Zhou; Xu, Xiaoqin; Squazzo, Sharon; Oberley, Matthew J; Inman, David; Singer, Michael A; Richmond, Todd A; Munn, Kyle J; Rada-Iglesias, Alvaro; Wallerman, Ola; Komorowski, Jan; Fowler, Joanna C; Couttet, Phillippe; Bruce, Alexander W; Dovey, Oliver M; Ellis, Peter D; Langford, Cordelia F; Nix, David A; Euskirchen, Ghia; Hartman, Stephen; Urban, Alexander E; Kraus, Peter; Van Calcar, Sara; Heintzman, Nate; Kim, Tae Hoon; Wang, Kun; Qu, Chunxu; Hon, Gary; Luna, Rosa; Glass, Christopher K; Rosenfeld, M Geoff; Aldred, Shelley Force; Cooper, Sara J; Halees, Anason; Lin, Jane M; Shulha, Hennady P; Zhang, Xiaoling; Xu, Mousheng; Haidar, Jaafar N S; Yu, Yong; Ruan, Yijun; Iyer, Vishwanath R; Green, Roland D; Wadelius, Claes; Farnham, Peggy J; Ren, Bing; Harte, Rachel A; Hinrichs, Angie S; Trumbower, Heather; Clawson, Hiram; Hillman-Jackson, Jennifer; Zweig, Ann S; Smith, Kayla; Thakkapallayil, Archana; Barber, Galt; Kuhn, Robert M; Karolchik, Donna; Armengol, Lluis; Bird, Christine P; de Bakker, Paul I W; Kern, Andrew D; Lopez-Bigas, Nuria; Martin, Joel D; Stranger, Barbara E; Woodroffe, Abigail; Davydov, Eugene; Dimas, Antigone; Eyras, Eduardo; Hallgrímsdóttir, Ingileif B; Huppert, Julian; Zody, Michael C; Abecasis, Gonçalo R; Estivill, Xavier; Bouffard, Gerard G; Guan, Xiaobin; Hansen, Nancy F; Idol, Jacquelyn R; Maduro, Valerie V B; Maskeri, Baishali; McDowell, Jennifer C; Park, Morgan; Thomas, Pamela J; Young, Alice C; Blakesley, Robert W; Muzny, Donna M; Sodergren, Erica; Wheeler, David A; Worley, Kim C; Jiang, Huaiyang; Weinstock, George M; Gibbs, Richard A; Graves, Tina; Fulton, Robert; Mardis, Elaine R; Wilson, Richard K; Clamp, Michele; Cuff, James; Gnerre, Sante; Jaffe, David B; Chang, Jean L; Lindblad-Toh, Kerstin; Lander, Eric S; Koriabine, Maxim; Nefedov, Mikhail; Osoegawa, Kazutoyo; Yoshinaga, Yuko; Zhu, Baoli; de Jong, Pieter J

    2007-06-14

    We report the generation and analysis of functional data from multiple, diverse experiments performed on a targeted 1% of the human genome as part of the pilot phase of the ENCODE Project. These data have been further integrated and augmented by a number of evolutionary and computational analyses. Together, our results advance the collective knowledge about human genome function in several major areas. First, our studies provide convincing evidence that the genome is pervasively transcribed, such that the majority of its bases can be found in primary transcripts, including non-protein-coding transcripts, and those that extensively overlap one another. Second, systematic examination of transcriptional regulation has yielded new understanding about transcription start sites, including their relationship to specific regulatory sequences and features of chromatin accessibility and histone modification. Third, a more sophisticated view of chromatin structure has emerged, including its inter-relationship with DNA replication and transcriptional regulation. Finally, integration of these new sources of information, in particular with respect to mammalian evolution based on inter- and intra-species sequence comparisons, has yielded new mechanistic and evolutionary insights concerning the functional landscape of the human genome. Together, these studies are defining a path for pursuit of a more comprehensive characterization of human genome function.

  10. Genome-wide transcriptional response of silkworm (Bombyx mori to infection by the microsporidian Nosema bombycis.

    Directory of Open Access Journals (Sweden)

    Zhengang Ma

    Full Text Available Microsporidia have attracted much attention because they infect a variety of species ranging from protists to mammals, including immunocompromised patients with AIDS or cancer. Aside from the study on Nosema ceranae, few works have focused on elucidating the mechanism in host response to microsporidia infection. Nosema bombycis is a pathogen of silkworm pébrine that causes great economic losses to the silkworm industry. Detailed understanding of the host (Bombyx mori response to infection by N. bombycis is helpful for prevention of this disease. A genome-wide survey of the gene expression profile at 2, 4, 6 and 8 days post-infection by N. bombycis was performed and results showed that 64, 244, 1,328, 1,887 genes were induced, respectively. Up to 124 genes, which are involved in basal metabolism pathways, were modulated. Notably, B. mori genes that play a role in juvenile hormone synthesis and metabolism pathways were induced, suggesting that the host may accumulate JH as a response to infection. Interestingly, N. bombycis can inhibit the silkworm serine protease cascade melanization pathway in hemolymph, which may be due to the secretion of serpins in the microsporidia. N. bombycis also induced up-regulation of several cellular immune factors, in which CTL11 has been suggested to be involved in both spore recognition and immune signal transduction. Microarray and real-time PCR analysis indicated the activation of silkworm Toll and JAK/STAT pathways. The notable up-regulation of antimicrobial peptides, including gloverins, lebocins and moricins, strongly indicated that antimicrobial peptide defense mechanisms were triggered to resist the invasive microsporidia. An analysis of N. bombycis-specific response factors suggested their important roles in anti-microsporidia defense. Overall, this study primarily provides insight into the potential molecular mechanisms for the host-parasite interaction between B. mori and N. bombycis and may provide a

  11. Integration of transcript expression, copy number and LOH analysis of infiltrating ductal carcinoma of the breast

    Directory of Open Access Journals (Sweden)

    Hawthorn Lesleyann

    2010-08-01

    Full Text Available Abstract Background A major challenge in the interpretation of genomic profiling data generated from breast cancer samples is the identification of driver genes as distinct from bystander genes which do not impact tumorigenesis. One way to assess the relative importance of alterations in the transcriptome profile is to combine parallel analyses that assess changes in the copy number alterations (CNAs. This integrated analysis permits the identification of genes with altered expression that map within specific chromosomal regions which demonstrate copy number alterations, providing a mechanistic approach to identify the 'driver genes'. Methods We have performed whole genome analysis of CNAs using the Affymetrix 250K Mapping array on 22 infiltrating ductal carcinoma samples (IDCs. Analysis of transcript expression alterations was performed using the Affymetrix U133 Plus2.0 array on 16 IDC samples. Fourteen IDC samples were analyzed using both platforms and the data integrated. We also incorporated data from loss of heterozygosity (LOH analysis to identify genes showing altered expression in LOH regions. Results Common chromosome gains and amplifications were identified at 1q21.3, 6p21.3, 7p11.2-p12.1, 8q21.11 and 8q24.3. A novel amplicon was identified at 5p15.33. Frequent losses were found at 1p36.22, 8q23.3, 11p13, 11q23, and 22q13. Over 130 genes were identified with concurrent increases or decreases in expression that mapped to these regions of copy number alterations. LOH analysis revealed three tumors with whole chromosome or p arm allelic loss of chromosome 17. Genes were identified that mapped to copy neutral LOH regions. LOH with accompanying copy loss was detected on Xp24 and Xp25 and genes mapping to these regions with decreased expression were identified. Gene expression data highlighted the PPARα/RXRα Activation Pathway as down-regulated in the tumor samples. Conclusion We have demonstrated the utility of the application of

  12. Calcium-Release Channels in Paramecium. Genomic Expansion, Differential Positioning and Partial Transcriptional Elimination

    Science.gov (United States)

    Ladenburger, Eva-Maria; Plattner, Helmut

    2011-01-01

    The release of Ca2+ from internal stores is a major source of signal Ca2+ in almost all cell types. The internal Ca2+ pools are activated via two main families of intracellular Ca2+-release channels, the ryanodine and the inositol 1,4,5-trisphosphate (InsP3) receptors. Among multicellular organisms these channel types are ubiquitous, whereas in most unicellular eukaryotes the identification of orthologs is impaired probably due to evolutionary sequence divergence. However, the ciliated protozoan Paramecium allowed us to prognosticate six groups, with a total of 34 genes, encoding proteins with characteristics typical of InsP3 and ryanodine receptors by BLAST search of the Paramecium database. We here report that these Ca2+-release channels may display all or only some of the characteristics of canonical InsP3 and ryanodine receptors. In all cases, prediction methods indicate the presence of six trans-membrane regions in the C-terminal domains, thus corresponding to canonical InsP3 receptors, while a sequence homologous to the InsP3-binding domain is present only in some types. Only two types have been analyzed in detail previously. We now show, by using antibodies and eventually by green fluorescent protein labeling, that the members of all six groups localize to distinct organelles known to participate in vesicle trafficking and, thus, may provide Ca2+ for local membrane-membrane interactions. Whole genome duplication can explain radiation within the six groups. Comparative and evolutionary evaluation suggests derivation from a common ancestor of canonical InsP3 and ryanodine receptors. With one group we could ascertain, to our knowledge for the first time, aberrant splicing in one thoroughly analyzed Paramecium gene. This yields truncated forms and, thus, may indicate a way to pseudogene formation. No comparable analysis is available for any other, free-living or parasitic/pathogenic protozoan. PMID:22102876

  13. Calcium-release channels in paramecium. Genomic expansion, differential positioning and partial transcriptional elimination.

    Directory of Open Access Journals (Sweden)

    Eva-Maria Ladenburger

    Full Text Available The release of Ca²⁺ from internal stores is a major source of signal Ca²⁺ in almost all cell types. The internal Ca²⁺ pools are activated via two main families of intracellular Ca²⁺-release channels, the ryanodine and the inositol 1,4,5-trisphosphate (InsP₃ receptors. Among multicellular organisms these channel types are ubiquitous, whereas in most unicellular eukaryotes the identification of orthologs is impaired probably due to evolutionary sequence divergence. However, the ciliated protozoan Paramecium allowed us to prognosticate six groups, with a total of 34 genes, encoding proteins with characteristics typical of InsP₃ and ryanodine receptors by BLAST search of the Paramecium database. We here report that these Ca²⁺-release channels may display all or only some of the characteristics of canonical InsP₃ and ryanodine receptors. In all cases, prediction methods indicate the presence of six trans-membrane regions in the C-terminal domains, thus corresponding to canonical InsP₃ receptors, while a sequence homologous to the InsP₃-binding domain is present only in some types. Only two types have been analyzed in detail previously. We now show, by using antibodies and eventually by green fluorescent protein labeling, that the members of all six groups localize to distinct organelles known to participate in vesicle trafficking and, thus, may provide Ca²⁺ for local membrane-membrane interactions. Whole genome duplication can explain radiation within the six groups. Comparative and evolutionary evaluation suggests derivation from a common ancestor of canonical InsP₃ and ryanodine receptors. With one group we could ascertain, to our knowledge for the first time, aberrant splicing in one thoroughly analyzed Paramecium gene. This yields truncated forms and, thus, may indicate a way to pseudogene formation. No comparable analysis is available for any other, free-living or parasitic/pathogenic protozoan.

  14. IMG: the integrated microbial genomes database and comparative analysis system

    Science.gov (United States)

    Markowitz, Victor M.; Chen, I-Min A.; Palaniappan, Krishna; Chu, Ken; Szeto, Ernest; Grechkin, Yuri; Ratner, Anna; Jacob, Biju; Huang, Jinghua; Williams, Peter; Huntemann, Marcel; Anderson, Iain; Mavromatis, Konstantinos; Ivanova, Natalia N.; Kyrpides, Nikos C.

    2012-01-01

    The Integrated Microbial Genomes (IMG) system serves as a community resource for comparative analysis of publicly available genomes in a comprehensive integrated context. IMG integrates publicly available draft and complete genomes from all three domains of life with a large number of plasmids and viruses. IMG provides tools and viewers for analyzing and reviewing the annotations of genes and genomes in a comparative context. IMG's data content and analytical capabilities have been continuously extended through regular updates since its first release in March 2005. IMG is available at http://img.jgi.doe.gov. Companion IMG systems provide support for expert review of genome annotations (IMG/ER: http://img.jgi.doe.gov/er), teaching courses and training in microbial genome analysis (IMG/EDU: http://img.jgi.doe.gov/edu) and analysis of genomes related to the Human Microbiome Project (IMG/HMP: http://www.hmpdacc-resources.org/img_hmp). PMID:22194640

  15. Comprehensive analysis of the specificity of transcription activator-like effector nucleases

    DEFF Research Database (Denmark)

    Juillerat, Alexandre; Dubois, Gwendoline; Valton, Julien

    2014-01-01

    A key issue when designing and using DNA-targeting nucleases is specificity. Ideally, an optimal DNA-targeting tool has only one recognition site within a genomic sequence. In practice, however, almost all designer nucleases available today can accommodate one to several mutations within...... their target site. The ability to predict the specificity of targeting is thus highly desirable. Here, we describe the first comprehensive experimental study focused on the specificity of the four commonly used repeat variable diresidues (RVDs; NI:A, HD:C, NN:G and NG:T) incorporated in transcription activator......-like effector nucleases (TALEN). The analysis of >15 500 unique TALEN/DNA cleavage profiles allowed us to monitor the specificity gradient of the RVDs along a TALEN/DNA binding array and to present a specificity scoring matrix for RVD/nucleotide association. Furthermore, we report that TALEN can only...

  16. Gene expression of herpes simplex virus. II. Uv radiological analysis of viral transcription units

    International Nuclear Information System (INIS)

    Millette, R. L.; Klaiber, R.

    1980-01-01

    The transcriptional organization of the genome of herpes simplex virus type 1 was analyzed by measuring the sensitivity of viral polypeptide synthesis to uv irradiation of the infecting virus. Herpes simplex virus type 1 was irradiated with various doses of uv light and used to infect xeroderma pigmentosum fibroblasts. Immediate early transcription units were analyzed by having cycloheximide present throughout the period of infection, removing the drug at 8 h postinfection, and pulse-labeling proteins with [355]methionine. Delayed early transcription units were analyzed in similar studies by having 9-beta-D-arabinofuranosyladenine present during the experiment to block replication of the input irradiated genome. The results indicate that none of the immediate early genes analyzed can be cotranscribed, whereas some of the delayed early genes might be cotranscribed. No evidence was found for the existence of large, multigene transcription units

  17. Big Data Analysis of Human Genome Variations

    KAUST Repository

    Gojobori, Takashi

    2016-01-01

    Since the human genome draft sequence was in public for the first time in 2000, genomic analyses have been intensively extended to the population level. The following three international projects are good examples for large-scale studies of human

  18. Cloning, nucleotide sequence and transcriptional analysis of the uvrA gene from Neisseria gonorrhoeae

    International Nuclear Information System (INIS)

    Black, C.G.; Fyfe, J.A.M.; Davies, J.K.

    1997-01-01

    A recombinant plasmid capable of restoring UV resistance to an Escherichia coli uvrA mutant was isolated from a genomic library of Neisseria gonorrhoeae. Sequence analysis revealed an open reading frame whose deduced amino acid sequence displayed significant similarity to those of the UvrA proteins of other bacterial species. A second open reading frame (ORF259) was identified upstream from, and in the opposite orientation to the gonococcal uvrA gene. Transcriptional fusions between portions of the gonococcal uvrA upstream region and a reporter gene were used to localise promoter activity in both E. coli and N. gonorrhoeae. The transcriptional starting points of uvrA and ORF259 were mapped in E. coli by primer extension analysis, and corresponding σ 70 promoters were identified. The arrangement of the uvrA-ORF259 intergenic region is similar to that of the gonococcal recA-aroD intergenic region. Both contain inverted copies of the 10 bp neisserial DNA uptake sequence situated between divergently transcribed genes. However, there is no evidence that either the uptake sequence or the proximity of the promoters influences expression of these genes. (author)

  19. Genomic profiling of rice sperm cell transcripts reveals conserved and distinct elements in the flowering plant male germ lineage.

    Science.gov (United States)

    Russell, Scott D; Gou, Xiaoping; Wong, Chui E; Wang, Xinkun; Yuan, Tong; Wei, Xiaoping; Bhalla, Prem L; Singh, Mohan B

    2012-08-01

    Genomic assay of sperm cell RNA provides insight into functional control, modes of regulation, and contributions of male gametes to double fertilization. Sperm cells of rice (Oryza sativa) were isolated from field-grown, disease-free plants and RNA was processed for use with the full-genome Affymetrix microarray. Comparison with Gene Expression Omnibus (GEO) reference arrays confirmed expressionally distinct gene profiles. A total of 10,732 distinct gene sequences were detected in sperm cells, of which 1668 were not expressed in pollen or seedlings. Pathways enriched in male germ cells included ubiquitin-mediated pathways, pathways involved in chromatin modeling including histones, histone modification and nonhistone epigenetic modification, and pathways related to RNAi and gene silencing. Genome-wide expression patterns in angiosperm sperm cells indicate common and divergent themes in the male germline that appear to be largely self-regulating through highly up-regulated chromatin modification pathways. A core of highly conserved genes appear common to all sperm cells, but evidence is still emerging that another class of genes have diverged in expression between monocots and dicots since their divergence. Sperm cell transcripts present at fusion may be transmitted through plasmogamy during double fertilization to effect immediate post-fertilization expression of early embryo and (or) endosperm development. © 2012 The Authors. New Phytologist © 2012 New Phytologist Trust.

  20. Genome editing of bread wheat using biolistic delivery of CRISPR/Cas9 in vitro transcripts or ribonucleoproteins.

    Science.gov (United States)

    Liang, Zhen; Chen, Kunling; Zhang, Yi; Liu, Jinxing; Yin, Kangquan; Qiu, Jin-Long; Gao, Caixia

    2018-03-01

    This protocol is an extension to: Nat. Protoc. 9, 2395-2410 (2014); doi:10.1038/nprot.2014.157; published online 18 September 2014In recent years, CRISPR/Cas9 has emerged as a powerful tool for improving crop traits. Conventional plant genome editing mainly relies on plasmid-carrying cassettes delivered by Agrobacterium or particle bombardment. Here, we describe DNA-free editing of bread wheat by delivering in vitro transcripts (IVTs) or ribonucleoprotein complexes (RNPs) of CRISPR/Cas9 by particle bombardment. This protocol serves as an extension of our previously published protocol on genome editing in bread wheat using CRISPR/Cas9 plasmids delivered by particle bombardment. The methods we describe not only eliminate random integration of CRISPR/Cas9 into genomic DNA, but also reduce off-target effects. In this protocol extension article, we present detailed protocols for preparation of IVTs and RNPs; validation by PCR/restriction enzyme (RE) and next-generation sequencing; delivery by biolistics; and recovery of mutants and identification of mutants by pooling methods and Sanger sequencing. To use these protocols, researchers should have basic skills and experience in molecular biology and biolistic transformation. By using these protocols, plants edited without the use of any foreign DNA can be generated and identified within 9-11 weeks.

  1. Genome-wide strategies identify downstream target genes of chick connective tissue-associated transcription factors.

    Science.gov (United States)

    Orgeur, Mickael; Martens, Marvin; Leonte, Georgeta; Nassari, Sonya; Bonnin, Marie-Ange; Börno, Stefan T; Timmermann, Bernd; Hecht, Jochen; Duprez, Delphine; Stricker, Sigmar

    2018-03-29

    Connective tissues support organs and play crucial roles in development, homeostasis and fibrosis, yet our understanding of their formation is still limited. To gain insight into the molecular mechanisms of connective tissue specification, we selected five zinc-finger transcription factors - OSR1, OSR2, EGR1, KLF2 and KLF4 - based on their expression patterns and/or known involvement in connective tissue subtype differentiation. RNA-seq and ChIP-seq profiling of chick limb micromass cultures revealed a set of common genes regulated by all five transcription factors, which we describe as a connective tissue core expression set. This common core was enriched with genes associated with axon guidance and myofibroblast signature, including fibrosis-related genes. In addition, each transcription factor regulated a specific set of signalling molecules and extracellular matrix components. This suggests a concept whereby local molecular niches can be created by the expression of specific transcription factors impinging on the specification of local microenvironments. The regulatory network established here identifies common and distinct molecular signatures of limb connective tissue subtypes, provides novel insight into the signalling pathways governing connective tissue specification, and serves as a resource for connective tissue development. © 2018. Published by The Company of Biologists Ltd.

  2. Genome-wide transcriptional responses to carbon starvation in nongrowing Lactococcus lactis

    NARCIS (Netherlands)

    Ercan, O.; Wels, M.; Smid, E.J.; Kleerebezem, M.

    2015-01-01

    This paper describes the transcriptional adaptations of nongrowing, retentostat cultures of Lactococcus lactis to starvation. Near-zero-growth cultures (µ = 0.0001 h-1) obtained by extended retentostat cultivation were exposed to starvation by termination of the medium supply for 24 h, followed by a

  3. Genome-wide organization and expression profiling of the R2R3-MYB transcription factor family in pineapple (Ananas comosus).

    Science.gov (United States)

    Liu, Chaoyang; Xie, Tao; Chen, Chenjie; Luan, Aiping; Long, Jianmei; Li, Chuhao; Ding, Yaqi; He, Yehua

    2017-07-01

    The MYB proteins comprise one of the largest families of plant transcription factors, which are involved in various plant physiological and biochemical processes. Pineapple (Ananas comosus) is one of three most important tropical fruits worldwide. The completion of pineapple genome sequencing provides a great opportunity to investigate the organization and evolutionary traits of pineapple MYB genes at the genome-wide level. In the present study, a total of 94 pineapple R2R3-MYB genes were identified and further phylogenetically classified into 26 subfamilies, as supported by the conserved gene structures and motif composition. Collinearity analysis indicated that the segmental duplication events played a crucial role in the expansion of pineapple MYB gene family. Further comparative phylogenetic analysis suggested that there have been functional divergences of MYB gene family during plant evolution. RNA-seq data from different tissues and developmental stages revealed distinct temporal and spatial expression profiles of the AcMYB genes. Further quantitative expression analysis showed the specific expression patterns of the selected putative stress-related AcMYB genes in response to distinct abiotic stress and hormonal treatments. The comprehensive expression analysis of the pineapple MYB genes, especially the tissue-preferential and stress-responsive genes, could provide valuable clues for further function characterization. In this work, we systematically identified AcMYB genes by analyzing the pineapple genome sequence using a set of bioinformatics approaches. Our findings provide a global insight into the organization, phylogeny and expression patterns of the pineapple R2R3-MYB genes, and hence contribute to the greater understanding of their biological roles in pineapple.

  4. The complete mitochondrial genome of Gossypium hirsutum and evolutionary analysis of higher plant mitochondrial genomes.

    Science.gov (United States)

    Liu, Guozheng; Cao, Dandan; Li, Shuangshuang; Su, Aiguo; Geng, Jianing; Grover, Corrinne E; Hu, Songnian; Hua, Jinping

    2013-01-01

    Mitochondria are the main manufacturers of cellular ATP in eukaryotes. The plant mitochondrial genome contains large number of foreign DNA and repeated sequences undergone frequently intramolecular recombination. Upland Cotton (Gossypium hirsutum L.) is one of the main natural fiber crops and also an important oil-producing plant in the world. Sequencing of the cotton mitochondrial (mt) genome could be helpful for the evolution research of plant mt genomes. We utilized 454 technology for sequencing and combined with Fosmid library of the Gossypium hirsutum mt genome screening and positive clones sequencing and conducted a series of evolutionary analysis on Cycas taitungensis and 24 angiosperms mt genomes. After data assembling and contigs joining, the complete mitochondrial genome sequence of G. hirsutum was obtained. The completed G.hirsutum mt genome is 621,884 bp in length, and contained 68 genes, including 35 protein genes, four rRNA genes and 29 tRNA genes. Five gene clusters are found conserved in all plant mt genomes; one and four clusters are specifically conserved in monocots and dicots, respectively. Homologous sequences are distributed along the plant mt genomes and species closely related share the most homologous sequences. For species that have both mt and chloroplast genome sequences available, we checked the location of cp-like migration and found several fragments closely linked with mitochondrial genes. The G. hirsutum mt genome possesses most of the common characters of higher plant mt genomes. The existence of syntenic gene clusters, as well as the conservation of some intergenic sequences and genic content among the plant mt genomes suggest that evolution of mt genomes is consistent with plant taxonomy but independent among different species.

  5. Phylogenomic Analysis and Dynamic Evolution of Chloroplast Genomes in Salicaceae

    Directory of Open Access Journals (Sweden)

    Yuan Huang

    2017-06-01

    Full Text Available Chloroplast genomes of plants are highly conserved in both gene order and gene content. Analysis of the whole chloroplast genome is known to provide much more informative DNA sites and thus generates high resolution for plant phylogenies. Here, we report the complete chloroplast genomes of three Salix species in family Salicaceae. Phylogeny of Salicaceae inferred from complete chloroplast genomes is generally consistent with previous studies but resolved with higher statistical support. Incongruences of phylogeny, however, are observed in genus Populus, which most likely results from homoplasy. By comparing three Salix chloroplast genomes with the published chloroplast genomes of other Salicaceae species, we demonstrate that the synteny and length of chloroplast genomes in Salicaceae are highly conserved but experienced dynamic evolution among species. We identify seven positively selected chloroplast genes in Salicaceae, which might be related to the adaptive evolution of Salicaceae species. Comparative chloroplast genome analysis within the family also indicates that some chloroplast genes are lost or became pseudogenes, infer that the chloroplast genes horizontally transferred to the nucleus genome. Based on the complete nucleus genome sequences from two Salicaceae species, we remarkably identify that the entire chloroplast genome is indeed transferred and integrated to the nucleus genome in the individual of the reference genome of P. trichocarpa at least once. This observation, along with presence of the large nuclear plastid DNA (NUPTs and NUPTs-containing multiple chloroplast genes in their original order in the chloroplast genome, favors the DNA-mediated hypothesis of organelle to nucleus DNA transfer. Overall, the phylogenomic analysis using chloroplast complete genomes clearly elucidates the phylogeny of Salicaceae. The identification of positively selected chloroplast genes and dynamic chloroplast-to-nucleus gene transfers in

  6. Transcript and metabolite analysis in Trincadeira cultivar reveals novel information regarding the dynamics of grape ripening.

    Science.gov (United States)

    Fortes, Ana M; Agudelo-Romero, Patricia; Silva, Marta S; Ali, Kashif; Sousa, Lisete; Maltese, Federica; Choi, Young H; Grimplet, Jerome; Martinez-Zapater, José M; Verpoorte, Robert; Pais, Maria S

    2011-11-02

    results were integrated with transcriptional profiling obtained using genome array to provide new information regarding the network of events leading to grape ripening. Altogether the data obtained provides the most extensive survey obtained so far for gene expression and metabolites accumulated during grape ripening. Moreover, it highlighted information obtained in a poorly known variety exhibiting particular characteristics that may be cultivar specific or dependent upon climatic conditions. Several genes were identified that had not been previously reported in the context of grape ripening namely genes involved in carbohydrate and amino acid metabolisms as well as in growth regulators; metabolism, epigenetic factors and signaling pathways. Some of these genes were annotated as receptors, transcription factors, and kinases and constitute good candidates for functional analysis in order to establish a model for ripening control of a non-climacteric fruit.

  7. Controllability analysis of transcriptional regulatory networks reveals circular control patterns among transcription factors

    DEFF Research Database (Denmark)

    Österlund, Tobias; Bordel, Sergio; Nielsen, Jens

    2015-01-01

    % for the human network. The high controllability (low number of drivers needed to control the system) in yeast, mouse and human is due to the presence of internal loops in their regulatory networks where the TFs regulate each other in a circular fashion. We refer to these internal loops as circular control...... motifs (CCM). The E. coli transcriptional regulatory network, which does not have any CCMs, shows a hierarchical structure of the transcriptional regulatory network in contrast to the eukaryal networks. The presence of CCMs also has influence on the stability of these networks, as the presence of cycles...

  8. Combined genome-wide expression profiling and targeted RNA interference in primary mouse macrophages reveals perturbation of transcriptional networks associated with interferon signalling

    Directory of Open Access Journals (Sweden)

    Craigon Marie

    2009-08-01

    Full Text Available Abstract Background Interferons (IFNs are potent antiviral cytokines capable of reprogramming the macrophage phenotype through the induction of interferon-stimulated genes (ISGs. Here we have used targeted RNA interference to suppress the expression of a number of key genes associated with IFN signalling in murine macrophages prior to stimulation with interferon-gamma. Genome-wide changes in transcript abundance caused by siRNA activity were measured using exon-level microarrays in the presence or absence of IFNγ. Results Transfection of murine bone-marrow derived macrophages (BMDMs with a non-targeting (control siRNA and 11 sequence-specific siRNAs was performed using a cationic lipid transfection reagent (Lipofectamine2000 prior to stimulation with IFNγ. Total RNA was harvested from cells and gene expression measured on Affymetrix GeneChip Mouse Exon 1.0 ST Arrays. Network-based analysis of these data revealed six siRNAs to cause a marked shift in the macrophage transcriptome in the presence or absence IFNγ. These six siRNAs targeted the Ifnb1, Irf3, Irf5, Stat1, Stat2 and Nfkb2 transcripts. The perturbation of the transcriptome by the six siRNAs was highly similar in each case and affected the expression of over 600 downstream transcripts. Regulated transcripts were clustered based on co-expression into five major groups corresponding to transcriptional networks associated with the type I and II IFN response, cell cycle regulation, and NF-KB signalling. In addition we have observed a significant non-specific immune stimulation of cells transfected with siRNA using Lipofectamine2000, suggesting use of this reagent in BMDMs, even at low concentrations, is enough to induce a type I IFN response. Conclusion Our results provide evidence that the type I IFN response in murine BMDMs is dependent on Ifnb1, Irf3, Irf5, Stat1, Stat2 and Nfkb2, and that siRNAs targeted to these genes results in perturbation of key transcriptional networks associated

  9. Millstone: software for multiplex microbial genome analysis and engineering.

    Science.gov (United States)

    Goodman, Daniel B; Kuznetsov, Gleb; Lajoie, Marc J; Ahern, Brian W; Napolitano, Michael G; Chen, Kevin Y; Chen, Changping; Church, George M

    2017-05-25

    Inexpensive DNA sequencing and advances in genome editing have made computational analysis a major rate-limiting step in adaptive laboratory evolution and microbial genome engineering. We describe Millstone, a web-based platform that automates genotype comparison and visualization for projects with up to hundreds of genomic samples. To enable iterative genome engineering, Millstone allows users to design oligonucleotide libraries and create successive versions of reference genomes. Millstone is open source and easily deployable to a cloud platform, local cluster, or desktop, making it a scalable solution for any lab.

  10. Microarray analysis of a salamander hopeful monster reveals transcriptional signatures of paedomorphic brain development

    Science.gov (United States)

    2010-01-01

    Background The Mexican axolotl (Ambystoma mexicanum) is considered a hopeful monster because it exhibits an adaptive and derived mode of development - paedomorphosis - that has evolved rapidly and independently among tiger salamanders. Unlike related tiger salamanders that undergo metamorphosis, axolotls retain larval morphological traits into adulthood and thus present an adult body plan that differs dramatically from the ancestral (metamorphic) form. The basis of paedomorphic development was investigated by comparing temporal patterns of gene transcription between axolotl and tiger salamander larvae (Ambystoma tigrinum tigrinum) that typically undergo a metamorphosis. Results Transcript abundances from whole brain and pituitary were estimated via microarray analysis on four different days post hatching (42, 56, 70, 84 dph) and regression modeling was used to independently identify genes that were differentially expressed as a function of time in both species. Collectively, more differentially expressed genes (DEGs) were identified as unique to the axolotl (n = 76) and tiger salamander (n = 292) than were identified as shared (n = 108). All but two of the shared DEGs exhibited the same temporal pattern of expression and the unique genes tended to show greater changes later in the larval period when tiger salamander larvae were undergoing anatomical metamorphosis. A second, complementary analysis that directly compared the expression of 1320 genes between the species identified 409 genes that differed as a function of species or the interaction between time and species. Of these 409 DEGs, 84% exhibited higher abundances in tiger salamander larvae at all sampling times. Conclusions Many of the unique tiger salamander transcriptional responses are probably associated with metamorphic biological processes. However, the axolotl also showed unique patterns of transcription early in development. In particular, the axolotl showed a genome-wide reduction in mRNA abundance

  11. Microarray analysis of a salamander hopeful monster reveals transcriptional signatures of paedomorphic brain development

    Directory of Open Access Journals (Sweden)

    Putta Srikrishna

    2010-06-01

    Full Text Available Abstract Background The Mexican axolotl (Ambystoma mexicanum is considered a hopeful monster because it exhibits an adaptive and derived mode of development - paedomorphosis - that has evolved rapidly and independently among tiger salamanders. Unlike related tiger salamanders that undergo metamorphosis, axolotls retain larval morphological traits into adulthood and thus present an adult body plan that differs dramatically from the ancestral (metamorphic form. The basis of paedomorphic development was investigated by comparing temporal patterns of gene transcription between axolotl and tiger salamander larvae (Ambystoma tigrinum tigrinum that typically undergo a metamorphosis. Results Transcript abundances from whole brain and pituitary were estimated via microarray analysis on four different days post hatching (42, 56, 70, 84 dph and regression modeling was used to independently identify genes that were differentially expressed as a function of time in both species. Collectively, more differentially expressed genes (DEGs were identified as unique to the axolotl (n = 76 and tiger salamander (n = 292 than were identified as shared (n = 108. All but two of the shared DEGs exhibited the same temporal pattern of expression and the unique genes tended to show greater changes later in the larval period when tiger salamander larvae were undergoing anatomical metamorphosis. A second, complementary analysis that directly compared the expression of 1320 genes between the species identified 409 genes that differed as a function of species or the interaction between time and species. Of these 409 DEGs, 84% exhibited higher abundances in tiger salamander larvae at all sampling times. Conclusions Many of the unique tiger salamander transcriptional responses are probably associated with metamorphic biological processes. However, the axolotl also showed unique patterns of transcription early in development. In particular, the axolotl showed a genome

  12. Genome-wide systematic characterization of the bZIP transcriptional factor family in tomato (Solanum lycopersicum L.).

    Science.gov (United States)

    Li, Dayong; Fu, Fuyou; Zhang, Huijuan; Song, Fengming

    2015-10-12

    Transcription factors of the basic leucine zipper (bZIP) family represent exclusively in eukaryotes and have been shown to regulate diverse biological processes in plant growth and development as well as in abiotic and biotic stress responses. However, little is known about the bZIP family in tomato (Solanum lycopersicum L.). The SlbZIP genes were identified using local BLAST and hidden Markov model profile searches. The phylogenetic trees, conserved motifs and gene structures were generated by MEGA6.06, MEME tool and gene Structure Display Server, respectively. The syntenic block diagrams were generated by the Circos software. The transcriptional gene expression profiles were obtained using Genevestigator tool and quantitative RT-PCR. In the present study, we carried out a genome-wide identification and systematic analyses of 69 SlbZIP genes that distributes unevenly on the tomato chromosomes. This family can be divided into 9 groups according to the phylogenetic relationship among the SlbZIP proteins. Six kinds of intron patterns (a-f) within the basic and hinge regions are defined. The additional conserved motifs and their presence of the group specificity were also identified. Further, we predicted the DNA-binding patterns and the dimerization property on the basis of the characteristic features in the basic and hinge regions and the leucine zipper, respectively, which supports our classification greatly and helps to classify 24 distinct subfamilies. Within the SlbZIP family, a total of 40 SlbZIP genes are located in the segmental duplicate regions in the tomato genome, suggesting that the segment chromosomal duplications contribute greatly to the expansion of the tomato SlbZIP family. Expression profiling analyses of 59 SlbZIP genes using quantitative RT-PCR and publicly available microarray data indicate that the tomato SlbZIP genes have distinct and diverse expression patterns in different tissues and developmental stages and many of the tomato bZIP genes

  13. Integration of multi-omics data of a genome-reduced bacterium: Prevalence of post-transcriptional regulation and its correlation with protein abundances

    Science.gov (United States)

    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

  14. Transcriptional analysis of the multicopy hao gene coding for hydroxylamine oxidoreductase in Nitrosomonas sp. strain ENI-11.

    Science.gov (United States)

    Hirota, Ryuichi; Kuroda, Akio; Ikeda, Tsukasa; Takiguchi, Noboru; Ohtake, Hisao; Kato, Junichi

    2006-08-01

    The nitrifying bacterium Nitrosomonas sp. strain ENI-11 has three copies of the gene encoding hydroxylamine oxidoreductase (hao(1), hao(2), and hao(3)) on its genome. Broad-host-range reporter plasmids containing transcriptional fusion genes between hao copies and lacZ were constructed to analyze the expression of each hydroxylamine oxidoreductase gene (hao) copy individually and quantitatively. beta-Galactosidase assays of ENI-11 harboring reporter plasmids revealed that all hao copies were transcribed in the wild-type strain. Promoter analysis of hao copies revealed that transcription of hao(3) was highest among the hao copies. Expression levels of hao(1) and hao(2) were 40% and 62% of that of hao(3) respectively. Transcription of hao(1) was negatively regulated, whereas a portion of hao(3) transcription was read through transcription from the rpsT promoter. When energy-depleted cells were incubated in the growth medium, only hao(3) expression increased. This result suggests that it is hao(3) that is responsible for recovery from energy-depleted conditions in Nitrosomonas sp. strain ENI-11.

  15. Genome-wide analysis of Tol2 transposon reintegration in zebrafish

    Directory of Open Access Journals (Sweden)

    Parinov Sergey

    2009-09-01

    Full Text Available Abstract Background Tol2, a member of the hAT family of transposons, has become a useful tool for genetic manipulation of model animals, but information about its interactions with vertebrate genomes is still limited. Furthermore, published reports on Tol2 have mainly been based on random integration of the transposon system after co-injection of a plasmid DNA harboring the transposon and a transposase mRNA. It is important to understand how Tol2 would behave upon activation after integration into the genome. Results We performed a large-scale enhancer trap (ET screen and generated 338 insertions of the Tol2 transposon-based ET cassette into the zebrafish genome. These insertions were generated by remobilizing the transposon from two different donor sites in two transgenic lines. We found that 39% of Tol2 insertions occurred in transcription units, mostly into introns. Analysis of the transposon target sites revealed no strict specificity at the DNA sequence level. However, Tol2 was prone to target AT-rich regions with weak palindromic consensus sequences centered at the insertion site. Conclusion Our systematic analysis of sequential remobilizations of the Tol2 transposon from two independent sites within a vertebrate genome has revealed properties such as a tendency to integrate into transcription units and into AT-rich palindrome-like sequences. This information will influence the development of various applications involving DNA transposons and Tol2 in particular.

  16. Genome-wide analysis of Tol2 transposon reintegration in zebrafish.

    Science.gov (United States)

    Kondrychyn, Igor; Garcia-Lecea, Marta; Emelyanov, Alexander; Parinov, Sergey; Korzh, Vladimir

    2009-09-08

    Tol2, a member of the hAT family of transposons, has become a useful tool for genetic manipulation of model animals, but information about its interactions with vertebrate genomes is still limited. Furthermore, published reports on Tol2 have mainly been based on random integration of the transposon system after co-injection of a plasmid DNA harboring the transposon and a transposase mRNA. It is important to understand how Tol2 would behave upon activation after integration into the genome. We performed a large-scale enhancer trap (ET) screen and generated 338 insertions of the Tol2 transposon-based ET cassette into the zebrafish genome. These insertions were generated by remobilizing the transposon from two different donor sites in two transgenic lines. We found that 39% of Tol2 insertions occurred in transcription units, mostly into introns. Analysis of the transposon target sites revealed no strict specificity at the DNA sequence level. However, Tol2 was prone to target AT-rich regions with weak palindromic consensus sequences centered at the insertion site. Our systematic analysis of sequential remobilizations of the Tol2 transposon from two independent sites within a vertebrate genome has revealed properties such as a tendency to integrate into transcription units and into AT-rich palindrome-like sequences. This information will influence the development of various applications involving DNA transposons and Tol2 in particular.

  17. Comparative Genomic Analysis of Clinical and Environmental Vibrio Vulnificus Isolates Revealed Biotype 3 Evolutionary Relationships

    Directory of Open Access Journals (Sweden)

    Yael eKotton

    2015-01-01

    Full Text Available In 1996 a common-source outbreak of severe soft tissue and bloodstream infections erupted among Israeli fish farmers and fish consumers due to changes in fish marketing policies. The causative pathogen was a new strain of Vibrio vulnificus, named biotype 3, which displayed a unique biochemical and genotypic profile. Initial observations suggested that the pathogen erupted as a result of genetic recombination between two distinct populations. We applied a whole genome shotgun sequencing approach using several V. vulnificus strains from Israel in order to study the pan genome of V. vulnificus and determine the phylogenetic relationship of biotype 3 with existing populations. The core genome of V. vulnificus based on 16 draft and complete genomes consisted of 3068 genes, representing between 59% and 78% of the whole genome of 16 strains. The accessory genome varied in size from 781 kbp to 2044 kbp. Phylogenetic analysis based on whole, core, and accessory genomes displayed similar clustering patterns with two main clusters, clinical (C and environmental (E, all biotype 3 strains formed a distinct group within the E cluster. Annotation of accessory genomic regions found in biotype 3 strains and absent from the core genome yielded 1732 genes, of which the vast majority encoded hypothetical proteins, phage-related proteins, and mobile element proteins. A total of 1916 proteins (including 713 hypothetical proteins were present in all human pathogenic strains (both biotype 3 and non-biotype 3 and absent from the environmental strains. Clustering analysis of the non-hypothetical proteins revealed 148 protein clusters shared by all human pathogenic strains; these included transcriptional regulators, arylsulfatases, methyl-accepting chemotaxis proteins, acetyltransferases, GGDEF family proteins, transposases, type IV secretory system (T4SS proteins, and integrases. Our study showed that V. vulnificus biotype 3 evolved from environmental populations and

  18. SIGMA: A System for Integrative Genomic Microarray Analysis of Cancer Genomes

    Directory of Open Access Journals (Sweden)

    Davies Jonathan J

    2006-12-01

    Full Text Available Abstract Background The prevalence of high resolution profiling of genomes has created a need for the integrative analysis of information generated from multiple methodologies and platforms. Although the majority of data in the public domain are gene expression profiles, and expression analysis software are available, the increase of array CGH studies has enabled integration of high throughput genomic and gene expression datasets. However, tools for direct mining and analysis of array CGH data are limited. Hence, there is a great need for analytical and display software tailored to cross platform integrative analysis of cancer genomes. Results We have created a user-friendly java application to facilitate sophisticated visualization and analysis such as cross-tumor and cross-platform comparisons. To demonstrate the utility of this software, we assembled array CGH data representing Affymetrix SNP chip, Stanford cDNA arrays and whole genome tiling path array platforms for cross comparison. This cancer genome database contains 267 profiles from commonly used cancer cell lines representing 14 different tissue types. Conclusion In this study we have developed an application for the visualization and analysis of data from high resolution array CGH platforms that can be adapted for analysis of multiple types of high throughput genomic datasets. Furthermore, we invite researchers using array CGH technology to deposit both their raw and processed data, as this will be a continually expanding database of cancer genomes. This publicly available resource, the System for Integrative Genomic Microarray Analysis (SIGMA of cancer genomes, can be accessed at http://sigma.bccrc.ca.

  19. Comprehensive Behavioral Analysis of Activating Transcription Factor 5-Deficient Mice

    Directory of Open Access Journals (Sweden)

    Mariko Umemura

    2017-07-01

    Full Text Available Activating transcription factor 5 (ATF5 is a member of the CREB/ATF family of basic leucine zipper transcription factors. We previously reported that ATF5-deficient (ATF5-/- mice demonstrated abnormal olfactory bulb development due to impaired interneuron supply. Furthermore, ATF5-/- mice were less aggressive than ATF5+/+ mice. Although ATF5 is widely expressed in the brain, and involved in the regulation of proliferation and development of neurons, the physiological role of ATF5 in the higher brain remains unknown. Our objective was to investigate the physiological role of ATF5 in the higher brain. We performed a comprehensive behavioral analysis using ATF5-/- mice and wild type littermates. ATF5-/- mice exhibited abnormal locomotor activity in the open field test. They also exhibited abnormal anxiety-like behavior in the light/dark transition test and open field test. Furthermore, ATF5-/- mice displayed reduced social interaction in the Crawley’s social interaction test and increased pain sensitivity in the hot plate test compared with wild type. Finally, behavioral flexibility was reduced in the T-maze test in ATF5-/- mice compared with wild type. In addition, we demonstrated that ATF5-/- mice display disturbances of monoamine neurotransmitter levels in several brain regions. These results indicate that ATF5 deficiency elicits abnormal behaviors and the disturbance of monoamine neurotransmitter levels in the brain. The behavioral abnormalities of ATF5-/- mice may be due to the disturbance of monoamine levels. Taken together, these findings suggest that ATF5-/- mice may be a unique animal model of some psychiatric disorders.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2010-03-15

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

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

    International Nuclear Information System (INIS)

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

    2010-03-01

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

  2. Barcode server: a visualization-based genome analysis system.

    Directory of Open Access Journals (Sweden)

    Fenglou Mao

    Full Text Available We have previously developed a computational method for representing a genome as a barcode image, which makes various genomic features visually apparent. We have demonstrated that this visual capability has made some challenging genome analysis problems relatively easy to solve. We have applied this capability to a number of challenging problems, including (a identification of horizontally transferred genes, (b identification of genomic islands with special properties and (c binning of metagenomic sequences, and achieved highly encouraging results. These application results inspired us to develop this barcode-based genome analysis server for public service, which supports the following capabilities: (a calculation of the k-mer based barcode image for a provided DNA sequence; (b detection of sequence fragments in a given genome with distinct barcodes from those of the majority of the genome, (c clustering of provided DNA sequences into groups having similar barcodes; and (d homology-based search using Blast against a genome database for any selected genomic regions deemed to have interesting barcodes. The barcode server provides a job management capability, allowing processing of a large number of analysis jobs for barcode-based comparative genome analyses. The barcode server is accessible at http://csbl1.bmb.uga.edu/Barcode.

  3. Functional Genomic investigation of Peroxisome Proliferator-Activated Receptor Gamma (PPARG mediated transcription response in gastric cancer

    Directory of Open Access Journals (Sweden)

    Karthikeyan Selvarasu

    2017-10-01

    Full Text Available Cancer is a complex and progressive multi-step disorder that results from the transformation of normal cells to malignant derivatives. Several oncogenic signaling pathways are involved in this transformation. PPARG (Peroxisome proliferator-activated receptor gamma mediated transcription and signaling is involved in few cancers. We have investigated the PPARG in gastric tumors. The objective of the present study was to investigate the PPARG mediated transcriptional response in gastric tumors. Gene-set based and pathway focused gene-set enrichment analysis of available PPARG signatures in gastric tumor mRNA profiles shows that PPARG mediated transcription is highly activated in intestinal sub-type of gastric tumors. Further, we have derived the PPARG associated genes in gastric cancer and their expression was identified for the association with the better survival of the patients. Analysis of the PPARG associated genes reveals their involvement in mitotic cell cycle process, chromosome organization and nuclear division. Towards identifying the association with other oncogenic signaling process, E2F regulated genes were found associated with PPARG mediated transcription. The current results reveal the possible stratification of gastric tumors based on the PPARG gene expression and the possible development of PPARG targeted gastric cancer therapeutics. The identified PPARG regulated genes were identified to be targetable by pioglitazone and rosiglitazone. The identification of PPARG genes also in the normal stomach tissues reveal the possible involvement of these genes in the normal physiology of stomach and needs to be investigated.

  4. A Genomics Approach to Tumor Gemome Analysis

    National Research Council Canada - National Science Library

    Collins, Colin

    2002-01-01

    Genomes of solid tumors are often highly rearranged and these rearrangements promote cancer progression through disruption of genes mediating immortality, survival, metastasis, and resistance to therapy...

  5. Cloning and functional analysis of human mTERFL encoding a novel mitochondrial transcription termination factor-like protein

    International Nuclear Information System (INIS)

    Chen Yao; Zhou Guangjin; Yu Min; He Yungang; Tang Wei; Lai Jianhua; He Jie; Liu Wanguo; Tan Deyong

    2005-01-01

    Serum plays an important role in the regulation of cell cycle and cell growth. To identify novel serum-inhibitory factors and study their roles in cell cycle regulation, we performed mRNA differential display analysis of U251 cells in the presence or absence of serum and cloned a novel gene encoding the human mitochondrial transcription termination factor-like protein (mTERFL). The full-length mTERFL cDNA has been isolated and the genomic structure determined. The mTERFL gene consists of three exons and encodes 385 amino acids with 52% sequence similarity to the human mitochondrial transcription termination factor (mTERF). However, mTERFL and mTERF have an opposite expression pattern in response to serum. The expression of mTERFL is dramatically inhibited by the addition of serum in serum-starved cells while the mTERF is rather induced. Northern blot analysis detected three mTERFL transcripts of 1.7, 3.2, and 3.5 kb. Besides the 3.2 kb transcript that is unique to skeletal muscle, other two transcripts express predominant in heart, liver, pancreas, and skeletal muscle. Expression of the GFP-mTERFL fusion protein in HeLa cells localized it to the mitochondria. Furthermore, ectopic expression of mTERFL suppresses cell growth and arrests cells in the G1 stage demonstrated by MTT and flow cytometry analysis. Collectively, our data suggest that mTERFL is a novel mTERF family member and a serum-inhibitory factor probably participating in the regulation of cell growth through the modulation of mitochondrial transcription

  6. Global analysis of WRKY transcription factor superfamily in Setaria identifies potential candidates involved in abiotic stress signalling

    Directory of Open Access Journals (Sweden)

    Mehanathan eMuthamilarasan

    2015-10-01

    Full Text Available Transcription factors (TFs are major players in stress signalling and constitute an integral part of signalling networks. Among the major TFs, WRKY proteins play pivotal roles in regulation of transcriptional reprogramming associated with stress responses. In view of this, genome- and transcriptome-wide identification of WRKY TF family was performed in the C4 model plants, Setaria italica (SiWRKY and S. viridis (SvWRKY, respectively. The study identified 105 SiWRKY and 44 SvWRKY proteins that were computationally analysed for their physicochemical properties. Sequence alignment and phylogenetic analysis classified these proteins into three major groups, namely I, II and III with majority of WRKY proteins belonging to group II (53 SiWRKY and 23 SvWRKY, followed by group III (39 SiWRKY and 11 SvWRKY and group I (10 SiWRKY and 6 SvWRKY. Group II proteins were further classified into 5 subgroups (IIa to IIe based on their phylogeny. Domain analysis showed the presence of WRKY motif and zinc finger-like structures in these proteins along with additional domains in a few proteins. All SiWRKY genes were physically mapped on the S. italica genome and their duplication analysis revealed that 10 and 8 gene pairs underwent tandem and segmental duplications, respectively. Comparative mapping of SiWRKY and SvWRKY genes in related C4 panicoid genomes demonstrated the orthologous relationships between these genomes. In silico expression analysis of SiWRKY and SvWRKY genes showed their differential expression patterns in different tissues and stress conditions. Expression profiling of candidate SiWRKY genes in response to stress (dehydration and salinity and hormone treatments (abscisic acid, salicylic acid and methyl jasmonate suggested the putative involvement of SiWRKY066 and SiWRKY082 in stress and hormone signalling. These genes could be potential candidates for further characterization to delineate their functional roles in abiotic stress signalling.

  7. Global analysis of WRKY transcription factor superfamily in Setaria identifies potential candidates involved in abiotic stress signaling.

    Science.gov (United States)

    Muthamilarasan, Mehanathan; Bonthala, Venkata S; Khandelwal, Rohit; Jaishankar, Jananee; Shweta, Shweta; Nawaz, Kashif; Prasad, Manoj

    2015-01-01

    Transcription factors (TFs) are major players in stress signaling and constitute an integral part of signaling networks. Among the major TFs, WRKY proteins play pivotal roles in regulation of transcriptional reprogramming associated with stress responses. In view of this, genome- and transcriptome-wide identification of WRKY TF family was performed in the C4model plants, Setaria italica (SiWRKY) and S. viridis (SvWRKY), respectively. The study identified 105 SiWRKY and 44 SvWRKY proteins that were computationally analyzed for their physicochemical properties. Sequence alignment and phylogenetic analysis classified these proteins into three major groups, namely I, II, and III with majority of WRKY proteins belonging to group II (53 SiWRKY and 23 SvWRKY), followed by group III (39 SiWRKY and 11 SvWRKY) and group I (10 SiWRKY and 6 SvWRKY). Group II proteins were further classified into 5 subgroups (IIa to IIe) based on their phylogeny. Domain analysis showed the presence of WRKY motif and zinc finger-like structures in these proteins along with additional domains in a few proteins. All SiWRKY genes were physically mapped on the S. italica genome and their duplication analysis revealed that 10 and 8 gene pairs underwent tandem and segmental duplications, respectively. Comparative mapping of SiWRKY and SvWRKY genes in related C4 panicoid genomes demonstrated the orthologous relationships between these genomes. In silico expression analysis of SiWRKY and SvWRKY genes showed their differential expression patterns in different tissues and stress conditions. Expression profiling of candidate SiWRKY genes in response to stress (dehydration and salinity) and hormone treatments (abscisic acid, salicylic acid, and methyl jasmonate) suggested the putative involvement of SiWRKY066 and SiWRKY082 in stress and hormone signaling. These genes could be potential candidates for further characterization to delineate their functional roles in abiotic stress signaling.

  8. Global analysis of WRKY transcription factor superfamily in Setaria identifies potential candidates involved in abiotic stress signaling

    Science.gov (United States)

    Muthamilarasan, Mehanathan; Bonthala, Venkata S.; Khandelwal, Rohit; Jaishankar, Jananee; Shweta, Shweta; Nawaz, Kashif; Prasad, Manoj

    2015-01-01

    Transcription factors (TFs) are major players in stress signaling and constitute an integral part of signaling networks. Among the major TFs, WRKY proteins play pivotal roles in regulation of transcriptional reprogramming associated with stress responses. In view of this, genome- and transcriptome-wide identification of WRKY TF family was performed in the C4model plants, Setaria italica (SiWRKY) and S. viridis (SvWRKY), respectively. The study identified 105 SiWRKY and 44 SvWRKY proteins that were computationally analyzed for their physicochemical properties. Sequence alignment and phylogenetic analysis classified these proteins into three major groups, namely I, II, and III with majority of WRKY proteins belonging to group II (53 SiWRKY and 23 SvWRKY), followed by group III (39 SiWRKY and 11 SvWRKY) and group I (10 SiWRKY and 6 SvWRKY). Group II proteins were further classified into 5 subgroups (IIa to IIe) based on their phylogeny. Domain analysis showed the presence of WRKY motif and zinc finger-like structures in these proteins along with additional domains in a few proteins. All SiWRKY genes were physically mapped on the S. italica genome and their duplication analysis revealed that 10 and 8 gene pairs underwent tandem and segmental duplications, respectively. Comparative mapping of SiWRKY and SvWRKY genes in related C4 panicoid genomes demonstrated the orthologous relationships between these genomes. In silico expression analysis of SiWRKY and SvWRKY genes showed their differential expression patterns in different tissues and stress conditions. Expression profiling of candidate SiWRKY genes in response to stress (dehydration and salinity) and hormone treatments (abscisic acid, salicylic acid, and methyl jasmonate) suggested the putative involvement of SiWRKY066 and SiWRKY082 in stress and hormone signaling. These genes could be potential candidates for further characterization to delineate their functional roles in abiotic stress signaling. PMID:26635818

  9. Bioinformatics Identification of Modules of Transcription Factor Binding Sites in Alzheimer's Disease-Related Genes by In Silico Promoter Analysis and Microarrays

    Directory of Open Access Journals (Sweden)

    Regina Augustin

    2011-01-01

    Full Text Available The molecular mechanisms and genetic risk factors underlying Alzheimer's disease (AD pathogenesis are only partly understood. To identify new factors, which may contribute to AD, different approaches are taken including proteomics, genetics, and functional genomics. Here, we used a bioinformatics approach and found that distinct AD-related genes share modules of transcription factor binding sites, suggesting a transcriptional coregulation. To detect additional coregulated genes, which may potentially contribute to AD, we established a new bioinformatics workflow with known multivariate methods like support vector machines, biclustering, and predicted transcription factor binding site modules by using in silico analysis and over 400 expression arrays from human and mouse. Two significant modules are composed of three transcription factor families: CTCF, SP1F, and EGRF/ZBPF, which are conserved between human and mouse APP promoter sequences. The specific combination of in silico promoter and multivariate analysis can identify regulation mechanisms of genes involved in multifactorial diseases.

  10. Genome-wide identification and characterization of Notch transcription complex-binding sequence paired sites in leukemia cells

    Science.gov (United States)

    Severson, Eric; Arnett, Kelly L.; Wang, Hongfang; Zang, Chongzhi; Taing, Len; Liu, Hudan; Pear, Warren S.; Liu, X. Shirley; Blacklow, Stephen C.; Aster, Jon C.

    2018-01-01

    Notch transcription complexes (NTCs) drive target gene expression by binding to two distinct types of genomic response elements, NTC monomer-binding sites and sequence-paired sites (SPSs) that bind NTC dimers. SPSs are conserved and are linked to the Notch-responsiveness of a few genes, but their overall contribution to Notch-dependent gene regulation is unknown. To address this issue, we determined the DNA sequence requirements for NTC dimerization using a fluorescence resonance energy transfer (FRET) assay, and applied insights from these in vitro studies to Notch-“addicted” leukemia cells. We find that SPSs contribute to the regulation of approximately a third of direct Notch target genes. While originally described in promoters, SPSs are present mainly in long-range enhancers, including an enhancer containing a newly described SPS that regulates HES5. Our work provides a general method for identifying sequence-paired sites in genome-wide data sets and highlights the widespread role of NTC dimerization in Notch-transformed leukemia cells. PMID:28465412

  11. Pathway and network analysis of cancer genomes

    DEFF Research Database (Denmark)

    Creixell, Pau; Reimand, Jueri; Haider, Syed

    2015-01-01

    Genomic information on tumors from 50 cancer types cataloged by the International Cancer Genome Consortium (ICGC) shows that only a few well-studied driver genes are frequently mutated, in contrast to many infrequently mutated genes that may also contribute to tumor biology. Hence there has been...

  12. Analysis of Genome-Scale Data

    NARCIS (Netherlands)

    Kemmeren, P.P.C.W.

    2005-01-01

    The genetic material of every cell in an organism is stored inside DNA in the form of genes, which together form the genome. The information stored in the DNA is translated to RNA and subsequently to proteins, which form complex biological systems. The availability of whole genome sequences has

  13. GENOME ANALYSIS OF BURKHOLDERIA CEPACIA AC1100

    Science.gov (United States)

    Burkholderia cepacia is an important organism in bioremediation of environmental pollutants and it is also of increasing interest as a human pathogen. The genomic organization of B. cepacia is being studied in order to better understand its unusual adaptive capacity and genome pl...

  14. Genomics-enabled analysis of the emergent disease cotton bacterial blight.

    Directory of Open Access Journals (Sweden)

    Anne Z Phillips

    2017-09-01

    Full Text Available Cotton bacterial blight (CBB, an important disease of (Gossypium hirsutum in the early 20th century, had been controlled by resistant germplasm for over half a century. Recently, CBB re-emerged as an agronomic problem in the United States. Here, we report analysis of cotton variety planting statistics that indicate a steady increase in the percentage of susceptible cotton varieties grown each year since 2009. Phylogenetic analysis revealed that strains from the current outbreak cluster with race 18 Xanthomonas citri pv. malvacearum (Xcm strains. Illumina based draft genomes were generated for thirteen Xcm isolates and analyzed along with 4 previously published Xcm genomes. These genomes encode 24 conserved and nine variable type three effectors. Strains in the race 18 clade contain 3 to 5 more effectors than other Xcm strains. SMRT sequencing of two geographically and temporally diverse strains of Xcm yielded circular chromosomes and accompanying plasmids. These genomes encode eight and thirteen distinct transcription activator-like effector genes. RNA-sequencing revealed 52 genes induced within two cotton cultivars by both tested Xcm strains. This gene list includes a homeologous pair of genes, with homology to the known susceptibility gene, MLO. In contrast, the two strains of Xcm induce different clade III SWEET sugar transporters. Subsequent genome wide analysis revealed patterns in the overall expression of homeologous gene pairs in cotton after inoculation by Xcm. These data reveal important insights into the Xcm-G. hirsutum disease complex and strategies for future development of resistant cultivars.

  15. Exogenous reference gene normalization for real-time reverse transcription-polymerase chain reaction analysis under dynamic endogenous transcription.

    Science.gov (United States)

    Johnston, Stephen; Gallaher, Zachary; Czaja, Krzysztof

    2012-05-15

    Quantitative real-time reverse transcription-polymerase chain reaction (qPCR) is widely used to investigate transcriptional changes following experimental manipulations to the nervous system. Despite the widespread utilization of qPCR, the interpretation of results is marred by the lack of a suitable reference gene due to the dynamic nature of endogenous transcription. To address this inherent deficiency, we investigated the use of an exogenous spike-in mRNA, luciferase, as an internal reference gene for the 2(-∆∆Ct) normalization method. To induce dynamic transcription, we systemically administered capsaicin, a neurotoxin selective for C-type sensory neurons expressing the TRPV-1 receptor, to adult male Sprague-Dawley rats. We later isolated nodose ganglia for qPCR analysis with the reference being either exogenous luciferase mRNA or the commonly used endogenous reference β-III tubulin. The exogenous luciferase mRNA reference clearly demonstrated the dynamic expression of the endogenous reference. Furthermore, variability of the endogenous reference would lead to misinterpretation of other genes of interest. In conclusion, traditional reference genes are often unstable under physiologically normal situations, and certainly unstable following the damage to the nervous system. The use of exogenous spike-in reference provides a consistent and easily implemented alternative for the analysis of qPCR data.

  16. Transcriptional analysis of ESAT-6 cluster 3 in Mycobacterium smegmatis

    Directory of Open Access Journals (Sweden)

    Riccardi Giovanna

    2009-03-01

    Full Text Available Abstract Background The ESAT-6 (early secreted antigenic target, 6 kDa family collects small mycobacterial proteins secreted by Mycobacterium tuberculosis, particularly in the early phase of growth. There are 23 ESAT-6 family members in M. tuberculosis H37Rv. In a previous work, we identified the Zur- dependent regulation of five proteins of the ESAT-6/CFP-10 family (esxG, esxH, esxQ, esxR, and esxS. esxG and esxH are part of ESAT-6 cluster 3, whose expression was already known to be induced by iron starvation. Results In this research, we performed EMSA experiments and transcriptional analysis of ESAT-6 cluster 3 in Mycobacterium smegmatis (msmeg0615-msmeg0625 and M. tuberculosis. In contrast to what we had observed in M. tuberculosis, we found that in M. smegmatis ESAT-6 cluster 3 responds only to iron and not to zinc. In both organisms we identified an internal promoter, a finding which suggests the presence of two transcriptional units and, by consequence, a differential expression of cluster 3 genes. We compared the expression of msmeg0615 and msmeg0620 in different growth and stress conditions by means of relative quantitative PCR. The expression of msmeg0615 and msmeg0620 genes was essentially similar; they appeared to be repressed in most of the tested conditions, with the exception of acid stress (pH 4.2 where msmeg0615 was about 4-fold induced, while msmeg0620 was repressed. Analysis revealed that in acid stress conditions M. tuberculosis rv0282 gene was 3-fold induced too, while rv0287 induction was almost insignificant. Conclusion In contrast with what has been reported for M. tuberculosis, our results suggest that in M. smegmatis only IdeR-dependent regulation is retained, while zinc has no effect on gene expression. The role of cluster 3 in M. tuberculosis virulence is still to be defined; however, iron- and zinc-dependent expression strongly suggests that cluster 3 is highly expressed in the infective process, and that the cluster

  17. Genomic profiling of neutrophil transcripts in Asian Qigong practitioners: a pilot study in gene regulation by mind-body interaction.

    Science.gov (United States)

    Li, Quan-Zhen; Li, Ping; Garcia, Gabriela E; Johnson, Richard J; Feng, Lili

    2005-02-01

    The great similarity of the genomes of humans and other species stimulated us to search for genes regulated by elements associated with human uniqueness, such as the mind-body interaction. DNA microarray technology offers the advantage of analyzing thousands of genes simultaneously, with the potential to determine healthy phenotypic changes in gene expression. The aim of this study was to determine the genomic profile and function of neutrophils in Falun Gong (FLG, an ancient Chinese Qigong) practitioners, with healthy subjects as controls. Six (6) Asian FLG practitioners and 6 Asian normal healthy controls were recruited for our study. The practitioners have practiced FLG for at least 1 year (range, 1-5 years). The practice includes daily reading of FLG books and daily practice of exercises lasting 1-2 hours. Selected normal healthy controls did not perform Qigong, yoga, t'ai chi, or any other type of mind-body practice, and had not followed any conventional physical exercise program for at least 1 year. Neutrophils were isolated from fresh blood and assayed for gene expression, using microarrays and RNase protection assay (RPA), as well as for function (phagocytosis) and survival (apoptosis). The changes in gene expression of FLG practitioners in contrast to normal healthy controls were characterized by enhanced immunity, downregulation of cellular metabolism, and alteration of apoptotic genes in favor of a rapid resolution of inflammation. The lifespan of normal neutrophils was prolonged, while the inflammatory neutrophils displayed accelerated cell death in FLG practitioners as determined by enzyme-linked immunosorbent assay. Correlating with enhanced immunity reflected by microarray data, neutrophil phagocytosis was significantly increased in Qigong practitioners. Some of the altered genes observed by microarray were confirmed by RPA. Qigong practice may regulate immunity, metabolic rate, and cell death, possibly at the transcriptional level. Our pilot study

  18. Genome-Wide Identification and Expression Analysis of WRKY Gene Family in Capsicum annuum L.

    Science.gov (United States)

    Diao, Wei-Ping; Snyder, John C; Wang, Shu-Bin; Liu, Jin-Bing; Pan, Bao-Gui; Guo, Guang-Jun; Wei, Ge

    2016-01-01

    The WRKY family of transcription factors is one of the most important families of plant transcriptional regulators with members regulating multiple biological processes, especially in regulating defense against biotic and abiotic stresses. However, little information is available about WRKYs in pepper (Capsicum annuum L.). The recent release of completely assembled genome sequences of pepper allowed us to perform a genome-wide investigation for pepper WRKY proteins. In the present study, a total of 71 WRKY genes were identified in the pepper genome. According to structural features of their encoded proteins, the pepper WRKY genes (CaWRKY) were classified into three main groups, with the second group further divided into five subgroups. Genome mapping analysis revealed that CaWRKY were enriched on four chromosomes, especially on chromosome 1, and 15.5% of the family members were tandemly duplicated genes. A phylogenetic tree was constructed depending on WRKY domain' sequences derived from pepper and Arabidopsis. The expression of 21 selected CaWRKY genes in response to seven different biotic and abiotic stresses (salt, heat shock, drought, Phytophtora capsici, SA, MeJA, and ABA) was evaluated by quantitative RT-PCR; Some CaWRKYs were highly expressed and up-regulated by stress treatment. Our results will provide a platform for functional identification and molecular breeding studies of WRKY genes in pepper.

  19. Genomic analysis of mouse retinal development.

    Directory of Open Access Journals (Sweden)

    Seth Blackshaw

    2004-09-01

    Full Text Available The vertebrate retina is comprised of seven major cell types that are generated in overlapping but well-defined intervals. To identify genes that might regulate retinal development, gene expression in the developing retina was profiled at multiple time points using serial analysis of gene expression (SAGE. The expression patterns of 1,051 genes that showed developmentally dynamic expression by SAGE were investigated using in situ hybridization. A molecular atlas of gene expression in the developing and mature retina was thereby constructed, along with a taxonomic classification of developmental gene expression patterns. Genes were identified that label both temporal and spatial subsets of mitotic progenitor cells. For each developing and mature major retinal cell type, genes selectively expressed in that cell type were identified. The gene expression profiles of retinal Müller glia and mitotic progenitor cells were found to be highly similar, suggesting that Müller glia might serve to produce multiple retinal cell types under the right conditions. In addition, multiple transcripts that were evolutionarily conserved that did not appear to encode open reading frames of more than 100 amino acids in length ("noncoding RNAs" were found to be dynamically and specifically expressed in developing and mature retinal cell types. Finally, many photoreceptor-enriched genes that mapped to chromosomal intervals containing retinal disease genes were identified. These data serve as a starting point for functional investigations of the roles of these genes in retinal development and physiology.

  20. Decoding genome-wide GadEWX-transcriptional regulatory networks reveals multifaceted cellular responses to acid stress in Escherichia coli

    DEFF Research Database (Denmark)

    Seo, Sang Woo; Kim, Donghyuk; O'Brien, Edward J.

    2015-01-01

    The regulators GadE, GadW and GadX (which we refer to as GadEWX) play a critical role in the transcriptional regulation of the glutamate-dependent acid resistance (GDAR) system in Escherichia coli K-12 MG1655. However, the genome-wide regulatory role of GadEWX is still unknown. Here we comprehens...

  1. Chromatin immunoprecipitation (ChIP) of plant transcription factors followed by sequencing (ChIP-SEQ) or hybridization to whole genome arrays (ChIP-CHIP)

    NARCIS (Netherlands)

    Kaufmann, K.; Muiño, J.M.; Østerås, M.; Farinelli, L.; Krajewski, P.; Angenent, G.C.

    2010-01-01

    Chromatin immunoprecipitation (ChIP) is a powerful technique to study interactions between transcription factors (TFs) and DNA in vivo. For genome-wide de novo discovery of TF-binding sites, the DNA that is obtained in ChIP experiments needs to be processed for sequence identification. The sequences

  2. Genome-wide Reconstruction of OxyR and SoxRS Transcriptional Regulatory Networks under Oxidative Stress in Escherichia coli K-12 MG1655

    DEFF Research Database (Denmark)

    Seo, Sang Woo; Kim, Donghyuk; Szubin, Richard

    2015-01-01

    Three transcription factors (TFs), OxyR, SoxR, and SoxS, play a critical role in transcriptional regulation of the defense system for oxidative stress in bacteria. However, their full genome-wide regulatory potential is unknown. Here, we perform a genome-scale reconstruction of the OxyR, SoxR, an...

  3. GenomePeek—an online tool for prokaryotic genome and metagenome analysis

    Directory of Open Access Journals (Sweden)

    Katelyn McNair

    2015-06-01

    Full Text Available As more and more prokaryotic sequencing takes place, a method to quickly and accurately analyze this data is needed. Previous tools are mainly designed for metagenomic analysis and have limitations; such as long runtimes and significant false positive error rates. The online tool GenomePeek (edwards.sdsu.edu/GenomePeek was developed to analyze both single genome and metagenome sequencing files, quickly and with low error rates. GenomePeek uses a sequence assembly approach where reads to a set of conserved genes are extracted, assembled and then aligned against the highly specific reference database. GenomePeek was found to be faster than traditional approaches while still keeping error rates low, as well as offering unique data visualization options.

  4. Exploratory analysis of genomic segmentations with Segtools

    Directory of Open Access Journals (Sweden)

    Buske Orion J

    2011-10-01

    Full Text Available Abstract Background As genome-wide experiments and annotations become more prevalent, researchers increasingly require tools to help interpret data at this scale. Many functional genomics experiments involve partitioning the genome into labeled segments, such that segments sharing the same label exhibit one or more biochemical or functional traits. For example, a collection of ChlP-seq experiments yields a compendium of peaks, each labeled with one or more associated DNA-binding proteins. Similarly, manually or automatically generated annotations of functional genomic elements, including cis-regulatory modules and protein-coding or RNA genes, can also be summarized as genomic segmentations. Results We present a software toolkit called Segtools that simplifies and automates the exploration of genomic segmentations. The software operates as a series of interacting tools, each of which provides one mode of summarization. These various tools can be pipelined and summarized in a single HTML page. We describe the Segtools toolkit and demonstrate its use in interpreting a collection of human histone modification data sets and Plasmodium falciparum local chromatin structure data sets. Conclusions Segtools provides a convenient, powerful means of interpreting a genomic segmentation.

  5. EG-13GENOME-WIDE METHYLATION ANALYSIS IDENTIFIES GENOMIC DNA DEMETHYLATION DURING MALIGNANT PROGRESSION OF GLIOMAS

    Science.gov (United States)

    Saito, Kuniaki; Mukasa, Akitake; Nagae, Genta; Aihara, Koki; Otani, Ryohei; Takayanagi, Shunsaku; Omata, Mayu; Tanaka, Shota; Shibahara, Junji; Takahashi, Miwako; Momose, Toshimitsu; Shimamura, Teppei; Miyano, Satoru; Narita, Yoshitaka; Ueki, Keisuke; Nishikawa, Ryo; Nagane, Motoo; Aburatani, Hiroyuki; Saito, Nobuhito

    2014-01-01

    Low-grade gliomas often undergo malignant progression, and these transformations are a leading cause of death in patients with low-grade gliomas. However, the molecular mechanisms underlying malignant tumor progression are still not well understood. Recent evidence indicates that epigenetic deregulation is an important cause of gliomagenesis; therefore, we examined the impact of epigenetic changes during malignant progression of low-grade gliomas. Specifically, we used the Illumina Infinium Human Methylation 450K BeadChip to perform genome-wide DNA methylation analysis of 120 gliomas and four normal brains. This study sample included 25 matched-pairs of initial low-grade gliomas and recurrent tumors (temporal heterogeneity) and 20 of the 25 recurring tumors recurred as malignant progressions, and one matched-pair of newly emerging malignant lesions and pre-existing lesions (spatial heterogeneity). Analyses of methylation profiles demonstrated that most low-grade gliomas in our sample (43/51; 84%) had a CpG island methylator phenotype (G-CIMP). Remarkably, approximately 50% of secondary glioblastomas that had progressed from low-grade tumors with the G-CIMP status exhibited a characteristic partial demethylation of genomic DNA during malignant progression, but other recurrent gliomas showed no apparent change in DNA methylation pattern. Interestingly, we found that most loci that were demethylated during malignant progression were located outside of CpG islands. The information of histone modifications patterns in normal human astrocytes and embryonal stem cells also showed that the ratio of active marks at the site corresponding to DNA demethylated loci in G-CIMP-demethylated tumors was significantly lower; this finding indicated that most demethylated loci in G-CIMP-demethylated tumors were likely transcriptionally inactive. A small number of the genes that were upregulated and had demethylated CpG islands were associated with cell cycle-related pathway. In

  6. Analysis of intra-genomic GC content homogeneity within prokaryotes

    DEFF Research Database (Denmark)

    Bohlin, J; Snipen, L; Hardy, S.P.

    2010-01-01

    the GC content varies within microbial genomes to assess whether this property can be associated with certain biological functions related to the organism's environment and phylogeny. We utilize a new quantity GCVAR, the intra-genomic GC content variability with respect to the average GC content......Bacterial genomes possess varying GC content (total guanines (Gs) and cytosines (Cs) per total of the four bases within the genome) but within a given genome, GC content can vary locally along the chromosome, with some regions significantly more or less GC rich than on average. We have examined how...... both aerobic and facultative microbes. Although an association has previously been found between mean genomic GC content and oxygen requirement, our analysis suggests that no such association exits when phylogenetic bias is accounted for. A significant association between GCVAR and mean GC content...

  7. Creation and genomic analysis of irradiation hybrids in Populus

    Science.gov (United States)

    Matthew S. Zinkgraf; K. Haiby; M.C. Lieberman; L. Comai; I.M. Henry; Andrew Groover

    2016-01-01

    Establishing efficient functional genomic systems for creating and characterizing genetic variation in forest trees is challenging. Here we describe protocols for creating novel gene-dosage variation in Populus through gamma-irradiation of pollen, followed by genomic analysis to identify chromosomal regions that have been deleted or inserted in...

  8. Bayesian error analysis model for reconstructing transcriptional regulatory networks

    OpenAIRE

    Sun, Ning; Carroll, Raymond J.; Zhao, Hongyu

    2006-01-01

    Transcription regulation is a fundamental biological process, and extensive efforts have been made to dissect its mechanisms through direct biological experiments and regulation modeling based on physical–chemical principles and mathematical formulations. Despite these efforts, transcription regulation is yet not well understood because of its complexity and limitations in biological experiments. Recent advances in high throughput technologies have provided substantial amounts and diverse typ...

  9. The complexity of Rhipicephalus (Boophilus microplus genome characterised through detailed analysis of two BAC clones

    Directory of Open Access Journals (Sweden)

    Valle Manuel

    2011-07-01

    Full Text Available Abstract Background Rhipicephalus (Boophilus microplus (Rmi a major cattle ectoparasite and tick borne disease vector, impacts on animal welfare and industry productivity. In arthropod research there is an absence of a complete Chelicerate genome, which includes ticks, mites, spiders, scorpions and crustaceans. Model arthropod genomes such as Drosophila and Anopheles are too taxonomically distant for a reference in tick genomic sequence analysis. This study focuses on the de-novo assembly of two R. microplus BAC sequences from the understudied R microplus genome. Based on available R. microplus sequenced resources and comparative analysis, tick genomic structure and functional predictions identify complex gene structures and genomic targets expressed during tick-cattle interaction. Results In our BAC analyses we have assembled, using the correct positioning of BAC end sequences and transcript sequences, two challenging genomic regions. Cot DNA fractions compared to the BAC sequences confirmed a highly repetitive BAC sequence BM-012-E08 and a low repetitive BAC sequence BM-005-G14 which was gene rich and contained short interspersed elements (SINEs. Based directly on the BAC and Cot data comparisons, the genome wide frequency of the SINE Ruka element was estimated. Using a conservative approach to the assembly of the highly repetitive BM-012-E08, the sequence was de-convoluted into three repeat units, each unit containing an 18S, 5.8S and 28S ribosomal RNA (rRNA encoding gene sequence (rDNA, related internal transcribed spacer and complex intergenic region. In the low repetitive BM-005-G14, a novel gene complex was found between to 2 genes on the same strand. Nested in the second intron of a large 9 Kb papilin gene was a helicase gene. This helicase overlapped in two exonic regions with the papilin. Both these genes were shown expressed in different tick life stage important in ectoparasite interaction with the host. Tick specific sequence

  10. Analysis of Genome-Scale Data

    OpenAIRE

    Kemmeren, P.P.C.W.

    2005-01-01

    The genetic material of every cell in an organism is stored inside DNA in the form of genes, which together form the genome. The information stored in the DNA is translated to RNA and subsequently to proteins, which form complex biological systems. The availability of whole genome sequences has given rise to the parallel development of other high-throughput approaches such as determining mRNA expression level changes, gene-deletion phenotypes, chromosomal location of DNA binding proteins, cel...

  11. Structure and transcription of the Helicoverpa armigera densovirus (HaDV2) genome and its expression strategy in LD652 cells.

    Science.gov (United States)

    Xu, Pengjun; Graham, Robert I; Wilson, Kenneth; Wu, Kongming

    2017-02-07

    Densoviruses (DVs) are highly pathogenic to their hosts. However, we previously reported a mutualistic DV (HaDV2). Very little was known about the characteristics of this virus, so herein we undertook a series of experiments to explore the molecular biology of HaDV2 further. Phylogenetic analysis showed that HaDV2 was similar to members of the genus Iteradensovirus. However, compared to current members of the genus Iteradensovirus, the sequence identity of HaDV2 is less than 44% at the nucleotide-level, and lower than 36, 28 and 19% at the amino-acid-level of VP, NS1 and NS2 proteins, respectively. Moreover, NS1 and NS2 proteins from HaDV2 were smaller than those from other iteradensoviruses due to their shorter N-terminal sequences. Two transcripts of about 2.2 kb coding for the NS proteins and the VP proteins were identified by Northern Blot and RACE analysis. Using specific anti-NS1 and anti-NS2 antibodies, Western Blot analysis revealed a 78 kDa and a 48 kDa protein, respectively. Finally, the localization of both NS1 and NS2 proteins within the cell nucleus was determined by using Green Fluorescent Protein (GFP) labelling. The genome organization, terminal hairpin structure, transcription and expression strategies as well as the mutualistic relationship with its host, suggested that HaDV2 was a novel member of the genus Iteradensovirus within the subfamily Densovirinae.

  12. Functional regression method for whole genome eQTL epistasis analysis with sequencing data.

    Science.gov (United States)

    Xu, Kelin; Jin, Li; Xiong, Momiao

    2017-05-18

    Epistasis plays an essential rule in understanding the regulation mechanisms and is an essential component of the genetic architecture of the gene expressions. However, interaction analysis of gene expressions remains fundamentally unexplored due to great computational challenges and data availability. Due to variation in splicing, transcription start sites, polyadenylation sites, post-transcriptional RNA editing across the entire gene, and transcription rates of the cells, RNA-seq measurements generate large expression variability and collectively create the observed position level read count curves. A single number for measuring gene expression which is widely used for microarray measured gene expression analysis is highly unlikely to sufficiently account for large expression variation across the gene. Simultaneously analyzing epistatic architecture using the RNA-seq and whole genome sequencing (WGS) data poses enormous challenges. We develop a nonlinear functional regression model (FRGM) with functional responses where the position-level read counts within a gene are taken as a function of genomic position, and functional predictors where genotype profiles are viewed as a function of genomic position, for epistasis analysis with RNA-seq data. Instead of testing the interaction of all possible pair-wises SNPs, the FRGM takes a gene as a basic unit for epistasis analysis, which tests for the interaction of all possible pairs of genes and use all the information that can be accessed to collectively test interaction between all possible pairs of SNPs within two genome regions. By large-scale simulations, we demonstrate that the proposed FRGM for epistasis analysis can achieve the correct type 1 error and has higher power to detect the interactions between genes than the existing methods. The proposed methods are applied to the RNA-seq and WGS data from the 1000 Genome Project. The numbers of pairs of significantly interacting genes after Bonferroni correction

  13. Evolution of a Pathogen: A Comparative Genomics Analysis Identifies a Genetic Pathway to Pathogenesis in Acinetobacter

    Science.gov (United States)

    Sahl, Jason W.; Gillece, John D.; Schupp, James M.; Waddell, Victor G.; Driebe, Elizabeth M.; Engelthaler, David M.; Keim, Paul

    2013-01-01

    Acinetobacter baumannii is an emergent and global nosocomial pathogen. In addition to A. baumannii, other Acinetobacter species, especially those in the Acinetobacter calcoaceticus-baumannii (Acb) complex, have also been associated with serious human infection. Although mechanisms of attachment, persistence on abiotic surfaces, and pathogenesis in A. baumannii have been identified, the genetic mechanisms that explain the emergence of A. baumannii as the most widespread and virulent Acinetobacter species are not fully understood. Recent whole genome sequencing has provided insight into the phylogenetic structure of the genus Acinetobacter. However, a global comparison of genomic features between Acinetobacter spp. has not been described in the literature. In this study, 136 Acinetobacter genomes, including 67 sequenced in this study, were compared to identify the acquisition and loss of genes in the expansion of the Acinetobacter genus. A whole genome phylogeny confirmed that A. baumannii is a monophyletic clade and that the larger Acb complex is also a well-supported monophyletic group. The whole genome phylogeny provided the framework for a global genomic comparison based on a blast score ratio (BSR) analysis. The BSR analysis demonstrated that specific genes have been both lost and acquired in the evolution of A. baumannii. In addition, several genes associated with A. baumannii pathogenesis were found to be more conserved in the Acb complex, and especially in A. baumannii, than in other Acinetobacter genomes; until recently, a global analysis of the distribution and conservation of virulence factors across the genus was not possible. The results demonstrate that the acquisition of specific virulence factors has likely contributed to the widespread persistence and virulence of A. baumannii. The identification of novel features associated with transcriptional regulation and acquired by clades in the Acb complex presents targets for better understanding the

  14. Detailed analysis of putative genes encoding small proteins in legume genomes

    Directory of Open Access Journals (Sweden)

    Gabriel eGuillén

    2013-06-01

    Full Text Available Diverse plant genome sequencing projects coupled with powerful bioinformatics tools have facilitated massive data analysis to construct specialized databases classified according to cellular function. However, there are still a considerable number of genes encoding proteins whose function has not yet been characterized. Included in this category are small proteins (SPs, 30-150 amino acids encoded by short open reading frames (sORFs. SPs play important roles in plant physiology, growth, and development. Unfortunately, protocols focused on the genome-wide identification and characterization of sORFs are scarce or remain poorly implemented. As a result, these genes are underrepresented in many genome annotations. In this work, we exploited publicly available genome sequences of Phaseolus vulgaris, Medicago truncatula, Glycine max and Lotus japonicus to analyze the abundance of annotated SPs in plant legumes. Our strategy to uncover bona fide sORFs at the genome level was centered in bioinformatics analysis of characteristics such as evidence of expression (transcription, presence of known protein regions or domains, and identification of orthologous genes in the genomes explored. We collected 6170, 10461, 30521, and 23599 putative sORFs from P. vulgaris, G. max, M. truncatula, and L. japonicus genomes, respectively. Expressed sequence tags (ESTs available in the DFCI Gene Index database provided evidence that ~one-third of the predicted legume sORFs are expressed. Most potential SPs have a counterpart in a different plant species and counterpart regions or domains in larger proteins. Potential functional sORFs were also classified according to a reduced set of GO categories, and the expression of 13 of them during P. vulgaris nodule ontogeny was confirmed by qPCR. This analysis provides a collection of sORFs that potentially encode for meaningful SPs, and offers the possibility of their further functional evaluation.

  15. Selection of reference genes for transcriptional analysis of edible tubers of potato (Solanum tuberosum L..

    Directory of Open Access Journals (Sweden)

    Roberta Fogliatto Mariot

    Full Text Available Potato (Solanum tuberosum yield has increased dramatically over the last 50 years and this has been achieved by a combination of improved agronomy and biotechnology efforts. Gene studies are taking place to improve new qualities and develop new cultivars. Reverse transcriptase quantitative polymerase chain reaction (RT-qPCR is a bench-marking analytical tool for gene expression analysis, but its accuracy is highly dependent on a reliable normalization strategy of an invariant reference genes. For this reason, the goal of this work was to select and validate reference genes for transcriptional analysis of edible tubers of potato. To do so, RT-qPCR primers were designed for ten genes with relatively stable expression in potato tubers as observed in RNA-Seq experiments. Primers were designed across exon boundaries to avoid genomic DNA contamination. Differences were observed in the ranking of candidate genes identified by geNorm, NormFinder and BestKeeper algorithms. The ranks determined by geNorm and NormFinder were very similar and for all samples the most stable candidates were C2, exocyst complex component sec3 (SEC3 and ATCUL3/ATCUL3A/CUL3/CUL3A (CUL3A. According to BestKeeper, the importin alpha and ubiquitin-associated/ts-n genes were the most stable. Three genes were selected as reference genes for potato edible tubers in RT-qPCR studies. The first one, called C2, was selected in common by NormFinder and geNorm, the second one is SEC3, selected by NormFinder, and the third one is CUL3A, selected by geNorm. Appropriate reference genes identified in this work will help to improve the accuracy of gene expression quantification analyses by taking into account differences that may be observed in RNA quality or reverse transcription efficiency across the samples.

  16. Selection of reference genes for transcriptional analysis of edible tubers of potato (Solanum tuberosum L.).

    Science.gov (United States)

    Mariot, Roberta Fogliatto; de Oliveira, Luisa Abruzzi; Voorhuijzen, Marleen M; Staats, Martijn; Hutten, Ronald C B; Van Dijk, Jeroen P; Kok, Esther; Frazzon, Jeverson

    2015-01-01

    Potato (Solanum tuberosum) yield has increased dramatically over the last 50 years and this has been achieved by a combination of improved agronomy and biotechnology efforts. Gene studies are taking place to improve new qualities and develop new cultivars. Reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) is a bench-marking analytical tool for gene expression analysis, but its accuracy is highly dependent on a reliable normalization strategy of an invariant reference genes. For this reason, the goal of this work was to select and validate reference genes for transcriptional analysis of edible tubers of potato. To do so, RT-qPCR primers were designed for ten genes with relatively stable expression in potato tubers as observed in RNA-Seq experiments. Primers were designed across exon boundaries to avoid genomic DNA contamination. Differences were observed in the ranking of candidate genes identified by geNorm, NormFinder and BestKeeper algorithms. The ranks determined by geNorm and NormFinder were very similar and for all samples the most stable candidates were C2, exocyst complex component sec3 (SEC3) and ATCUL3/ATCUL3A/CUL3/CUL3A (CUL3A). According to BestKeeper, the importin alpha and ubiquitin-associated/ts-n genes were the most stable. Three genes were selected as reference genes for potato edible tubers in RT-qPCR studies. The first one, called C2, was selected in common by NormFinder and geNorm, the second one is SEC3, selected by NormFinder, and the third one is CUL3A, selected by geNorm. Appropriate reference genes identified in this work will help to improve the accuracy of gene expression quantification analyses by taking into account differences that may be observed in RNA quality or reverse transcription efficiency across the samples.

  17. A Resource for the Transcriptional Signature of Bona Fide Trophoblast Stem Cells and Analysis of Their Embryonic Persistence

    Directory of Open Access Journals (Sweden)

    Georg Kuales

    2015-01-01

    Full Text Available Trophoblast stem cells (TSCs represent the multipotent progenitors that give rise to the different cells of the embryonic portion of the placenta. Here, we analysed the expression of key TSC transcription factors Cdx2, Eomes, and Elf5 in the early developing placenta of mouse embryos and in cultured TSCs and reveal surprising heterogeneity in protein levels. We analysed persistence of TSCs in the early placenta and find that TSCs remain in the chorionic hinge until E9.5 and are lost shortly afterwards. To define the transcriptional signature of bona fide TSCs, we used inducible gain- and loss-of-function alleles of Eomes or Cdx2, and EomesGFP, to manipulate and monitor the core maintenance factors of TSCs, followed by genome-wide expression profiling. Combinatorial analysis of resulting expression profiles allowed for defining novel TSC marker genes that might functionally contribute to the maintenance of the TSC state. Analyses by qRT-PCR and in situ hybridisation validated novel TSC- and chorion-specific marker genes, such as Bok/Mtd, Cldn26, Duox2, Duoxa2, Nr0b1, and Sox21. Thus, these expression data provide a valuable resource for the transcriptional signature of bona fide and early differentiating TSCs and may contribute to an increased understanding of the transcriptional circuitries that maintain and/or establish stemness of TSCs.

  18. Two alternatively spliced GPR39 transcripts in seabream: molecular cloning, genomic organization, and regulation of gene expression by metabolic signals.

    Science.gov (United States)

    Zhang, Yong; Liu, Yun; Huang, Xigui; Liu, Xiaochun; Jiao, Baowei; Meng, Zining; Zhu, Pei; Li, Shuisheng; Lin, Haoran; Cheng, Christopher H K

    2008-12-01

    Two GPR39 transcripts, designated as sbGPR39-1a and sbGPR39-1b, were identified in black seabream (Acanthopagrus schlegeli). The deduced amino acid (aa) sequence of sbGPR39-1a contains 423 residues with seven putative transmembrane (TM) domains. On the other hand, sbGPR39-1b contains 284 aa residues with only five putative TM domains. Northern blot analysis confirmed the presence of two GPR39 transcripts in the seabream intestine, stomach, and liver. Apart from seabream, the presence of two GPR39 transcripts was also found to exist in a number of teleosts (zebrafish and pufferfish) and mammals (human and mouse). Analysis of the GPR39 gene structure in different species suggests that the two GPR39 transcripts are generated by alternative splicing. When the seabream receptors were expressed in cultured HEK293 cells, Zn(2)(+) could trigger sbGPR39-1a signaling through the serum response element pathway, but no such functionality could be detected for the sbGPR39-1b receptor. The two receptors were found to be differentially expressed in seabream tissues. sbGPR39-1a is predominantly expressed in the gastrointestinal tract. On the other hand, sbGPR39-1b is widely expressed in most central and peripheral tissues except muscle and ovary. The expression of sbGPR39-1a in the intestine and the expression of sbGPR39-1b in the hypothalamus were decreased significantly during food deprivation in seabream. On the contrary, the expression of the GH secretagogue receptors (sbGHSR-1a and sbGHSR-1b) was significantly increased in the hypothalamus of the food-deprived seabream. The reciprocal regulatory patterns of expression of these two genes suggest that both of them are involved in controlling the physiological response of the organism during starvation.

  19. Genomics of a Metamorphic Timing QTL: met1 Maps to a Unique Genomic Position and Regulates Morph and Species-Specific Patterns of Brain Transcription

    Science.gov (United States)

    Page, Robert B.; Boley, Meredith A.; Kump, David K.; Voss, Stephen R.

    2013-01-01

    Very little is known about genetic factors that regulate life history transitions during ontogeny. Closely related tiger salamanders (Ambystoma species complex) show extreme variation in metamorphic timing, with some species foregoing metamorphosis altogether, an adaptive trait called paedomorphosis. Previous studies identified a major effect quantitative trait locus (met1) for metamorphic timing and expression of paedomorphosis in hybrid crosses between the biphasic Eastern tiger salamander (Ambystoma tigrinum tigrinum) and the paedomorphic Mexican axolotl (Ambystoma mexicanum). We used existing hybrid mapping panels and a newly created hybrid cross to map the met1 genomic region and determine the effect of met1 on larval growth, metamorphic timing, and gene expression in the brain. We show that met1 maps to the position of a urodele-specific chromosome rearrangement on linkage group 2 that uniquely brought functionally associated genes into linkage. Furthermore, we found that more than 200 genes were differentially expressed during larval development as a function of met1 genotype. This list of differentially expressed genes is enriched for proteins that function in the mitochondria, providing evidence of a link between met1, thyroid hormone signaling, and mitochondrial energetics associated with metamorphosis. Finally, we found that met1 significantly affected metamorphic timing in hybrids, but not early larval growth rate. Collectively, our results show that met1 regulates species and morph-specific patterns of brain transcription and life history variation. PMID:23946331

  20. High resolution analysis of the human transcriptome: detection of extensive alternative splicing independent of transcriptional activity

    Directory of Open Access Journals (Sweden)

    Rouet Fabien

    2009-10-01

    Full Text Available Abstract Background Commercially available microarrays have been used in many settings to generate expression profiles for a variety of applications, including target selection for disease detection, classification, profiling for pharmacogenomic response to therapeutics, and potential disease staging. However, many commercially available microarray platforms fail to capture transcript diversity produced by alternative splicing, a major mechanism for driving proteomic diversity through transcript heterogeneity. Results The human Genome-Wide SpliceArray™ (GWSA, a novel microarray platform, utilizes an existing probe design concept to monitor such transcript diversity on a genome scale. The human GWSA allows the detection of alternatively spliced events within the human genome through the use of exon body and exon junction probes to provide a direct measure of each transcript, through simple calculations derived from expression data. This report focuses on the performance and validation of the array when measured against standards recently published by the Microarray Quality Control (MAQC Project. The array was shown to be highly quantitative, and displayed greater than 85% correlation with the HG-U133 Plus 2.0 array at the gene level while providing more extensive coverage of each gene. Almost 60% of splice events among genes demonstrating differential expression of greater than 3 fold also contained extensive splicing alterations. Importantly, almost 10% of splice events within the gene set displaying constant overall expression values had evidence of transcript diversity. Two examples illustrate the types of events identified: LIM domain 7 showed no differential expression at the gene level, but demonstrated deregulation of an exon skip event, while erythrocyte membrane protein band 4.1 -like 3 was differentially expressed and also displayed deregulation of a skipped exon isoform. Conclusion Significant changes were detected independent of

  1. A genome-wide survey on basic helix-loop-helix transcription factors in giant panda.

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    Chunwang Dang

    Full Text Available The giant panda (Ailuropoda melanoleuca is a critically endangered mammalian species. Studies on functions of regulatory proteins involved in developmental processes would facilitate understanding of specific behavior in giant panda. The basic helix-loop-helix (bHLH proteins play essential roles in a wide range of developmental processes in higher organisms. bHLH family members have been identified in over 20 organisms, including fruit fly, zebrafish, mouse and human. Our present study identified 107 bHLH family members being encoded in giant panda genome. Phylogenetic analyses revealed that they belong to 44 bHLH families with 46, 25, 15, 4, 11 and 3 members in group A, B, C, D, E and F, respectively, while the remaining 3 members were assigned into "orphan". Compared to mouse, the giant panda does not encode seven bHLH proteins namely Beta3a, Mesp2, Sclerax, S-Myc, Hes5 (or Hes6, EBF4 and Orphan 1. These results provide useful background information for future studies on structure and function of bHLH proteins in the regulation of giant panda development.

  2. Genome-Wide Transcription Study of Cryptococcus neoformans H99 Clinical Strain versus Environmental Strains.

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    Elaheh Movahed

    Full Text Available The infection of Cryptococcus neoformans is acquired through the inhalation of desiccated yeast cells and basidiospores originated from the environment, particularly from bird's droppings and decaying wood. Three environmental strains of C. neoformans originated from bird droppings (H4, S48B and S68B and C. neoformans reference clinical strain (H99 were used for intranasal infection in C57BL/6 mice. We showed that the H99 strain demonstrated higher virulence compared to H4, S48B and S68B strains. To examine if gene expression contributed to the different degree of virulence among these strains, a genome-wide microarray study was performed to inspect the transcriptomic profiles of all four strains. Our results revealed that out of 7,419 genes (22,257 probes examined, 65 genes were significantly up-or down-regulated in H99 versus H4, S48B and S68B strains. The up-regulated genes in H99 strain include Hydroxymethylglutaryl-CoA synthase (MVA1, Mitochondrial matrix factor 1 (MMF1, Bud-site-selection protein 8 (BUD8, High affinity glucose transporter 3 (SNF3 and Rho GTPase-activating protein 2 (RGA2. Pathway annotation using DAVID bioinformatics resource showed that metal ion binding and sugar transmembrane transporter activity pathways were highly expressed in the H99 strain. We suggest that the genes and pathways identified may possibly play crucial roles in the fungal pathogenesis.

  3. Analysis of the highly diverse gene borders in Ebola virus reveals a distinct mechanism of transcriptional regulation.

    Science.gov (United States)

    Brauburger, Kristina; Boehmann, Yannik; Tsuda, Yoshimi; Hoenen, Thomas; Olejnik, Judith; Schümann, Michael; Ebihara, Hideki; Mühlberger, Elke

    2014-11-01

    Ebola virus (EBOV) belongs to the group of nonsegmented negative-sense RNA viruses. The seven EBOV genes are separated by variable gene borders, including short (4- or 5-nucleotide) intergenic regions (IRs), a single long (144-nucleotide) IR, and gene overlaps, where the neighboring gene end and start signals share five conserved nucleotides. The unique structure of the gene overlaps and the presence of a single long IR are conserved among all filoviruses. Here, we sought to determine the impact of the EBOV gene borders during viral transcription. We show that readthrough mRNA synthesis occurs in EBOV-infected cells irrespective of the structure of the gene border, indicating that the gene overlaps do not promote recognition of the gene end signal. However, two consecutive gene end signals at the VP24 gene might improve termination at the VP24-L gene border, ensuring efficient L gene expression. We further demonstrate that the long IR is not essential for but regulates transcription reinitiation in a length-dependent but sequence-independent manner. Mutational analysis of bicistronic minigenomes and recombinant EBOVs showed no direct correlation between IR length and reinitiation rates but demonstrated that specific IR lengths not found naturally in filoviruses profoundly inhibit downstream gene expression. Intriguingly, although truncation of the 144-nucleotide-long IR to 5 nucleotides did not substantially affect EBOV transcription, it led to a significant reduction of viral growth. Our current understanding of EBOV transcription regulation is limited due to the requirement for high-containment conditions to study this highly pathogenic virus. EBOV is thought to share many mechanistic features with well-analyzed prototype nonsegmented negative-sense RNA viruses. A single polymerase entry site at the 3' end of the genome determines that transcription of the genes is mainly controlled by gene order and cis-acting signals found at the gene borders. Here, we examined

  4. GWAMA: software for genome-wide association meta-analysis

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    Mägi Reedik

    2010-05-01

    Full Text Available Abstract Background Despite the recent success of genome-wide association studies in identifying novel loci contributing effects to complex human traits, such as type 2 diabetes and obesity, much of the genetic component of variation in these phenotypes remains unexplained. One way to improving power to detect further novel loci is through meta-analysis of studies from the same population, increasing the sample size over any individual study. Although statistical software analysis packages incorporate routines for meta-analysis, they are ill equipped to meet the challenges of the scale and complexity of data generated in genome-wide association studies. Results We have developed flexible, open-source software for the meta-analysis of genome-wide association studies. The software incorporates a variety of error trapping facilities, and provides a range of meta-analysis summary statistics. The software is distributed with scripts that allow simple formatting of files containing the results of each association study and generate graphical summaries of genome-wide meta-analysis results. Conclusions The GWAMA (Genome-Wide Association Meta-Analysis software has been developed to perform meta-analysis of summary statistics generated from genome-wide association studies of dichotomous phenotypes or quantitative traits. Software with source files, documentation and example data files are freely available online at http://www.well.ox.ac.uk/GWAMA.

  5. msCentipede: Modeling Heterogeneity across Genomic Sites and Replicates Improves Accuracy in the Inference of Transcription Factor Binding.

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    Anil Raj

    Full Text Available Understanding global gene regulation depends critically on accurate annotation of regulatory elements that are functional in a given cell type. CENTIPEDE, a powerful, probabilistic framework for identifying transcription factor binding sites from tissue-specific DNase I cleavage patterns and genomic sequence content, leverages the hypersensitivity of factor-bound chromatin and the information in the DNase I spatial cleavage profile characteristic of each DNA binding protein to accurately infer functional factor binding sites. However, the model for the spatial profile in this framework fails to account for the substantial variation in the DNase I cleavage profiles across different binding sites. Neither does it account for variation in the profiles at the same binding site across multiple replicate DNase I experiments, which are increasingly available. In this work, we introduce new methods, based on multi-scale models for inhomogeneous Poisson processes, to account for such variation in DNase I cleavage patterns both within and across binding sites. These models account for the spatial structure in the heterogeneity in DNase I cleavage patterns for each factor. Using DNase-seq measurements assayed in a lymphoblastoid cell line, we demonstrate the improved performance of this model for several transcription factors by comparing against the Chip-seq peaks for those factors. Finally, we explore the effects of DNase I sequence bias on inference of factor binding using a simple extension to our framework that allows for a more flexible background model. The proposed model can also be easily applied to paired-end ATAC-seq and DNase-seq data. msCentipede, a Python implementation of our algorithm, is available at http://rajanil.github.io/msCentipede.

  6. msCentipede: Modeling Heterogeneity across Genomic Sites and Replicates Improves Accuracy in the Inference of Transcription Factor Binding.

    Science.gov (United States)

    Raj, Anil; Shim, Heejung; Gilad, Yoav; Pritchard, Jonathan K; Stephens, Matthew

    2015-01-01

    Understanding global gene regulation depends critically on accurate annotation of regulatory elements that are functional in a given cell type. CENTIPEDE, a powerful, probabilistic framework for identifying transcription factor binding sites from tissue-specific DNase I cleavage patterns and genomic sequence content, leverages the hypersensitivity of factor-bound chromatin and the information in the DNase I spatial cleavage profile characteristic of each DNA binding protein to accurately infer functional factor binding sites. However, the model for the spatial profile in this framework fails to account for the substantial variation in the DNase I cleavage profiles across different binding sites. Neither does it account for variation in the profiles at the same binding site across multiple replicate DNase I experiments, which are increasingly available. In this work, we introduce new methods, based on multi-scale models for inhomogeneous Poisson processes, to account for such variation in DNase I cleavage patterns both within and across binding sites. These models account for the spatial structure in the heterogeneity in DNase I cleavage patterns for each factor. Using DNase-seq measurements assayed in a lymphoblastoid cell line, we demonstrate the improved performance of this model for several transcription factors by comparing against the Chip-seq peaks for those factors. Finally, we explore the effects of DNase I sequence bias on inference of factor binding using a simple extension to our framework that allows for a more flexible background model. The proposed model can also be easily applied to paired-end ATAC-seq and DNase-seq data. msCentipede, a Python implementation of our algorithm, is available at http://rajanil.github.io/msCentipede.

  7. Trimming of mammalian transcriptional networks using network component analysis

    Directory of Open Access Journals (Sweden)

    Liao James C

    2010-10-01

    Full Text Available Abstract Background Network Component Analysis (NCA has been used to deduce the activities of transcription factors (TFs from gene expression data and the TF-gene binding relationship. However, the TF-gene interaction varies in different environmental conditions and tissues, but such information is rarely available and cannot be predicted simply by motif analysis. Thus, it is beneficial to identify key TF-gene interactions under the experimental condition based on transcriptome data. Such information would be useful in identifying key regulatory pathways and gene markers of TFs in further studies. Results We developed an algorithm to trim network connectivity such that the important regulatory interactions between the TFs and the genes were retained and the regulatory signals were deduced. Theoretical studies demonstrated that the regulatory signals were accurately reconstructed even in the case where only three independent transcriptome datasets were available. At least 80% of the main target genes were correctly predicted in the extreme condition of high noise level and small number of datasets. Our algorithm was tested with transcriptome data taken from mice under rapamycin treatment. The initial network topology from the literature contains 70 TFs, 778 genes, and 1423 edges between the TFs and genes. Our method retained 1074 edges (i.e. 75% of the original edge number and identified 17 TFs as being significantly perturbed under the experimental condition. Twelve of these TFs are involved in MAPK signaling or myeloid leukemia pathways defined in the KEGG database, or are known to physically interact with each other. Additionally, four of these TFs, which are Hif1a, Cebpb, Nfkb1, and Atf1, are known targets of rapamycin. Furthermore, the trimmed network was able to predict Eno1 as an important target of Hif1a; this key interaction could not be detected without trimming the regulatory network. Conclusions The advantage of our new algorithm

  8. Identification and functional analysis of two alternatively spliced transcripts of ABSCISIC ACID INSENSITIVE3 (ABI3) in linseed flax (Linum usitatissimum L.).

    Science.gov (United States)

    Wang, Yanyan; Zhang, Tianbao; Song, Xiaxia; Zhang, Jianping; Dang, Zhanhai; Pei, Xinwu; Long, Yan

    2018-01-01

    Alternative splicing is a popular phenomenon in different types of plants. It can produce alternative spliced transcripts that encode proteins with altered functions. Previous studies have shown that one transcription factor, ABSCISIC ACID INSENSITIVE3 (ABI3), which encodes an important component in abscisic acid (ABA) signaling, is subjected to alternative splicing in both mono- and dicotyledons. In the current study, we identified two homologs of ABI3 in the genome of linseed flax. We screened two alternatively spliced flax LuABI3 transcripts, LuABI3-2 and LuABI3-3, and one normal flax LuABI3 transcript, LuABI3-1. Sequence analysis revealed that one of the alternatively spliced transcripts, LuABI3-3, retained a 6 bp intron. RNA accumulation analysis showed that all three transcripts were expressed during seed development, while subcellular localization and transgene experiments showed that LuABI3-3 had no biological function. The two normal transcripts, LuABI3-1 and LuABI3-2, are the important functional isoforms in flax and play significant roles in the ABA regulatory pathway during seed development, germination, and maturation.

  9. Identification and functional analysis of two alternatively spliced transcripts of ABSCISIC ACID INSENSITIVE3 (ABI3 in linseed flax (Linum usitatissimum L..

    Directory of Open Access Journals (Sweden)

    Yanyan Wang

    Full Text Available Alternative splicing is a popular phenomenon in different types of plants. It can produce alternative spliced transcripts that encode proteins with altered functions. Previous studies have shown that one transcription factor, ABSCISIC ACID INSENSITIVE3 (ABI3, which encodes an important component in abscisic acid (ABA signaling, is subjected to alternative splicing in both mono- and dicotyledons. In the current study, we identified two homologs of ABI3 in the genome of linseed flax. We screened two alternatively spliced flax LuABI3 transcripts, LuABI3-2 and LuABI3-3, and one normal flax LuABI3 transcript, LuABI3-1. Sequence analysis revealed that one of the alternatively spliced transcripts, LuABI3-3, retained a 6 bp intron. RNA accumulation analysis showed that all three transcripts were expressed during seed development, while subcellular localization and transgene experiments showed that LuABI3-3 had no biological function. The two normal transcripts, LuABI3-1 and LuABI3-2, are the important functional isoforms in flax and play significant roles in the ABA regulatory pathway during seed development, germination, and maturation.

  10. Bradyrhizobium elkanii nod regulon: insights through genomic analysis

    Directory of Open Access Journals (Sweden)

    Luciane M. P. Passaglia

    2017-07-01

    Full Text Available Abstract A successful symbiotic relationship between soybean [Glycine max (L. Merr.] and Bradyrhizobium species requires expression of the bacterial structural nod genes that encode for the synthesis of lipochitooligosaccharide nodulation signal molecules, known as Nod factors (NFs. Bradyrhizobium diazoefficiens USDA 110 possesses a wide nodulation gene repertoire that allows NF assembly and modification, with transcription of the nodYABCSUIJnolMNOnodZ operon depending upon specific activators, i.e., products of regulatory nod genes that are responsive to signaling molecules such as flavonoid compounds exuded by host plant roots. Central to this regulatory circuit of nod gene expression are NodD proteins, members of the LysR-type regulator family. In this study, publicly available Bradyrhizobium elkanii sequenced genomes were compared with the closely related B. diazoefficiens USDA 110 reference genome to determine the similarities between those genomes, especially with regards to the nod operon and nod regulon. Bioinformatics analyses revealed a correlation between functional mechanisms and key elements that play an essential role in the regulation of nod gene expression. These analyses also revealed new genomic features that had not been clearly explored before, some of which were unique for some B. elkanii genomes.

  11. Genome-wide comparative analysis of codon usage bias and codon context patterns among cyanobacterial genomes.

    Science.gov (United States)

    Prabha, Ratna; Singh, Dhananjaya P; Sinha, Swati; Ahmad, Khurshid; Rai, Anil

    2017-04-01

    With the increasing accumulation of genomic sequence information of prokaryotes, the study of codon usage bias has gained renewed attention. The purpose of this study was to examine codon selection pattern within and across cyanobacterial species belonging to diverse taxonomic orders and habitats. We performed detailed comparative analysis of cyanobacterial genomes with respect to codon bias. Our analysis reflects that in cyanobacterial genomes, A- and/or T-ending codons were used predominantly in the genes whereas G- and/or C-ending codons were largely avoided. Variation in the codon context usage of cyanobacterial genes corresponded to the clustering of cyanobacteria as per their GC content. Analysis of codon adaptation index (CAI) and synonymous codon usage order (SCUO) revealed that majority of genes are associated with low codon bias. Codon selection pattern in cyanobacterial genomes reflected compositional constraints as major influencing factor. It is also identified that although, mutational constraint may play some role in affecting codon usage bias in cyanobacteria, compositional constraint in terms of genomic GC composition coupled with environmental factors affected codon selection pattern in cyanobacterial genomes. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Genome and transcriptome analysis of the food-yeast Candida utilis.

    Directory of Open Access Journals (Sweden)

    Yasuyuki Tomita

    Full Text Available The industrially important food-yeast Candida utilis is a Crabtree effect-negative yeast used to produce valuable chemicals and recombinant proteins. In the present study, we conducted whole genome sequencing and phylogenetic analysis of C. utilis, which showed that this yeast diverged long before the formation of the CUG and Saccharomyces/Kluyveromyces clades. In addition, we performed comparative genome and transcriptome analyses using next-generation sequencing, which resulted in the identification of genes important for characteristic phenotypes of C. utilis such as those involved in nitrate assimilation, in addition to the gene encoding the functional hexose transporter. We also found that an antisense transcript of the alcohol dehydrogenase gene, which in silico analysis did not predict to be a functional gene, was transcribed in the stationary-phase, suggesting a novel system of repression of ethanol production. These findings should facilitate the development of more sophisticated systems for the production of useful reagents using C. utilis.

  13. RESEARCH NOTE Genome-based exome-sequencing analysis ...

    Indian Academy of Sciences (India)

    Navya

    2017-02-22

    Feb 22, 2017 ... Genome-based exome-sequencing analysis identifies GYG1, DIS3L, DDRGK1 genes ... Cardiology Division, Department of Internal Medicine, Severance .... with p values of <0.05 byanalyzing differences in allele distribution.

  14. Genome inventory and analysis of nuclear hormone receptors in ...

    Indian Academy of Sciences (India)

    Prakash

    2006-12-20

    Dec 20, 2006 ... progestins, as well as lipids, cholesterol metabolites, and. Genome ... Gene structure analysis shows strong conservation of exon structures among orthologoues. ..... earlier subfamily classification of NRs (Nuclear Receptors.

  15. Human · mouse genome analysis and radiation biology. Proceedings

    International Nuclear Information System (INIS)

    Hori, Tada-aki

    1994-03-01

    This issue is the collection of the papers presented at the 25th NIRS symposium on Human, Mouse Genome Analysis and Radiation Biology. The 14 of the presented papers are indexed individually. (J.P.N.)

  16. Sequence and transcription analysis of the human cytomegalovirus DNA polymerase gene

    International Nuclear Information System (INIS)

    Kouzarides, T.; Bankier, A.T.; Satchwell, S.C.; Weston, K.; Tomlinson, P.; Barrell, B.G.

    1987-01-01

    DNA sequence analysis has revealed that the gene coding for the human cytomegalovirus (HCMV) DNA polymerase is present within the long unique region of the virus genome. Identification is based on extensive amino acid homology between the predicted HCMV open reading frame HFLF2 and the DNA polymerase of herpes simplex virus type 1. The authors present here a 5280 base-pair DNA sequence containing the HCMV pol gene, along with the analysis of transcripts encoded within this region. Since HCMV pol also shows homology to the predicted Epstein-Barr virus pol, they were able to analyze the extent of homology between the DNA polymerases of three distantly related herpes viruses, HCMV, Epstein-Barr virus, and herpes simplex virus. The comparison shows that these DNA polymerases exhibit considerable amino acid homology and highlights a number of highly conserved regions; two such regions show homology to sequences within the adenovirus type 2 DNA polymerase. The HCMV pol gene is flanked by open reading frames with homology to those of other herpes viruses; upstream, there is a reading frame homologous to the glycoprotein B gene of herpes simplex virus type I and Epstein-Barr virus, and downstream there is a reading frame homologous to BFLF2 of Epstein-Barr virus

  17. Comparative analysis of rosaceous genomes and the reconstruction of a putative ancestral genome for the family.

    Science.gov (United States)

    Illa, Eudald; Sargent, Daniel J; Lopez Girona, Elena; Bushakra, Jill; Cestaro, Alessandro; Crowhurst, Ross; Pindo, Massimo; Cabrera, Antonio; van der Knaap, Esther; Iezzoni, Amy; Gardiner, Susan; Velasco, Riccardo; Arús, Pere; Chagné, David; Troggio, Michela

    2011-01-12

    Comparative genome mapping studies in Rosaceae have been conducted until now by aligning genetic maps within the same genus, or closely related genera and using a limited number of common markers. The growing body of genomics resources and sequence data for both Prunus and Fragaria permits detailed comparisons between these genera and the recently released Malus × domestica genome sequence. We generated a comparative analysis using 806 molecular markers that are anchored genetically to the Prunus and/or Fragaria reference maps, and physically to the Malus genome sequence. Markers in common for Malus and Prunus, and Malus and Fragaria, respectively were 784 and 148. The correspondence between marker positions was high and conserved syntenic blocks were identified among the three genera in the Rosaceae. We reconstructed a proposed ancestral genome for the Rosaceae. A genome containing nine chromosomes is the most likely candidate for the ancestral Rosaceae progenitor. The number of chromosomal translocations observed between the three genera investigated was low. However, the number of inversions identified among Malus and Prunus was much higher than any reported genome comparisons in plants, suggesting that small inversions have played an important role in the evolution of these two genera or of the Rosaceae.

  18. Comparative analysis of rosaceous genomes and the reconstruction of a putative ancestral genome for the family

    Directory of Open Access Journals (Sweden)

    Velasco Riccardo

    2011-01-01

    Full Text Available Abstract Background Comparative genome mapping studies in Rosaceae have been conducted until now by aligning genetic maps within the same genus, or closely related genera and using a limited number of common markers. The growing body of genomics resources and sequence data for both Prunus and Fragaria permits detailed comparisons between these genera and the recently released Malus × domestica genome sequence. Results We generated a comparative analysis using 806 molecular markers that are anchored genetically to the Prunus and/or Fragaria reference maps, and physically to the Malus genome sequence. Markers in common for Malus and Prunus, and Malus and Fragaria, respectively were 784 and 148. The correspondence between marker positions was high and conserved syntenic blocks were identified among the three genera in the Rosaceae. We reconstructed a proposed ancestral genome for the Rosaceae. Conclusions A genome containing nine chromosomes is the most likely candidate for the ancestral Rosaceae progenitor. The number of chromosomal translocations observed between the three genera investigated was low. However, the number of inversions identified among Malus and Prunus was much higher than any reported genome comparisons in plants, suggesting that small inversions have played an important role in the evolution of these two genera or of the Rosaceae.

  19. Genome-wide transcriptional profiling of skin and dorsal root ganglia after ultraviolet-B-induced inflammation.

    Directory of Open Access Journals (Sweden)

    John M Dawes

    Full Text Available Ultraviolet-B (UVB-induced inflammation produces a dose-dependent mechanical and thermal hyperalgesia in both humans and rats, most likely via inflammatory mediators acting at the site of injury. Previous work has shown that the gene expression of cytokines and chemokines is positively correlated between species and that these factors can contribute to UVB-induced pain. In order to investigate other potential pain mediators in this model we used RNA-seq to perform genome-wide transcriptional profiling in both human and rat skin at the peak of hyperalgesia. In addition we have also measured transcriptional changes in the L4 and L5 DRG of the rat model. Our data show that UVB irradiation produces a large number of transcriptional changes in the skin: 2186 and 3888 genes are significantly dysregulated in human and rat skin, respectively. The most highly up-regulated genes in human skin feature those encoding cytokines (IL6 and IL24, chemokines (CCL3, CCL20, CXCL1, CXCL2, CXCL3 and CXCL5, the prostanoid synthesising enzyme COX-2 and members of the keratin gene family. Overall there was a strong positive and significant correlation in gene expression between the human and rat (R = 0.8022. In contrast to the skin, only 39 genes were significantly dysregulated in the rat L4 and L5 DRGs, the majority of which had small fold change values. Amongst the most up-regulated genes in DRG were REG3B, CCL2 and VGF. Overall, our data shows that numerous genes were up-regulated in UVB irradiated skin at the peak of hyperalgesia in both human and rats. Many of the top up-regulated genes were cytokines and chemokines, highlighting again their potential as pain mediators. However many other genes were also up-regulated and might play a role in UVB-induced hyperalgesia. In addition, the strong gene expression correlation between species re-emphasises the value of the UVB model as translational tool to study inflammatory pain.

  20. Comparative Pan-Genome Analysis of Piscirickettsia salmonis Reveals Genomic Divergences within Genogroups

    Directory of Open Access Journals (Sweden)

    Guillermo Nourdin-Galindo

    2017-10-01

    Full Text Available Piscirickettsia salmonis is the etiological agent of salmonid rickettsial septicemia, a disease that seriously affects the salmonid industry. Despite efforts to genomically characterize P. salmonis, functional information on the life cycle, pathogenesis mechanisms, diagnosis, treatment, and control of this fish pathogen remain lacking. To address this knowledge gap, the present study conducted an in silico pan-genome analysis of 19 P. salmonis strains from distinct geographic locations and genogroups. Results revealed an expected open pan-genome of 3,463 genes and a core-genome of 1,732 genes. Two marked genogroups were identified, as confirmed by phylogenetic and phylogenomic relationships to the LF-89 and EM-90 reference strains, as well as by assessments of genomic structures. Different structural configurations were found for the six identified copies of the ribosomal operon in the P. salmonis genome, indicating translocation throughout the genetic material. Chromosomal divergences in genomic localization and quantity of genetic cassettes were also found for the Dot/Icm type IVB secretion system. To determine divergences between core-genomes, additional pan-genome descriptions were compiled for the so-termed LF and EM genogroups. Open pan-genomes composed of 2,924 and 2,778 genes and core-genomes composed of 2,170 and 2,228 genes were respectively found for the LF and EM genogroups. The core-genomes were functionally annotated using the Gene Ontology, KEGG, and Virulence Factor databases, revealing the presence of several shared groups of genes related to basic function of intracellular survival and bacterial pathogenesis. Additionally, the specific pan-genomes for the LF and EM genogroups were defined, resulting in the identification of 148 and 273 exclusive proteins, respectively. Notably, specific virulence factors linked to adherence, colonization, invasion factors, and endotoxins were established. The obtained data suggest that these

  1. Short and long-term genome stability analysis of prokaryotic genomes.

    Science.gov (United States)

    Brilli, Matteo; Liò, Pietro; Lacroix, Vincent; Sagot, Marie-France

    2013-05-08

    Gene organization dynamics is actively studied because it provides useful evolutionary information, makes functional annotation easier and often enables to characterize pathogens. There is therefore a strong interest in understanding the variability of this trait and the possible correlations with life-style. Two kinds of events affect genome organization: on one hand translocations and recombinations change the relative position of genes shared by two genomes (i.e. the backbone gene order); on the other, insertions and deletions leave the backbone gene order unchanged but they alter the gene neighborhoods by breaking the syntenic regions. A complete picture about genome organization evolution therefore requires to account for both kinds of events. We developed an approach where we model chromosomes as graphs on which we compute different stability estimators; we consider genome rearrangements as well as the effect of gene insertions and deletions. In a first part of the paper, we fit a measure of backbone gene order conservation (hereinafter called backbone stability) against phylogenetic distance for over 3000 genome comparisons, improving existing models for the divergence in time of backbone stability. Intra- and inter-specific comparisons were treated separately to focus on different time-scales. The use of multiple genomes of a same species allowed to identify genomes with diverging gene order with respect to their conspecific. The inter-species analysis indicates that pathogens are more often unstable with respect to non-pathogens. In a second part of the text, we show that in pathogens, gene content dynamics (insertions and deletions) have a much more dramatic effect on genome organization stability than backbone rearrangements. In this work, we studied genome organization divergence taking into account the contribution of both genome order rearrangements and genome content dynamics. By studying species with multiple sequenced genomes available, we were

  2. Yeast Sub1 and human PC4 are G-quadruplex binding proteins that suppress genome instability at co-transcriptionally formed G4 DNA.

    Science.gov (United States)

    Lopez, Christopher R; Singh, Shivani; Hambarde, Shashank; Griffin, Wezley C; Gao, Jun; Chib, Shubeena; Yu, Yang; Ira, Grzegorz; Raney, Kevin D; Kim, Nayun

    2017-06-02

    G-quadruplex or G4 DNA is a non-B secondary DNA structure consisting of a stacked array of guanine-quartets that can disrupt critical cellular functions such as replication and transcription. When sequences that can adopt Non-B structures including G4 DNA are located within actively transcribed genes, the reshaping of DNA topology necessary for transcription process stimulates secondary structure-formation thereby amplifying the potential for genome instability. Using a reporter assay designed to study G4-induced recombination in the context of an actively transcribed locus in Saccharomyces cerevisiae, we tested whether co-transcriptional activator Sub1, recently identified as a G4-binding factor, contributes to genome maintenance at G4-forming sequences. Our data indicate that, upon Sub1-disruption, genome instability linked to co-transcriptionally formed G4 DNA in Top1-deficient cells is significantly augmented and that its highly conserved DNA binding domain or the human homolog PC4 is sufficient to suppress G4-associated genome instability. We also show that Sub1 interacts specifically with co-transcriptionally formed G4 DNA in vivo and that yeast cells become highly sensitivity to G4-stabilizing chemical ligands by the loss of Sub1. Finally, we demonstrate the physical and genetic interaction of Sub1 with the G4-resolving helicase Pif1, suggesting a possible mechanism by which Sub1 suppresses instability at G4 DNA. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  3. Data on genome analysis of Bacillus velezensis LS69.

    Science.gov (United States)

    Liu, Guoqiang; Kong, Yingying; Fan, Yajing; Geng, Ce; Peng, Donghai; Sun, Ming

    2017-08-01

    The data presented in this article are related to the published entitled "Whole-genome sequencing of Bacillus velezensis LS69, a strain with a broad inhibitory spectrum against pathogenic bacteria" (Liu et al., 2017) [1]. Genome analysis revealed B. velezensis LS69 has a good potential for biocontrol and plant growth promotion. This article provides an extended analysis of the genetic islands, core genes and amylolysin loci of B. velezensis LS69.

  4. Data on genome analysis of Bacillus velezensis LS69

    OpenAIRE

    Liu, Guoqiang; Kong, Yingying; Fan, Yajing; Geng, Ce; Peng, Donghai; Sun, Ming

    2017-01-01

    The data presented in this article are related to the published entitled “Whole-genome sequencing of Bacillus velezensis LS69, a strain with a broad inhibitory spectrum against pathogenic bacteria” (Liu et al., 2017) [1]. Genome analysis revealed B. velezensis LS69 has a good potential for biocontrol and plant growth promotion. This article provides an extended analysis of the genetic islands, core genes and amylolysin loci of B. velezensis LS69.

  5. Data on genome analysis of Bacillus velezensis LS69

    Directory of Open Access Journals (Sweden)

    Guoqiang Liu

    2017-08-01

    Full Text Available The data presented in this article are related to the published entitled “Whole-genome sequencing of Bacillus velezensis LS69, a strain with a broad inhibitory spectrum against pathogenic bacteria” (Liu et al., 2017 [1]. Genome analysis revealed B. velezensis LS69 has a good potential for biocontrol and plant growth promotion. This article provides an extended analysis of the genetic islands, core genes and amylolysin loci of B. velezensis LS69.

  6. Genomic Analysis of Complex Microbial Communities in Wounds

    Science.gov (United States)

    2012-01-01

    Permutation Multivariate Analysis of Variance ( PerMANOVA ). We used PerMANOVA to test the null-hypothesis of no... permutation -based version of the multivariate analysis of variance (MANOVA). PerMANOVA uses the distances between samples to partition variance and...coli. Antibiotics, bacteria, community analysis , diabetes, pyrosequencing, wound, wound therapy, 16S rRNA gene Genomic Analysis of Complex

  7. Mycobacterial species as case-study of comparative genome analysis.

    Science.gov (United States)

    Zakham, F; Belayachi, L; Ussery, D; Akrim, M; Benjouad, A; El Aouad, R; Ennaji, M M

    2011-02-08

    The genus Mycobacterium represents more than 120 species including important pathogens of human and cause major public health problems and illnesses. Further, with more than 100 genome sequences from this genus, comparative genome analysis can provide new insights for better understanding the evolutionary events of these species and improving drugs, vaccines, and diagnostics tools for controlling Mycobacterial diseases. In this present study we aim to outline a comparative genome analysis of fourteen Mycobacterial genomes: M. avium subsp. paratuberculosis K—10, M. bovis AF2122/97, M. bovis BCG str. Pasteur 1173P2, M. leprae Br4923, M. marinum M, M. sp. KMS, M. sp. MCS, M. tuberculosis CDC1551, M. tuberculosis F11, M. tuberculosis H37Ra, M. tuberculosis H37Rv, M. tuberculosis KZN 1435 , M. ulcerans Agy99,and M. vanbaalenii PYR—1, For this purpose a comparison has been done based on their length of genomes, GC content, number of genes in different data bases (Genbank, Refseq, and Prodigal). The BLAST matrix of these genomes has been figured to give a lot of information about the similarity between species in a simple scheme. As a result of multiple genome analysis, the pan and core genome have been defined for twelve Mycobacterial species. We have also introduced the genome atlas of the reference strain M. tuberculosis H37Rv which can give a good overview of this genome. And for examining the phylogenetic relationships among these bacteria, a phylogenic tree has been constructed from 16S rRNA gene for tuberculosis and non tuberculosis Mycobacteria to understand the evolutionary events of these species.

  8. Whole genome transcript profiling from fingerstick blood samples: a comparison and feasibility study

    Directory of Open Access Journals (Sweden)

    Williams Adam R

    2009-12-01

    Full Text Available Abstract Background Whole genome gene expression profiling has revolutionized research in the past decade especially with the advent of microarrays. Recently, there have been significant improvements in whole blood RNA isolation techniques which, through stabilization of RNA at the time of sample collection, avoid bias and artifacts introduced during sample handling. Despite these improvements, current human whole blood RNA stabilization/isolation kits are limited by the requirement of a venous blood sample of at least 2.5 mL. While fingerstick blood collection has been used for many different assays, there has yet to be a kit developed to isolate high quality RNA for use in gene expression studies from such small human samples. The clinical and field testing advantages of obtaining reliable and reproducible gene expression data from a fingerstick are many; it is less invasive, time saving, more mobile, and eliminates the need of a trained phlebotomist. Furthermore, this method could also be employed in small animal studies, i.e. mice, where larger sample collections often require sacrificing the animal. In this study, we offer a rapid and simple method to extract sufficient amounts of high quality total RNA from approximately 70 μl of whole blood collected via a fingerstick using a modified protocol of the commercially available Qiagen PAXgene RNA Blood Kit. Results From two sets of fingerstick collections, about 70 uL whole blood collected via finger lancet and capillary tube, we recovered an average of 252.6 ng total RNA with an average RIN of 9.3. The post-amplification yields for 50 ng of total RNA averaged at 7.0 ug cDNA. The cDNA hybridized to Affymetrix HG-U133 Plus 2.0 GeneChips had an average % Present call of 52.5%. Both fingerstick collections were highly correlated with r2 values ranging from 0.94 to 0.97. Similarly both fingerstick collections were highly correlated to the venous collection with r2 values ranging from 0.88 to 0

  9. The transcriptional landscape

    DEFF Research Database (Denmark)

    Nielsen, Henrik

    2011-01-01

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

  10. Analysis Of Transcriptomes In A Porcine Tissue Collection Using RNA-Seq And Genome Assembly 10

    DEFF Research Database (Denmark)

    Hornshøj, Henrik; Thomsen, Bo; Hedegaard, Jakob

    2011-01-01

    The release of Sus scrofa genome assembly 10 supports improvement of the pig genome annotation and in depth transcriptome analyses using next-generation sequencing technologies. In this study we analyze RNA-seq reads from a tissue collection, including 10 separate tissues from Duroc boars and 10...... short read alignment software we mapped the reads to the genome assembly 10. We extracted contig sequences of gene transcripts using the Cufflinks software. Based on this information we identified expressed genes that are present in the genome assembly. The portion of these genes being previously known...... was roughly estimated by sequence comparison to known genes. Similarly, we searched for genes that are expressed in the tissues but not present in the genome assembly by aligning the non-genome-mapped reads to known gene transcripts. For the genes predicted to have alternative transcript variants by Cufflinks...

  11. Genome-Wide Investigation of WRKY Transcription Factors Involved in Terminal Drought Stress Response in Common Bean.

    Science.gov (United States)

    Wu, Jing; Chen, Jibao; Wang, Lanfen; Wang, Shumin

    2017-01-01

    WRKY transcription factor plays a key role in drought stress. However, the characteristics of the WRKY gene family in the common bean ( Phaseolus vulgaris L.) are unknown. In this study, we identified 88 complete WRKY proteins from the draft genome sequence of the "G19833" common bean. The predicted genes were non-randomly distributed in all chromosomes. Basic information, amino acid motifs, phylogenetic tree and the expression patterns of PvWRKY genes were analyzed, and the proteins were classified into groups 1, 2, and 3. Group 2 was further divided into five subgroups: 2a, 2b, 2c, 2d, and 2e. Finally, we detected 19 WRKY genes that were responsive to drought stress using qRT-PCR; 11 were down-regulated, and 8 were up-regulated under drought stress. This study comprehensively examines WRKY proteins in the common bean, a model food legume, and it provides a foundation for the functional characterization of the WRKY family and opportunities for understanding the mechanisms of drought stress tolerance in this plant.

  12. Identification of novel candidate genes involved in mineralization of dental enamel by genome-wide transcript profiling.

    Science.gov (United States)

    Lacruz, Rodrigo S; Smith, Charles E; Bringas, Pablo; Chen, Yi-Bu; Smith, Susan M; Snead, Malcolm L; Kurtz, Ira; Hacia, Joseph G; Hubbard, Michael J; Paine, Michael L

    2012-05-01

    The gene repertoire regulating vertebrate biomineralization is poorly understood. Dental enamel, the most highly mineralized tissue in mammals, differs from other calcifying systems in that the formative cells (ameloblasts) lack remodeling activity and largely degrade and resorb the initial extracellular matrix. Enamel mineralization requires that ameloblasts undergo a profound functional switch from matrix-secreting to maturational (calcium transport, protein resorption) roles as mineralization progresses. During the maturation stage, extracellular pH decreases markedly, placing high demands on ameloblasts to regulate acidic environments present around the growing hydroxyapatite crystals. To identify the genetic events driving enamel mineralization, we conducted genome-wide transcript profiling of the developing enamel organ from rat incisors and highlight over 300 genes differentially expressed during maturation. Using multiple bioinformatics analyses, we identified groups of maturation-associated genes whose functions are linked to key mineralization processes including pH regulation, calcium handling, and matrix turnover. Subsequent qPCR and Western blot analyses revealed that a number of solute carrier (SLC) gene family members were up-regulated during maturation, including the novel protein Slc24a4 involved in calcium handling as well as other proteins of similar function (Stim1). By providing the first global overview of the cellular machinery required for enamel maturation, this study provide a strong foundation for improving basic understanding of biomineralization and its practical applications in healthcare. Copyright © 2011 Wiley Periodicals, Inc.

  13. Analysis of carboxylesterase 2 transcript variants in cynomolgus macaque liver.

    Science.gov (United States)

    Uno, Yasuhiro; Igawa, Yoshiyuki; Tanaka, Maori; Ohura, Kayoko; Hosokawa, Masakiyo; Imai, Teruko

    2018-04-27

    Carboxylesterase (CES) is important for the detoxification of a wide range of drugs and xenobiotics. In this study, the hepatic level of CES2 mRNA was examined in cynomolgus macaques used widely in preclinical studies for drug metabolism. Three CES2 mRNAs were present in cynomolgus macaque liver. The mRNA level was highest for cynomolgus CES2A (formerly CES2v3), much lower for cynomolgus CES2B (formerly CES2v1) and extremely low for cynomolgus CES2C (formerly CES2v2). Most various transcript variants produced from cynomolgus CES2B gene did not contain a complete coding region. Thus, CES2A is the major CES2 enzyme in cynomolgus liver. A new transcript variant of CES2A, CES2Av2, was identified. CES2Av2 contained exon 3 region different from wild-type (CES2Av1). In cynomolgus macaques expressing only CES2Av2 transcript, CES2A contained the sequence of CES2B in exon 3 and vicinity, probably due to gene conversion. On genotyping, this CES2Av2 allele was prevalent in Indochinese cynomolgus macaques, but not in Indonesian cynomolgus or rhesus macaques. CES2Av2 recombinant protein showed similar activity to CES2Av1 protein for several substrates. It is concluded that CES2A is the major cynomolgus hepatic CES2, and new transcript variant, CES2Av2, has similar functions to CES2Av1.

  14. Applied bioinformatics: Genome annotation and transcriptome analysis

    DEFF Research Database (Denmark)

    Gupta, Vikas

    agricultural and biological importance. Its capacity to form symbiotic relationships with rhizobia and microrrhizal fungi has fascinated researchers for years. Lotus has a small genome of approximately 470 Mb and a short life cycle of 2 to 3 months, which has made Lotus a model legume plant for many molecular...

  15. Comparative genome analysis of trypanotolerance QTL | Nganga ...

    African Journals Online (AJOL)

    Homologous sequences were used in the definition of synteny relationships and subsequent identification of the shared disease response genes. The homologous genes within the human genome were then identified and aligned to the bovine radiation hybrid map in order to identify the mouse/bovine homologous regions.

  16. Cellular promoters incorporated into the adenovirus genome: effects of viral regulatory elements on transcription rates and cell specificity of albumin and beta-globin promoters.

    OpenAIRE

    Babiss, L E; Friedman, J M; Darnell, J E

    1986-01-01

    In the accompanying paper (Friedman et al., Mol. Cell. Biol. 6:3791-3797, 1986), hepatoma-specific expression of the rat albumin promoter within the adenovirus genome was demonstrated. However, the rate of transcription was very low compared with that of the endogenous chromosomal albumin gene. Here we show that in hepatoma cells the adenovirus E1A enhancer, especially in the presence of E1A protein, greatly stimulates transcription from the albumin promoter but not the mouse beta-globin prom...

  17. Genome analysis methods - PGDBj Registered plant list, Marker list, QTL list, Plant DB link & Genome analysis methods | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available List Contact us PGDBj Registered plant list, Marker list, QTL list, Plant DB link & Genome analysis methods Genome analysis... methods Data detail Data name Genome analysis methods DOI 10.18908/lsdba.nbdc01194-01-005 De...scription of data contents The current status and related information of the genomic analysis about each org...anism (March, 2014). In the case of organisms carried out genomic analysis, the d...e File name: pgdbj_dna_marker_linkage_map_genome_analysis_methods_en.zip File URL: ftp://ftp.biosciencedbc.j

  18. REGIA, An EU Project on Functional Genomics of Transcription Factors from Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Javier Paz-Ares

    2002-01-01

    and metabolic profiling; 5. the systematic analysis of interactions between TFs; and 6. the generation of a bioinformatics infrastructure to access and integrate all this information. We expect that this programme will establish the full biotechnological potential of plant TFs, and provide insights into hierarchies, redundancies, and interdependencies, and their evolution. The project involves the preparation of both a TF gene array for expression analysis and a normalised full length open reading frame (ORF library of TFs in a yeast two hybrid vector; the applications of these resources should extend beyond the scope of this programme.

  19. Genome-wide investigation and transcriptome analysis of the WRKY gene family in Gossypium.

    Science.gov (United States)

    Ding, Mingquan; Chen, Jiadong; Jiang, Yurong; Lin, Lifeng; Cao, YueFen; Wang, Minhua; Zhang, Yuting; Rong, Junkang; Ye, Wuwei

    2015-02-01

    WRKY transcription factors play important roles in various stress responses in diverse plant species. In cotton, this family has not been well studied, especially in relation to fiber development. Here, the genomes and transcriptomes of Gossypium raimondii and Gossypium arboreum were investigated to identify fiber development related WRKY genes. This represents the first comprehensive comparative study of WRKY transcription factors in both diploid A and D cotton species. In total, 112 G. raimondii and 109 G. arboreum WRKY genes were identified. No significant gene structure or domain alterations were detected between the two species, but many SNPs distributed unequally in exon and intron regions. Physical mapping revealed that the WRKY genes in G. arboreum were not located in the corresponding chromosomes of G. raimondii, suggesting great chromosome rearrangement in the diploid cotton genomes. The cotton WRKY genes, especially subgroups I and II, have expanded through multiple whole genome duplications and tandem duplications compared with other plant species. Sequence comparison showed many functionally divergent sites between WRKY subgroups, while the genes within each group are under strong purifying selection. Transcriptome analysis suggested that many WRKY genes participate in specific fiber development processes such as fiber initiation, elongation and maturation with different expression patterns between species. Complex WRKY gene expression such as differential Dt and At allelic gene expression in G. hirsutum and alternative splicing events were also observed in both diploid and tetraploid cottons during fiber development process. In conclusion, this study provides important information on the evolution and function of WRKY gene family in cotton species.

  20. Integrative analysis of histone ChIP-seq and transcription data using Bayesian mixture models

    DEFF Research Database (Denmark)

    Klein, Hans-Ulrich; Schäfer, Martin; Porse, Bo T

    2014-01-01

    Histone modifications are a key epigenetic mechanism to activate or repress the transcription of genes. Datasets of matched transcription data and histone modification data obtained by ChIP-seq exist, but methods for integrative analysis of both data types are still rare. Here, we present a novel...

  1. RNA transcriptional biosignature analysis for identifying febrile infants with serious bacterial infections in the emergency department: a feasibility study.

    Science.gov (United States)

    Mahajan, Prashant; Kuppermann, Nathan; Suarez, Nicolas; Mejias, Asuncion; Casper, Charlie; Dean, J Michael; Ramilo, Octavio

    2015-01-01

    To develop the infrastructure and demonstrate the feasibility of conducting microarray-based RNA transcriptional profile analyses for the diagnosis of serious bacterial infections in febrile infants 60 days and younger in a multicenter pediatric emergency research network. We designed a prospective multicenter cohort study with the aim of enrolling more than 4000 febrile infants 60 days and younger. To ensure success of conducting complex genomic studies in emergency department (ED) settings, we established an infrastructure within the Pediatric Emergency Care Applied Research Network, including 21 sites, to evaluate RNA transcriptional profiles in young febrile infants. We developed a comprehensive manual of operations and trained site investigators to obtain and process blood samples for RNA extraction and genomic analyses. We created standard operating procedures for blood sample collection, processing, storage, shipping, and analyses. We planned to prospectively identify, enroll, and collect 1 mL blood samples for genomic analyses from eligible patients to identify logistical issues with study procedures. Finally, we planned to batch blood samples and determined RNA quantity and quality at the central microarray laboratory and organized data analysis with the Pediatric Emergency Care Applied Research Network data coordinating center. Below we report on establishment of the infrastructure and the feasibility success in the first year based on the enrollment of a limited number of patients. We successfully established the infrastructure at 21 EDs. Over the first 5 months we enrolled 79% (74 of 94) of eligible febrile infants. We were able to obtain and ship 1 mL of blood from 74% (55 of 74) of enrolled participants, with at least 1 sample per participating ED. The 55 samples were shipped and evaluated at the microarray laboratory, and 95% (52 of 55) of blood samples were of adequate quality and contained sufficient RNA for expression analysis. It is possible to

  2. Network based transcription factor analysis of regenerating axolotl limbs

    Directory of Open Access Journals (Sweden)

    Cameron Jo Ann

    2011-03-01

    Full Text Available Abstract Background Studies on amphibian limb regeneration began in the early 1700's but we still do not completely understand the cellular and molecular events of this unique process. Understanding a complex biological process such as limb regeneration is more complicated than the knowledge of the individual genes or proteins involved. Here we followed a systems biology approach in an effort to construct the networks and pathways of protein interactions involved in formation of the accumulation blastema in regenerating axolotl limbs. Results We used the human orthologs of proteins previously identified by our research team as bait to identify the transcription factor (TF pathways and networks that regulate blastema formation in amputated axolotl limbs. The five most connected factors, c-Myc, SP1, HNF4A, ESR1 and p53 regulate ~50% of the proteins in our data. Among these, c-Myc and SP1 regulate 36.2% of the proteins. c-Myc was the most highly connected TF (71 targets. Network analysis showed that TGF-β1 and fibronectin (FN lead to the activation of these TFs. We found that other TFs known to be involved in epigenetic reprogramming, such as Klf4, Oct4, and Lin28 are also connected to c-Myc and SP1. Conclusions Our study provides a systems biology approach to how different molecular entities inter-connect with each other during the formation of an accumulation blastema in regenerating axolotl limbs. This approach provides an in silico methodology to identify proteins that are not detected by experimental methods such as proteomics but are potentially important to blastema formation. We found that the TFs, c-Myc and SP1 and their target genes could potentially play a central role in limb regeneration. Systems biology has the potential to map out numerous other pathways that are crucial to blastema formation in regeneration-competent limbs, to compare these to the pathways that characterize regeneration-deficient limbs and finally, to identify stem

  3. Genome-wide analysis of poly(A) site selection in Schizosaccharomyces pombe

    KAUST Repository

    Schlackow, M.

    2013-10-23

    Polyadenylation of pre-mRNAs, a critical step in eukaryotic gene expression, is mediated by cis elements collectively called the polyadenylation signal. Genome-wide analysis of such polyadenylation signals was missing in fission yeast, even though it is an important model organism. We demonstrate that the canonical AATAAA motif is the most frequent and functional polyadenylation signal in Schizosaccharomyces pombe. Using analysis of RNA-Seq data sets from cells grown under various physiological conditions, we identify 3\\' UTRs for nearly 90% of the yeast genes. Heterogeneity of cleavage sites is common, as is alternative polyadenylation within and between conditions. We validated the computationally identified sequence elements likely to promote polyadenylation by functional assays, including qRT-PCR and 3\\'RACE analysis. The biological importance of the AATAAA motif is underlined by functional analysis of the genes containing it. Furthermore, it has been shown that convergent genes require trans elements, like cohesin for efficient transcription termination. Here we show that convergent genes lacking cohesin (on chromosome 2) are generally associated with longer overlapping mRNA transcripts. Our bioinformatic and experimental genome-wide results are summarized and can be accessed and customized in a user-friendly database Pomb(A).

  4. Genome-wide analysis of poly(A) site selection in Schizosaccharomyces pombe

    KAUST Repository

    Schlackow, M.; Marguerat, S.; Proudfoot, N. J.; Bahler, J.; Erban, R.; Gullerova, M.

    2013-01-01

    Polyadenylation of pre-mRNAs, a critical step in eukaryotic gene expression, is mediated by cis elements collectively called the polyadenylation signal. Genome-wide analysis of such polyadenylation signals was missing in fission yeast, even though it is an important model organism. We demonstrate that the canonical AATAAA motif is the most frequent and functional polyadenylation signal in Schizosaccharomyces pombe. Using analysis of RNA-Seq data sets from cells grown under various physiological conditions, we identify 3' UTRs for nearly 90% of the yeast genes. Heterogeneity of cleavage sites is common, as is alternative polyadenylation within and between conditions. We validated the computationally identified sequence elements likely to promote polyadenylation by functional assays, including qRT-PCR and 3'RACE analysis. The biological importance of the AATAAA motif is underlined by functional analysis of the genes containing it. Furthermore, it has been shown that convergent genes require trans elements, like cohesin for efficient transcription termination. Here we show that convergent genes lacking cohesin (on chromosome 2) are generally associated with longer overlapping mRNA transcripts. Our bioinformatic and experimental genome-wide results are summarized and can be accessed and customized in a user-friendly database Pomb(A).

  5. Genome-wide analysis of Polycomb targets in Drosophila

    Energy Technology Data Exchange (ETDEWEB)

    Schwartz, Yuri B.; Kahn, Tatyana G.; Nix, David A.; Li,Xiao-Yong; Bourgon, Richard; Biggin, Mark; Pirrotta, Vincenzo

    2006-04-01

    Polycomb Group (PcG) complexes are multiprotein assemblages that bind to chromatin and establish chromatin states leading to epigenetic silencing. PcG proteins regulate homeotic genes in flies and vertebrates but little is known about other PcG targets and the role of the PcG in development, differentiation and disease. We have determined the distribution of the PcG proteins PC, E(Z) and PSC and of histone H3K27 trimethylation in the Drosophila genome. At more than 200 PcG target genes, binding sites for the three PcG proteins colocalize to presumptive Polycomb Response Elements (PREs). In contrast, H3 me3K27 forms broad domains including the entire transcription unit and regulatory regions. PcG targets are highly enriched in genes encoding transcription factors but receptors, signaling proteins, morphogens and regulators representing all major developmental pathways are also included.

  6. Genome Sequencing and Comparative Analysis of the Biocontrol Agent Trichoderma harzianum sensu stricto TR274

    Energy Technology Data Exchange (ETDEWEB)

    Steindorff, Andrei S.; Noronha, Elilane F.; Ulhoa, Cirano J.; Kuo, Alan; Salamov, Asaf A.; Haridas, Sajeet; Riley, Robert W.; Druzhinina, Irina S.; Kubicek, Christian P.; Grigoriev, Igor V.

    2015-03-17

    Biological control is a complex process which requires many mechanisms and a high diversity of biochemical pathways. The species of Trichoderma harzianum are well known for their biocontrol activity against many plant pathogens. To gain new insights into the biocontrol mechanism used by T. harzianum, we sequenced the isolate TR274 genome using Illumina. The assembly was performed using AllPaths-LG with a maximum coverage of 100x. The assembly resulted in 2282 contigs with a N50 of 37033bp. The genome size generated was 40.8 Mb and the GC content was 47.7%, similar to other Trichoderma genomes. Using the JGI Annotation Pipeline we predicted 13,932 genes with a high transcriptome support. CEGMA tests suggested 100% genome completeness and 97.9% of RNA-SEQ reads were mapped to the genome. The phylogenetic comparison using orthologous proteins with all Trichoderma genomes sequenced at JGI, corroborates the Trichoderma (T. asperellum and T. atroviride), Longibrachiatum (T. reesei and T. longibrachiatum) and Pachibasium (T. harzianum and T. virens) section division described previously. The comparison between two Trichoderma harzianum species suggests a high genome similarity but some strain-specific expansions. Analyses of the secondary metabolites, CAZymes, transporters, proteases, transcription factors were performed. The Pachybasium section expanded virtually all categories analyzed compared with the other sections, specially Longibrachiatum section, that shows a clear contraction. These results suggests that these proteins families have an important role in their respective phenotypes. Future analysis will improve the understanding of this complex genus and give some insights about its lifestyle and the interactions with the environment.

  7. QTL Analysis and Functional Genomics of Animal Model

    DEFF Research Database (Denmark)

    Farajzadeh, Leila

    , for example, has enabled scientists to examine more complex interactions in connection with studies of properties and diseases. In her PhD project, Leila Farajzadeh integrated different organisational levels in biology, including genotype, phenotype, association studies, transcription profiles and genetic......In recent years, the use of functional genomics and next-generation sequencing technologies has increased the probability of success in studies of complex properties. The integration of large data sets from association studies, DNA resequencing, gene expression profiles and phenotypic data...

  8. Comparative analysis of the mitochondrial genomes in gastropods

    International Nuclear Information System (INIS)

    Arquez, Moises; Uribe, Juan Esteban; Castro, Lyda Raquel

    2012-01-01

    In this work we presented a comparative analysis of the mitochondrial genomes in gastropods. Nucleotide and amino acids composition was calculated and a comparative visual analysis of the start and termination codons was performed. The organization of the genome was compared calculating the number of intergenic sequences, the location of the genes and the number of reorganized genes (breakpoints) in comparison with the sequence that is presumed to be ancestral for the group. In order to calculate variations in the rates of molecular evolution within the group, the relative rate test was performed. In spite of the differences in the size of the genomes, the amino acids number is conserved. The nucleotide and amino acid composition is similar between Vetigastropoda, Ceanogastropoda and Neritimorpha in comparison to Heterobranchia and Patellogastropoda. The mitochondrial genomes of the group are very compact with few intergenic sequences, the only exception is the genome of Patellogastropoda with 26,828 bp. Start codons of the Heterobranchia and Patellogastropoda are very variable and there is also an increase in genome rearrangements for these two groups. Generally, the hypothesis of constant rates of molecular evolution between the groups is rejected, except when the genomes of Caenogastropoda and Vetigastropoda are compared.

  9. MIPS: analysis and annotation of proteins from whole genomes.

    Science.gov (United States)

    Mewes, H W; Amid, C; Arnold, R; Frishman, D; Güldener, U; Mannhaupt, G; Münsterkötter, M; Pagel, P; Strack, N; Stümpflen, V; Warfsmann, J; Ruepp, A

    2004-01-01

    The Munich Information Center for Protein Sequences (MIPS-GSF), Neuherberg, Germany, provides protein sequence-related information based on whole-genome analysis. The main focus of the work is directed toward the systematic organization of sequence-related attributes as gathered by a variety of algorithms, primary information from experimental data together with information compiled from the scientific literature. MIPS maintains automatically generated and manually annotated genome-specific databases, develops systematic classification schemes for the functional annotation of protein sequences and provides tools for the comprehensive analysis of protein sequences. This report updates the information on the yeast genome (CYGD), the Neurospora crassa genome (MNCDB), the database of complete cDNAs (German Human Genome Project, NGFN), the database of mammalian protein-protein interactions (MPPI), the database of FASTA homologies (SIMAP), and the interface for the fast retrieval of protein-associated information (QUIPOS). The Arabidopsis thaliana database, the rice database, the plant EST databases (MATDB, MOsDB, SPUTNIK), as well as the databases for the comprehensive set of genomes (PEDANT genomes) are described elsewhere in the 2003 and 2004 NAR database issues, respectively. All databases described, and the detailed descriptions of our projects can be accessed through the MIPS web server (http://mips.gsf.de).

  10. Transcriptional analysis of genetic region RvD1 of Mycobacterium bovis

    Directory of Open Access Journals (Sweden)

    Víctor Manuel Tibatá R.

    2004-07-01

    Full Text Available Mycobacterium bovis, shares 99.9% of genomic identity with M. tuberculosis, M. africanum and M. microti. Within this 0.1 % of difference, there are two genetic regions characteristics of M. bovis that are deleted in M. tuberculo­sis H37Rv: RvD1 and RvD2. According to bioinformatic analysis, these regions contain Open Reading Frames (ORFs. With the purpose of determining if the RvD1 region transcribes the ORFs predicted by bioinformatics (ORF1, ORF2 and Rv2024; total RNA was extracted from a culture of M. bovis BCG Pasteur, at different time points along the growth curve. The RNA samples were analyzed by Real Time Reverse Transcription - Poly-merase Chain Reaction (RTq-PCR. The findings show that ORF1, ORF2 and Rv2024, were transcribed consti-tutively, something that has not been reported previously. These results are a first step in order to determine the function of M. bovis RvD1 region, its possible role in pathogenesis and its interaction with both cattle and humans. Key words: Mycobacterium bovis, BCG, RNA, Real Time, RT-PCR, RvD1

  11. Molecular Evolution and Expansion Analysis of the NAC Transcription Factor in Zea mays

    Science.gov (United States)

    Fan, Kai; Wang, Ming; Miao, Ying; Ni, Mi; Bibi, Noreen; Yuan, Shuna; Li, Feng; Wang, Xuede

    2014-01-01

    NAC (NAM, ATAF1, 2 and CUC2) family is a plant-specific transcription factor and it controls various plant developmental processes. In the current study, 124 NAC members were identified in Zea mays and were phylogenetically clustered into 13 distinct subfamilies. The whole genome duplication (WGD), especially an additional WGD event, may lead to expanding ZmNAC members. Different subfamily has different expansion rate, and NAC subfamily preference was found during the expansion in maize. Moreover, the duplication events might occur after the divergence of the lineages of Z. mays and S. italica, and segmental duplication seemed to be the dominant pattern for the gene duplication in maize. Furthermore, the expansion of ZmNAC members may be also related to gain and loss of introns. Besides, the restriction of functional divergence was discovered after most of the gene duplication events. These results could provide novel insights into molecular evolution and expansion analysis of NAC family in maize, and advance the NAC researches in other plants, especially polyploid plants. PMID:25369196

  12. Stochasticity in the enterococcal sex pheromone response revealed by quantitative analysis of transcription in single cells.

    Science.gov (United States)

    Breuer, Rebecca J; Bandyopadhyay, Arpan; O'Brien, Sofie A; Barnes, Aaron M T; Hunter, Ryan C; Hu, Wei-Shou; Dunny, Gary M

    2017-07-01

    In Enterococcus faecalis, sex pheromone-mediated transfer of antibiotic resistance plasmids can occur under unfavorable conditions, for example, when inducing pheromone concentrations are low and inhibiting pheromone concentrations are high. To better understand this paradox, we adapted fluorescence in situ hybridization chain reaction (HCR) methodology for simultaneous quantification of multiple E. faecalis transcripts at the single cell level. We present direct evidence for variability in the minimum period, maximum response level, and duration of response of individual cells to a specific inducing condition. Tracking of induction patterns of single cells temporally using a fluorescent reporter supported HCR findings. It also revealed subpopulations of rapid responders, even under low inducing pheromone concentrations where the overall response of the entire population was slow. The strong, rapid induction of small numbers of cells in cultures exposed to low pheromone concentrations is in agreement with predictions of a stochastic model of the enterococcal pheromone response. The previously documented complex regulatory circuitry controlling the pheromone response likely contributes to stochastic variation in this system. In addition to increasing our basic understanding of the biology of a horizontal gene transfer system regulated by cell-cell signaling, demonstration of the stochastic nature of the pheromone response also impacts any future efforts to develop therapeutic agents targeting the system. Quantitative single cell analysis using HCR also has great potential to elucidate important bacterial regulatory mechanisms not previously amenable to study at the single cell level, and to accelerate the pace of functional genomic studies.

  13. A Mitochondrial Genome of Rhyparochromidae (Hemiptera: Heteroptera) and a Comparative Analysis of Related Mitochondrial Genomes.

    Science.gov (United States)

    Li, Teng; Yang, Jie; Li, Yinwan; Cui, Ying; Xie, Qiang; Bu, Wenjun; Hillis, David M

    2016-10-19

    The Rhyparochromidae, the largest family of Lygaeoidea, encompasses more than 1,850 described species, but no mitochondrial genome has been sequenced to date. Here we describe the first mitochondrial genome for Rhyparochromidae: a complete mitochondrial genome of Panaorus albomaculatus (Scott, 1874). This mitochondrial genome is comprised of 16,345 bp, and contains the expected 37 genes and control region. The majority of the control region is made up of a large tandem-repeat region, which has a novel pattern not previously observed in other insects. The tandem-repeats region of P. albomaculatus consists of 53 tandem duplications (including one partial repeat), which is the largest number of tandem repeats among all the known insect mitochondrial genomes. Slipped-strand mispairing during replication is likely to have generated this novel pattern of tandem repeats. Comparative analysis of tRNA gene families in sequenced Pentatomomorpha and Lygaeoidea species shows that the pattern of nucleotide conservation is markedly higher on the J-strand. Phylogenetic reconstruction based on mitochondrial genomes suggests that Rhyparochromidae is not the sister group to all the remaining Lygaeoidea, and supports the monophyly of Lygaeoidea.

  14. COGNAT: a web server for comparative analysis of genomic neighborhoods.

    Science.gov (United States)

    Klimchuk, Olesya I; Konovalov, Kirill A; Perekhvatov, Vadim V; Skulachev, Konstantin V; Dibrova, Daria V; Mulkidjanian, Armen Y

    2017-11-22

    In prokaryotic genomes, functionally coupled genes can be organized in conserved gene clusters enabling their coordinated regulation. Such clusters could contain one or several operons, which are groups of co-transcribed genes. Those genes that evolved from a common ancestral gene by speciation (i.e. orthologs) are expected to have similar genomic neighborhoods in different organisms, whereas those copies of the gene that are responsible for dissimilar functions (i.e. paralogs) could be found in dissimilar genomic contexts. Comparative analysis of genomic neighborhoods facilitates the prediction of co-regulated genes and helps to discern different functions in large protein families. We intended, building on the attribution of gene sequences to the clusters of orthologous groups of proteins (COGs), to provide a method for visualization and comparative analysis of genomic neighborhoods of evolutionary related genes, as well as a respective web server. Here we introduce the COmparative Gene Neighborhoods Analysis Tool (COGNAT), a web server for comparative analysis of genomic neighborhoods. The tool is based on the COG database, as well as the Pfam protein families database. As an example, we show the utility of COGNAT in identifying a new type of membrane protein complex that is formed by paralog(s) of one of the membrane subunits of the NADH:quinone oxidoreductase of type 1 (COG1009) and a cytoplasmic protein of unknown function (COG3002). This article was reviewed by Drs. Igor Zhulin, Uri Gophna and Igor Rogozin.

  15. Comparative genomic analysis of Drosophila melanogaster and vector mosquito developmental genes.

    Directory of Open Access Journals (Sweden)

    Susanta K Behura

    Full Text Available Genome sequencing projects have presented the opportunity for analysis of developmental genes in three vector mosquito species: Aedes aegypti, Culex quinquefasciatus, and Anopheles gambiae. A comparative genomic analysis of developmental genes in Drosophila melanogaster and these three important vectors of human disease was performed in this investigation. While the study was comprehensive, special emphasis centered on genes that 1 are components of developmental signaling pathways, 2 regulate fundamental developmental processes, 3 are critical for the development of tissues of vector importance, 4 function in developmental processes known to have diverged within insects, and 5 encode microRNAs (miRNAs that regulate developmental transcripts in Drosophila. While most fruit fly developmental genes are conserved in the three vector mosquito species, several genes known to be critical for Drosophila development were not identified in one or more mosquito genomes. In other cases, mosquito lineage-specific gene gains with respect to D. melanogaster were noted. Sequence analyses also revealed that numerous repetitive sequences are a common structural feature of Drosophila and mosquito developmental genes. Finally, analysis of predicted miRNA binding sites in fruit fly and mosquito developmental genes suggests that the repertoire of developmental genes targeted by miRNAs is species-specific. The results of this study provide insight into the evolution of developmental genes and processes in dipterans and other arthropods, serve as a resource for those pursuing analysis of mosquito development, and will promote the design and refinement of functional analysis experiments.

  16. Genome-wide analysis of Dongxiang wild rice (Oryza rufipogon Griff.) to investigate lost/acquired genes during rice domestication.

    Science.gov (United States)

    Zhang, Fantao; Xu, Tao; Mao, Linyong; Yan, Shuangyong; Chen, Xiwen; Wu, Zhenfeng; Chen, Rui; Luo, Xiangdong; Xie, Jiankun; Gao, Shan

    2016-04-26

    It is widely accepted that cultivated rice (Oryza sativa L.) was domesticated from common wild rice (Oryza rufipogon Griff.). Compared to other studies which concentrate on rice origin, this study is to genetically elucidate the substantially phenotypic and physiological changes from wild rice to cultivated rice at the whole genome level. Instead of comparing two assembled genomes, this study directly compared the Dongxiang wild rice (DXWR) Illumina sequencing reads with the Nipponbare (O. sativa) complete genome without assembly of the DXWR genome. Based on the results from the comparative genomics analysis, structural variations (SVs) between DXWR and Nipponbare were determined to locate deleted genes which could have been acquired by Nipponbare during rice domestication. To overcome the limit of the SV detection, the DXWR transcriptome was also sequenced and compared with the Nipponbare transcriptome to discover the genes which could have been lost in DXWR during domestication. Both 1591 Nipponbare-acquired genes and 206 DXWR-lost transcripts were further analyzed using annotations from multiple sources. The NGS data are available in the NCBI SRA database with ID SRP070627. These results help better understanding the domestication from wild rice to cultivated rice at the whole genome level and provide a genomic data resource for rice genetic research or breeding. One finding confirmed transposable elements contribute greatly to the genome evolution from wild rice to cultivated rice. Another finding suggested the photophosphorylation and oxidative phosphorylation system in cultivated rice could have adapted to environmental changes simultaneously during domestication.

  17. Asymmetric Modification of Hepatitis B Virus (HBV) Genomes by an Endogenous Cytidine Deaminase inside HBV Cores Informs a Model of Reverse Transcription.

    Science.gov (United States)

    Nair, Smita; Zlotnick, Adam

    2018-05-15

    Cytidine deaminases inhibit replication of a broad range of DNA viruses by deaminating cytidines on single-stranded DNA (ssDNA) to generate uracil. While several lines of evidence have revealed hepatitis B virus (HBV) genome editing by deamination, it is still unclear which nucleic acid intermediate of HBV is modified. Hepatitis B virus has a relaxed circular double-stranded DNA (rcDNA) genome that is reverse transcribed within virus cores from a RNA template. The HBV genome also persists as covalently closed circular DNA (cccDNA) in the nucleus of an infected cell. In the present study, we found that in HBV-producing HepAD38 and HepG2.2.15 cell lines, endogenous cytidine deaminases edited 10 to 25% of HBV rcDNA genomes, asymmetrically with almost all mutations on the 5' half of the minus strand. This region corresponds to the last half of the minus strand to be protected by plus-strand synthesis. Within this half of the genome, the number of mutations peaks in the middle. Overexpressed APOBEC3A and APOBEC3G could be packaged in HBV capsids but did not change the amount or distribution of mutations. We found no deamination on pregenomic RNA (pgRNA), indicating that an intact genome is encapsidated and deaminated during or after reverse transcription. The deamination pattern suggests a model of rcDNA synthesis in which pgRNA and then newly synthesized minus-sense single-stranded DNA are protected from deaminase by interaction with the virus capsid; during plus-strand synthesis, when enough dsDNA has been synthesized to displace the remaining minus strand from the capsid surface, the single-stranded DNA becomes deaminase sensitive. IMPORTANCE Host-induced mutation of the HBV genome by APOBEC proteins may be a path to clearing the virus. We examined cytidine-to-thymidine mutations in the genomes of HBV particles grown in the presence or absence of overexpressed APOBEC proteins. We found that genomes were subjected to deamination activity during reverse transcription

  18. Specificity and robustness in transcription control networks.

    Science.gov (United States)

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

    2002-02-19

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

  19. Diversity of Pseudomonas Genomes, Including Populus-Associated Isolates, as Revealed by Comparative Genome Analysis.

    Science.gov (United States)

    Jun, Se-Ran; Wassenaar, Trudy M; Nookaew, Intawat; Hauser, Loren; Wanchai, Visanu; Land, Miriam; Timm, Collin M; Lu, Tse-Yuan S; Schadt, Christopher W; Doktycz, Mitchel J; Pelletier, Dale A; Ussery, David W

    2016-01-01

    The Pseudomonas genus contains a metabolically versatile group of organisms that are known to occupy numerous ecological niches, including the rhizosphere and endosphere of many plants. Their diversity influences the phylogenetic diversity and heterogeneity of these communities. On the basis of average amino acid identity, comparative genome analysis of >1,000 Pseudomonas genomes, including 21 Pseudomonas strains isolated from the roots of native Populus deltoides (eastern cottonwood) trees resulted in consistent and robust genomic clusters with phylogenetic homogeneity. All Pseudomonas aeruginosa genomes clustered together, and these were clearly distinct from other Pseudomonas species groups on the basis of pangenome and core genome analyses. In contrast, the genomes of Pseudomonas fluorescens were organized into 20 distinct genomic clusters, representing enormous diversity and heterogeneity. Most of our 21 Populus-associated isolates formed three distinct subgroups within the major P. fluorescens group, supported by pathway profile analysis, while two isolates were more closely related to Pseudomonas chlororaphis and Pseudomonas putida. Genes specific to Populus-associated subgroups were identified. Genes specific to subgroup 1 include several sensory systems that act in two-component signal transduction, a TonB-dependent receptor, and a phosphorelay sensor. Genes specific to subgroup 2 contain hypothetical genes, and genes specific to subgroup 3 were annotated with hydrolase activit