Complexity and specificity of the maize (Zea mays L.) root hair transcriptome.

PubMed

Hey, Stefan; Baldauf, Jutta; Opitz, Nina; Lithio, Andrew; Pasha, Asher; Provart, Nicholas; Nettleton, Dan; Hochholdinger, Frank

2017-04-01

Root hairs are tubular extensions of epidermis cells. Transcriptome profiling demonstrated that the single cell-type root hair transcriptome was less complex than the transcriptome of multiple cell-type primary roots without root hairs. In total, 831 genes were exclusively and 5585 genes were preferentially expressed in root hairs [false discovery rate (FDR) ≤1%]. Among those, the most significantly enriched Gene Ontology (GO) functional terms were related to energy metabolism, highlighting the high energy demand for the development and function of root hairs. Subsequently, the maize homologs for 138 Arabidopsis genes known to be involved in root hair development were identified and their phylogenetic relationship and expression in root hairs were determined. This study indicated that the genetic regulation of root hair development in Arabidopsis and maize is controlled by common genes, but also shows differences which need to be dissected in future genetic experiments. Finally, a maize root view of the eFP browser was implemented including the root hair transcriptome of the present study and several previously published maize root transcriptome data sets. The eFP browser provides color-coded expression levels for these root types and tissues for any gene of interest, thus providing a novel resource to study gene expression and function in maize roots. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Transcriptome Profiling of Shewanella oneidensis Gene Expression following Exposure to Acidic and Alkaline pH†

PubMed Central

Leaphart, Adam B.; Thompson, Dorothea K.; Huang, Katherine; Alm, Eric; Wan, Xiu-Feng; Arkin, Adam; Brown, Steven D.; Wu, Liyou; Yan, Tingfen; Liu, Xueduan; Wickham, Gene S.; Zhou, Jizhong

2006-01-01

The molecular response of Shewanella oneidensis MR-1 to variations in extracellular pH was investigated based on genomewide gene expression profiling. Microarray analysis revealed that cells elicited both general and specific transcriptome responses when challenged with environmental acid (pH 4) or base (pH 10) conditions over a 60-min period. Global responses included the differential expression of genes functionally linked to amino acid metabolism, transcriptional regulation and signal transduction, transport, cell membrane structure, and oxidative stress protection. Response to acid stress included the elevated expression of genes encoding glycogen biosynthetic enzymes, phosphate transporters, and the RNA polymerase sigma-38 factor (rpoS), whereas the molecular response to alkaline pH was characterized by upregulation of nhaA and nhaR, which are predicted to encode an Na+/H+ antiporter and transcriptional activator, respectively, as well as sulfate transport and sulfur metabolism genes. Collectively, these results suggest that S. oneidensis modulates multiple transporters, cell envelope components, and pathways of amino acid consumption and central intermediary metabolism as part of its transcriptome response to changing external pH conditions. PMID:16452448

Integrated analysis of whole-exome sequencing and transcriptome profiling in males with autism spectrum disorders.

PubMed

Codina-Solà, Marta; Rodríguez-Santiago, Benjamín; Homs, Aïda; Santoyo, Javier; Rigau, Maria; Aznar-Laín, Gemma; Del Campo, Miguel; Gener, Blanca; Gabau, Elisabeth; Botella, María Pilar; Gutiérrez-Arumí, Armand; Antiñolo, Guillermo; Pérez-Jurado, Luis Alberto; Cuscó, Ivon

2015-01-01

Autism spectrum disorders (ASD) are a group of neurodevelopmental disorders with high heritability. Recent findings support a highly heterogeneous and complex genetic etiology including rare de novo and inherited mutations or chromosomal rearrangements as well as double or multiple hits. We performed whole-exome sequencing (WES) and blood cell transcriptome by RNAseq in a subset of male patients with idiopathic ASD (n = 36) in order to identify causative genes, transcriptomic alterations, and susceptibility variants. We detected likely monogenic causes in seven cases: five de novo (SCN2A, MED13L, KCNV1, CUL3, and PTEN) and two inherited X-linked variants (MAOA and CDKL5). Transcriptomic analyses allowed the identification of intronic causative mutations missed by the usual filtering of WES and revealed functional consequences of some rare mutations. These included aberrant transcripts (PTEN, POLR3C), deregulated expression in 1.7% of mutated genes (that is, SEMA6B, MECP2, ANK3, CREBBP), allele-specific expression (FUS, MTOR, TAF1C), and non-sense-mediated decay (RIT1, ALG9). The analysis of rare inherited variants showed enrichment in relevant pathways such as the PI3K-Akt signaling and the axon guidance. Integrative analysis of WES and blood RNAseq data has proven to be an efficient strategy to identify likely monogenic forms of ASD (19% in our cohort), as well as additional rare inherited mutations that can contribute to ASD risk in a multifactorial manner. Blood transcriptomic data, besides validating 88% of expressed variants, allowed the identification of missed intronic mutations and revealed functional correlations of genetic variants, including changes in splicing, expression levels, and allelic expression.

Transcriptome profiles of chicken intestinal intraepithelial lymphocytes altered by the intake of a multi-strain direct-fed microbials

USDA-ARS?s Scientific Manuscript database

The current study was conducted to investigate the effects of the direct-fed microbials (DFM) including three Bacillus subtilis strains on the modulation of transcriptional profile in chicken intestinal intraepithelial lymphocytes (IEL). The multiple-strain DFM product modified 453 probes from 1,98...

Transcriptome analysis of Pinus monticola primary needles by RNA-seq provides novel insight into host resistance to Cronartium ribicola

PubMed Central

2013-01-01

Background Five-needle pines are important forest species that have been devastated by white pine blister rust (WPBR, caused by Cronartium ribicola) across North America. Currently little transcriptomic and genomic data are available to understand molecular interactions in the WPBR pathosystem. Results We report here RNA-seq analysis results using Illumina deep sequencing of primary needles of western white pine (Pinus monticola) infected with WPBR. De novo gene assembly was used to generate the first P. monticola consensus transcriptome, which contained 39,439 unique transcripts with an average length of 1,303 bp and a total length of 51.4 Mb. About 23,000 P. monticola unigenes produced orthologous hits in the Pinus gene index (PGI) database (BLASTn with E values < e-100) and 6,300 genes were expressed actively (at RPKM ≥ 10) in the healthy tissues. Comparison of transcriptomes from WPBR-susceptible and -resistant genotypes revealed a total of 979 differentially expressed genes (DEGs) with a significant fold change > 1.5 during P. monticola- C. ribicola interactions. Three hundred and ten DEGs were regulated similarly in both susceptible and resistant seedlings and 275 DEGs showed regulatory differences between susceptible and resistant seedlings post infection by C. ribicola. The DEGs up-regulated in resistant seedlings included a set of putative signal receptor genes encoding disease resistance protein homologs, calcineurin B-like (CBL)-interacting protein kinases (CIPK), F-box family proteins (FBP), and abscisic acid (ABA) receptor; transcriptional factor (TF) genes of multiple families; genes homologous to apoptosis-inducing factor (AIF), flowering locus T-like protein (FT), and subtilisin-like protease. DEGs up-regulated in resistant seedlings also included a wide diversity of down-stream genes (encoding enzymes involved in different metabolic pathways, pathogenesis-related -PR proteins of multiple families, and anti-microbial proteins). A large proportion of the down-regulated DEGs were related to photosystems, the metabolic pathways of carbon fixation and flavonoid biosynthesis. Conclusions The novel P. monticola transcriptome data provide a basis for future studies of genetic resistance in a non-model, coniferous species. Our global gene expression profiling presents a comprehensive view of transcriptomic regulation in the WPBR pathosystem and yields novel insights on molecular and biochemical mechanisms of disease resistance in conifers. PMID:24341615

Mitochondria, oligodendrocytes and inflammation in bipolar disorder: evidence from transcriptome studies points to intriguing parallels with multiple sclerosis

PubMed Central

Konradi, Christine; Sillivan, Stephanie E.; Clay, Hayley B.

2011-01-01

Gene expression studies of bipolar disorder (BPD) have shown changes in transcriptome profiles in multiple brain regions. Here we summarize the most consistent findings in the scientific literature, and compare them to data from schizophrenia (SZ) and major depressive disorder (MDD). The transcriptome profiles of all three disorders overlap, making the existence of a BPD-specific profile unlikely. Three groups of functionally related genes are consistently expressed at altered levels in BPD, SZ and MDD. Genes involved in energy metabolism and mitochondrial function are downregulated, genes involved in immune response and inflammation are upregulated, and genes expressed in oligodendrocytes are downregulated. Experimental paradigms for multiple sclerosis demonstrate a tight link between energy metabolism, inflammation and demyelination. These studies also show variabilities in the extent of oligodendrocyte stress, which can vary from a downregulation of oligodendrocyte genes, such as observed in psychiatric disorders, to cell death and brain lesions seen in multiple sclerosis. We conclude that experimental models of multiple sclerosis could be of interest for the research of BPD, SZ and MDD. PMID:21310238

Pyrosequencing the Midgut Transcriptome of the Banana Weevil Cosmopolites sordidus (Germar) (Coleoptera: Curculionidae) Reveals Multiple Protease-Like Transcripts.

PubMed

Valencia, Arnubio; Wang, Haichuan; Soto, Alberto; Aristizabal, Manuel; Arboleda, Jorge W; Eyun, Seong-Il; Noriega, Daniel D; Siegfried, Blair

2016-01-01

The banana weevil Cosmopolites sordidus is an important and serious insect pest in most banana and plantain-growing areas of the world. In spite of the economic importance of this insect pest very little genomic and transcriptomic information exists for this species. In the present study, we characterized the midgut transcriptome of C. sordidus using massive 454-pyrosequencing. We generated over 590,000 sequencing reads that assembled into 30,840 contigs with more than 400 bp, representing a significant expansion of existing sequences available for this insect pest. Among them, 16,427 contigs contained one or more GO terms. In addition, 15,263 contigs were assigned an EC number. In-depth transcriptome analysis identified genes potentially involved in insecticide resistance, peritrophic membrane biosynthesis, immunity-related function and defense against pathogens, and Bacillus thuringiensis toxins binding proteins as well as multiple enzymes involved with protein digestion. This transcriptome will provide a valuable resource for understanding larval physiology and for identifying novel target sites and management approaches for this important insect pest.

Pyrosequencing the Midgut Transcriptome of the Banana Weevil Cosmopolites sordidus (Germar) (Coleoptera: Curculionidae) Reveals Multiple Protease-Like Transcripts

PubMed Central

Valencia, Arnubio; Wang, Haichuan; Soto, Alberto; Aristizabal, Manuel; Arboleda, Jorge W.; Eyun, Seong-il; Noriega, Daniel D.; Siegfried, Blair

2016-01-01

The banana weevil Cosmopolites sordidus is an important and serious insect pest in most banana and plantain-growing areas of the world. In spite of the economic importance of this insect pest very little genomic and transcriptomic information exists for this species. In the present study, we characterized the midgut transcriptome of C. sordidus using massive 454-pyrosequencing. We generated over 590,000 sequencing reads that assembled into 30,840 contigs with more than 400 bp, representing a significant expansion of existing sequences available for this insect pest. Among them, 16,427 contigs contained one or more GO terms. In addition, 15,263 contigs were assigned an EC number. In-depth transcriptome analysis identified genes potentially involved in insecticide resistance, peritrophic membrane biosynthesis, immunity-related function and defense against pathogens, and Bacillus thuringiensis toxins binding proteins as well as multiple enzymes involved with protein digestion. This transcriptome will provide a valuable resource for understanding larval physiology and for identifying novel target sites and management approaches for this important insect pest. PMID:26949943

Integrative FourD omics approach profiles the target network of the carbon storage regulatory system

PubMed Central

Sowa, Steven W.; Gelderman, Grant; Leistra, Abigail N.; Buvanendiran, Aishwarya; Lipp, Sarah; Pitaktong, Areen; Vakulskas, Christopher A.; Romeo, Tony; Baldea, Michael

2017-01-01

Abstract Multi-target regulators represent a largely untapped area for metabolic engineering and anti-bacterial development. These regulators are complex to characterize because they often act at multiple levels, affecting proteins, transcripts and metabolites. Therefore, single omics experiments cannot profile their underlying targets and mechanisms. In this work, we used an Integrative FourD omics approach (INFO) that consists of collecting and analyzing systems data throughout multiple time points, using multiple genetic backgrounds, and multiple omics approaches (transcriptomics, proteomics and high throughput sequencing crosslinking immunoprecipitation) to evaluate simultaneous changes in gene expression after imposing an environmental stress that accentuates the regulatory features of a network. Using this approach, we profiled the targets and potential regulatory mechanisms of a global regulatory system, the well-studied carbon storage regulatory (Csr) system of Escherichia coli, which is widespread among bacteria. Using 126 sets of proteomics and transcriptomics data, we identified 136 potential direct CsrA targets, including 50 novel ones, categorized their behaviors into distinct regulatory patterns, and performed in vivo fluorescence-based follow up experiments. The results of this work validate 17 novel mRNAs as authentic direct CsrA targets and demonstrate a generalizable strategy to integrate multiple lines of omics data to identify a core pool of regulator targets. PMID:28126921

The Need for Integrated Approaches in Metabolic Engineering

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

Lechner, Anna; Brunk, Elizabeth; Keasling, Jay D.

Highlights include state-of-the-art procedures for heterologous small-molecule biosynthesis, the associated bottlenecks, and new strategies that have the potential to accelerate future accomplishments in metabolic engineering. A combination of different approaches over multiple time and size scales must be considered for successful pathway engineering in a heterologous host. We have classified these optimization procedures based on the “system” that is being manipulated: transcriptome, translatome, proteome, or reactome. Here, by bridging multiple disciplines, including molecular biology, biochemistry, biophysics, and computational sciences, we can create an integral framework for the discovery and implementation of novel biosynthetic production routes.

The Need for Integrated Approaches in Metabolic Engineering

DOE PAGES

Lechner, Anna; Brunk, Elizabeth; Keasling, Jay D.

2016-08-15

Highlights include state-of-the-art procedures for heterologous small-molecule biosynthesis, the associated bottlenecks, and new strategies that have the potential to accelerate future accomplishments in metabolic engineering. A combination of different approaches over multiple time and size scales must be considered for successful pathway engineering in a heterologous host. We have classified these optimization procedures based on the “system” that is being manipulated: transcriptome, translatome, proteome, or reactome. Here, by bridging multiple disciplines, including molecular biology, biochemistry, biophysics, and computational sciences, we can create an integral framework for the discovery and implementation of novel biosynthetic production routes.

De novo transcriptome assembly of the calanoid copepod Neocalanus flemingeri: A new resource for emergence from diapause.

PubMed

Roncalli, Vittoria; Cieslak, Matthew C; Sommer, Stephanie A; Hopcroft, Russell R; Lenz, Petra H

2018-02-01

Copepods, small planktonic crustaceans, are key links between primary producers and upper trophic levels, including many economically important fishes. In the subarctic North Pacific, the life cycle of copepods like Neocalanus flemingeri includes an ontogenetic migration to depth followed by a period of diapause (a type of dormancy) characterized by arrested development and low metabolic activity. The end of diapause is marked by the production of the first brood of eggs. Recent temperature anomalies in the North Pacific have raised concerns about potential negative effects on N. flemingeri. Since diapause is a developmental program, its progress can be tracked using through global gene expression. Thus, a reference transcriptome was developed as a first step towards physiological profiling of diapausing females using high-throughput Illumina sequencing. The de novo transcriptome, the first for this species was designed to investigate the diapause period. RNA-Seq reads were obtained for dormant to reproductive N. flemingeri females. A high quality de novo transcriptome was obtained by first assembling reads from each individual using Trinity software followed by clustering with CAP3 Assembly Program. This assembly consisted of 140,841transcripts (contigs). Bench-marking universal single-copy orthologs analysis identified 85% of core eukaryotic genes, with 79% predicted to be complete. Comparison with other calanoid transcriptomes confirmed its quality and degree of completeness. Trinity assembly of reads originating from multiple individuals led to fragmentation. Thus, the workflow applied here differed from the one recommended by Trinity, but was required to obtain a good assembly. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Determining the optimal number of independent components for reproducible transcriptomic data analysis.

PubMed

Kairov, Ulykbek; Cantini, Laura; Greco, Alessandro; Molkenov, Askhat; Czerwinska, Urszula; Barillot, Emmanuel; Zinovyev, Andrei

2017-09-11

Independent Component Analysis (ICA) is a method that models gene expression data as an action of a set of statistically independent hidden factors. The output of ICA depends on a fundamental parameter: the number of components (factors) to compute. The optimal choice of this parameter, related to determining the effective data dimension, remains an open question in the application of blind source separation techniques to transcriptomic data. Here we address the question of optimizing the number of statistically independent components in the analysis of transcriptomic data for reproducibility of the components in multiple runs of ICA (within the same or within varying effective dimensions) and in multiple independent datasets. To this end, we introduce ranking of independent components based on their stability in multiple ICA computation runs and define a distinguished number of components (Most Stable Transcriptome Dimension, MSTD) corresponding to the point of the qualitative change of the stability profile. Based on a large body of data, we demonstrate that a sufficient number of dimensions is required for biological interpretability of the ICA decomposition and that the most stable components with ranks below MSTD have more chances to be reproduced in independent studies compared to the less stable ones. At the same time, we show that a transcriptomics dataset can be reduced to a relatively high number of dimensions without losing the interpretability of ICA, even though higher dimensions give rise to components driven by small gene sets. We suggest a protocol of ICA application to transcriptomics data with a possibility of prioritizing components with respect to their reproducibility that strengthens the biological interpretation. Computing too few components (much less than MSTD) is not optimal for interpretability of the results. The components ranked within MSTD range have more chances to be reproduced in independent studies.

Cited1 Deficiency Suppresses Intestinal Tumorigenesis

PubMed Central

Young, Madeleine; Poetz, Oliver; Parry, Lee; Jenkins, John R.; Williams, Geraint T.; Dunwoodie, Sally L.; Watson, Alastair; Clarke, Alan R.

2013-01-01

Conditional deletion of Apc in the murine intestine alters crypt-villus architecture and function. This process is accompanied by multiple changes in gene expression, including upregulation of Cited1, whose role in colorectal carcinogenesis is unknown. Here we explore the relevance of Cited1 to intestinal tumorigenesis. We crossed Cited1 null mice with ApcMin/+ and AhCre+Apcfl/fl mice and determined the impact of Cited1 deficiency on tumour growth/initiation including tumour multiplicity, cell proliferation, apoptosis and the transcriptome. We show that Cited1 is up-regulated in both human and murine tumours, and that constitutive deficiency of Cited1 increases survival in ApcMin/+ mice from 230.5 to 515 days. However, paradoxically, Cited1 deficiency accentuated nearly all aspects of the immediate phenotype 4 days after conditional deletion of Apc, including an increase in cell death and enhanced perturbation of differentiation, including of the stem cell compartment. Transcriptome analysis revealed multiple pathway changes, including p53, PI3K and Wnt. The activation of Wnt through Cited1 deficiency correlated with increased transcription of β-catenin and increased levels of dephosphorylated β-catenin. Hence, immediately following deletion of Apc, Cited1 normally restrains the Wnt pathway at the level of β-catenin. Thus deficiency of Cited1 leads to hyper-activation of Wnt signaling and an exaggerated Wnt phenotype including elevated cell death. Cited1 deficiency decreases intestinal tumourigenesis in ApcMin/+ mice and impacts upon a number of oncogenic signaling pathways, including Wnt. This restraint imposed by Cited1 is consistent with a requirement for Cited1 to constrain Wnt activity to a level commensurate with optimal adenoma formation and maintenance, and provides one mechanism for tumour repression in the absence of Cited1. PMID:23935526

The head-regeneration transcriptome of the planarian Schmidtea mediterranea.

PubMed

Sandmann, Thomas; Vogg, Matthias C; Owlarn, Suthira; Boutros, Michael; Bartscherer, Kerstin

2011-08-16

Planarian flatworms can regenerate their head, including a functional brain, within less than a week. Despite the enormous potential of these animals for medical research and regenerative medicine, the mechanisms of regeneration and the molecules involved remain largely unknown. To identify genes that are differentially expressed during early stages of planarian head regeneration, we generated a de novo transcriptome assembly from more than 300 million paired-end reads from planarian fragments regenerating the head at 16 different time points. The assembly yielded 26,018 putative transcripts, including very long transcripts spanning multiple genomic supercontigs, and thousands of isoforms. Using short-read data from two platforms, we analyzed dynamic gene regulation during the first three days of head regeneration. We identified at least five different temporal synexpression classes, including genes specifically induced within a few hours after injury. Furthermore, we characterized the role of a conserved Runx transcription factor, smed-runt-like1. RNA interference (RNAi) knockdown and immunofluorescence analysis of the regenerating visual system indicated that smed-runt-like1 encodes a transcriptional regulator of eye morphology and photoreceptor patterning. Transcriptome sequencing of short reads allowed for the simultaneous de novo assembly and differential expression analysis of transcripts, demonstrating highly dynamic regulation during head regeneration in planarians.

The head-regeneration transcriptome of the planarian Schmidtea mediterranea

PubMed Central

2011-01-01

Background Planarian flatworms can regenerate their head, including a functional brain, within less than a week. Despite the enormous potential of these animals for medical research and regenerative medicine, the mechanisms of regeneration and the molecules involved remain largely unknown. Results To identify genes that are differentially expressed during early stages of planarian head regeneration, we generated a de novo transcriptome assembly from more than 300 million paired-end reads from planarian fragments regenerating the head at 16 different time points. The assembly yielded 26,018 putative transcripts, including very long transcripts spanning multiple genomic supercontigs, and thousands of isoforms. Using short-read data from two platforms, we analyzed dynamic gene regulation during the first three days of head regeneration. We identified at least five different temporal synexpression classes, including genes specifically induced within a few hours after injury. Furthermore, we characterized the role of a conserved Runx transcription factor, smed-runt-like1. RNA interference (RNAi) knockdown and immunofluorescence analysis of the regenerating visual system indicated that smed-runt-like1 encodes a transcriptional regulator of eye morphology and photoreceptor patterning. Conclusions Transcriptome sequencing of short reads allowed for the simultaneous de novo assembly and differential expression analysis of transcripts, demonstrating highly dynamic regulation during head regeneration in planarians. PMID:21846378

Survival of Halophilic Archaea in the Stratosphere as a Mars Analog: A Transcriptomic Approach

NASA Astrophysics Data System (ADS)

DasSarma, S.; DasSarma, P.; Laye, V.; Harvey, J.; Reid, C.; Shultz, J.; Yarborough, A.; Lamb, A.; Koske-Phillips, A.; Herbst, A.; Molina, F.; Grah, O.; Phillips, T.

2016-05-01

On Earth, halophilic Archaea tolerate multiple extreme conditions similar to those on Mars. In order to study their survival, we launched live cultures into Earth’s stratosphere on helium balloons. The effects on survival and transcriptomes were interrogated in the lab.

Integrative FourD omics approach profiles the target network of the carbon storage regulatory system.

PubMed

Sowa, Steven W; Gelderman, Grant; Leistra, Abigail N; Buvanendiran, Aishwarya; Lipp, Sarah; Pitaktong, Areen; Vakulskas, Christopher A; Romeo, Tony; Baldea, Michael; Contreras, Lydia M

2017-02-28

Multi-target regulators represent a largely untapped area for metabolic engineering and anti-bacterial development. These regulators are complex to characterize because they often act at multiple levels, affecting proteins, transcripts and metabolites. Therefore, single omics experiments cannot profile their underlying targets and mechanisms. In this work, we used an Integrative FourD omics approach (INFO) that consists of collecting and analyzing systems data throughout multiple time points, using multiple genetic backgrounds, and multiple omics approaches (transcriptomics, proteomics and high throughput sequencing crosslinking immunoprecipitation) to evaluate simultaneous changes in gene expression after imposing an environmental stress that accentuates the regulatory features of a network. Using this approach, we profiled the targets and potential regulatory mechanisms of a global regulatory system, the well-studied carbon storage regulatory (Csr) system of Escherichia coli, which is widespread among bacteria. Using 126 sets of proteomics and transcriptomics data, we identified 136 potential direct CsrA targets, including 50 novel ones, categorized their behaviors into distinct regulatory patterns, and performed in vivo fluorescence-based follow up experiments. The results of this work validate 17 novel mRNAs as authentic direct CsrA targets and demonstrate a generalizable strategy to integrate multiple lines of omics data to identify a core pool of regulator targets. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

PIVOT: platform for interactive analysis and visualization of transcriptomics data.

PubMed

Zhu, Qin; Fisher, Stephen A; Dueck, Hannah; Middleton, Sarah; Khaladkar, Mugdha; Kim, Junhyong

2018-01-05

Many R packages have been developed for transcriptome analysis but their use often requires familiarity with R and integrating results of different packages requires scripts to wrangle the datatypes. Furthermore, exploratory data analyses often generate multiple derived datasets such as data subsets or data transformations, which can be difficult to track. Here we present PIVOT, an R-based platform that wraps open source transcriptome analysis packages with a uniform user interface and graphical data management that allows non-programmers to interactively explore transcriptomics data. PIVOT supports more than 40 popular open source packages for transcriptome analysis and provides an extensive set of tools for statistical data manipulations. A graph-based visual interface is used to represent the links between derived datasets, allowing easy tracking of data versions. PIVOT further supports automatic report generation, publication-quality plots, and program/data state saving, such that all analysis can be saved, shared and reproduced. PIVOT will allow researchers with broad background to easily access sophisticated transcriptome analysis tools and interactively explore transcriptome datasets.

Reptilian-transcriptome v1.0, a glimpse in the brain transcriptome of five divergent Sauropsida lineages and the phylogenetic position of turtles.

PubMed

Tzika, Athanasia C; Helaers, Raphaël; Schramm, Gerrit; Milinkovitch, Michel C

2011-09-26

Reptiles are largely under-represented in comparative genomics despite the fact that they are substantially more diverse in many respects than mammals. Given the high divergence of reptiles from classical model species, next-generation sequencing of their transcriptomes is an approach of choice for gene identification and annotation. Here, we use 454 technology to sequence the brain transcriptome of four divergent reptilian and one reference avian species: the Nile crocodile, the corn snake, the bearded dragon, the red-eared turtle, and the chicken. Using an in-house pipeline for recursive similarity searches of >3,000,000 reads against multiple databases from 7 reference vertebrates, we compile a reptilian comparative transcriptomics dataset, with homology assignment for 20,000 to 31,000 transcripts per species and a cumulated non-redundant sequence length of 248.6 Mbases. Our approach identifies the majority (87%) of chicken brain transcripts and about 50% of de novo assembled reptilian transcripts. In addition to 57,502 microsatellite loci, we identify thousands of SNP and indel polymorphisms for population genetic and linkage analyses. We also build very large multiple alignments for Sauropsida and mammals (two million residues per species) and perform extensive phylogenetic analyses suggesting that turtles are not basal living reptiles but are rather associated with Archosaurians, hence, potentially answering a long-standing question in the phylogeny of Amniotes. The reptilian transcriptome (freely available at http://www.reptilian-transcriptomes.org) should prove a useful new resource as reptiles are becoming important new models for comparative genomics, ecology, and evolutionary developmental genetics.

Comparative analysis of transcriptome in two wheat genotypes with contrasting levels of drought tolerance

USDA-ARS?s Scientific Manuscript database

Drought tolerance is a complex trait that is governed by multiple genes. To identify the potential candidate genes, comparative analysis of drought stress-responsive transcriptome between drought-tolerant (Triticum aestivum Cv. C306) and drought-sensitive (Triticum aestivum Cv. WL711) genotypes was ...

Correction to: Comparison of multiple transcriptomes exposes unified and divergent features of quiescent and activated skeletal muscle stem cells.

PubMed

Pietrosemoli, Natalia; Mella, Sébastien; Yennek, Siham; Baghdadi, Meryem B; Sakai, Hiroshi; Sambasivan, Ramkumar; Pala, Francesca; Di Girolamo, Daniela; Tajbakhsh, Shahragim

2018-06-06

After publication of this article [1], the authors noted that the legends for supplementary files Figures S3 and S4 were truncated in the production process, therefore lacking some information concerning these Figures. The complete legends are included in this Correction. The authors apologize for any inconvenience that this might have caused.

De Novo transcriptome assembly (NGS) of Curcuma longa L. rhizome reveals novel transcripts related to anticancer and antimalarial terpenoids.

PubMed

Annadurai, Ramasamy S; Neethiraj, Ramprasad; Jayakumar, Vasanthan; Damodaran, Anand C; Rao, Sudha Narayana; Katta, Mohan A V S K; Gopinathan, Sreeja; Sarma, Santosh Prasad; Senthilkumar, Vanitha; Niranjan, Vidya; Gopinath, Ashok; Mugasimangalam, Raja C

2013-01-01

Herbal remedies are increasingly being recognised in recent years as alternative medicine for a number of diseases including cancer. Curcuma longa L., commonly known as turmeric is used as a culinary spice in India and in many Asian countries has been attributed to lower incidences of gastrointestinal cancers. Curcumin, a secondary metabolite isolated from the rhizomes of this plant has been shown to have significant anticancer properties, in addition to antimalarial and antioxidant effects. We sequenced the transcriptome of the rhizome of the 3 varieties of Curcuma longa L. using Illumina reversible dye terminator sequencing followed by de novo transcriptome assembly. Multiple databases were used to obtain a comprehensive annotation and the transcripts were functionally classified using GO, KOG and PlantCyc. Special emphasis was given for annotating the secondary metabolite pathways and terpenoid biosynthesis pathways. We report for the first time, the presence of transcripts related to biosynthetic pathways of several anti-cancer compounds like taxol, curcumin, and vinblastine in addition to anti-malarial compounds like artemisinin and acridone alkaloids, emphasizing turmeric's importance as a highly potent phytochemical. Our data not only provides molecular signatures for several terpenoids but also a comprehensive molecular resource for facilitating deeper insights into the transcriptome of C. longa.

De Novo Transcriptome Assembly (NGS) of Curcuma longa L. Rhizome Reveals Novel Transcripts Related to Anticancer and Antimalarial Terpenoids

PubMed Central

Jayakumar, Vasanthan; Damodaran, Anand C.; Rao, Sudha Narayana; Katta, Mohan A. V. S. K.; Gopinathan, Sreeja; Sarma, Santosh Prasad; Senthilkumar, Vanitha; Niranjan, Vidya; Gopinath, Ashok; Mugasimangalam, Raja C.

2013-01-01

Herbal remedies are increasingly being recognised in recent years as alternative medicine for a number of diseases including cancer. Curcuma longa L., commonly known as turmeric is used as a culinary spice in India and in many Asian countries has been attributed to lower incidences of gastrointestinal cancers. Curcumin, a secondary metabolite isolated from the rhizomes of this plant has been shown to have significant anticancer properties, in addition to antimalarial and antioxidant effects. We sequenced the transcriptome of the rhizome of the 3 varieties of Curcuma longa L. using Illumina reversible dye terminator sequencing followed by de novo transcriptome assembly. Multiple databases were used to obtain a comprehensive annotation and the transcripts were functionally classified using GO, KOG and PlantCyc. Special emphasis was given for annotating the secondary metabolite pathways and terpenoid biosynthesis pathways. We report for the first time, the presence of transcripts related to biosynthetic pathways of several anti-cancer compounds like taxol, curcumin, and vinblastine in addition to anti-malarial compounds like artemisinin and acridone alkaloids, emphasizing turmeric's importance as a highly potent phytochemical. Our data not only provides molecular signatures for several terpenoids but also a comprehensive molecular resource for facilitating deeper insights into the transcriptome of C. longa. PMID:23468859

Transcriptome landscape of a bacterial pathogen under plant immunity.

PubMed

Nobori, Tatsuya; Velásquez, André C; Wu, Jingni; Kvitko, Brian H; Kremer, James M; Wang, Yiming; He, Sheng Yang; Tsuda, Kenichi

2018-03-27

Plant pathogens can cause serious diseases that impact global agriculture. The plant innate immunity, when fully activated, can halt pathogen growth in plants. Despite extensive studies into the molecular and genetic bases of plant immunity against pathogens, the influence of plant immunity in global pathogen metabolism to restrict pathogen growth is poorly understood. Here, we developed RNA sequencing pipelines for analyzing bacterial transcriptomes in planta and determined high-resolution transcriptome patterns of the foliar bacterial pathogen Pseudomonas syringae in Arabidopsis thaliana with a total of 27 combinations of plant immunity mutants and bacterial strains. Bacterial transcriptomes were analyzed at 6 h post infection to capture early effects of plant immunity on bacterial processes and to avoid secondary effects caused by different bacterial population densities in planta We identified specific "immune-responsive" bacterial genes and processes, including those that are activated in susceptible plants and suppressed by plant immune activation. Expression patterns of immune-responsive bacterial genes at the early time point were tightly linked to later bacterial growth levels in different host genotypes. Moreover, we found that a bacterial iron acquisition pathway is commonly suppressed by multiple plant immune-signaling pathways. Overexpression of a P. syringae sigma factor gene involved in iron regulation and other processes partially countered bacterial growth restriction during the plant immune response triggered by AvrRpt2. Collectively, this study defines the effects of plant immunity on the transcriptome of a bacterial pathogen and sheds light on the enigmatic mechanisms of bacterial growth inhibition during the plant immune response.

Allele Identification for Transcriptome-Based Population Genomics in the Invasive Plant Centaurea solstitialis

PubMed Central

Dlugosch, Katrina M.; Lai, Zhao; Bonin, Aurélie; Hierro, José; Rieseberg, Loren H.

2013-01-01

Transcriptome sequences are becoming more broadly available for multiple individuals of the same species, providing opportunities to derive population genomic information from these datasets. Using the 454 Life Science Genome Sequencer FLX and FLX-Titanium next-generation platforms, we generated 11−430 Mbp of sequence for normalized cDNA for 40 wild genotypes of the invasive plant Centaurea solstitialis, yellow starthistle, from across its worldwide distribution. We examined the impact of sequencing effort on transcriptome recovery and overlap among individuals. To do this, we developed two novel publicly available software pipelines: SnoWhite for read cleaning before assembly, and AllelePipe for clustering of loci and allele identification in assembled datasets with or without a reference genome. AllelePipe is designed specifically for cases in which read depth information is not appropriate or available to assist with disentangling closely related paralogs from allelic variation, as in transcriptome or previously assembled libraries. We find that modest applications of sequencing effort recover most of the novel sequences present in the transcriptome of this species, including single-copy loci and a representative distribution of functional groups. In contrast, the coverage of variable sites, observation of heterozygosity, and overlap among different libraries are all highly dependent on sequencing effort. Nevertheless, the information gained from overlapping regions was informative regarding coarse population structure and variation across our small number of population samples, providing the first genetic evidence in support of hypothesized invasion scenarios. PMID:23390612

Reptilian-transcriptome v1.0, a glimpse in the brain transcriptome of five divergent Sauropsida lineages and the phylogenetic position of turtles

PubMed Central

2011-01-01

Background Reptiles are largely under-represented in comparative genomics despite the fact that they are substantially more diverse in many respects than mammals. Given the high divergence of reptiles from classical model species, next-generation sequencing of their transcriptomes is an approach of choice for gene identification and annotation. Results Here, we use 454 technology to sequence the brain transcriptome of four divergent reptilian and one reference avian species: the Nile crocodile, the corn snake, the bearded dragon, the red-eared turtle, and the chicken. Using an in-house pipeline for recursive similarity searches of >3,000,000 reads against multiple databases from 7 reference vertebrates, we compile a reptilian comparative transcriptomics dataset, with homology assignment for 20,000 to 31,000 transcripts per species and a cumulated non-redundant sequence length of 248.6 Mbases. Our approach identifies the majority (87%) of chicken brain transcripts and about 50% of de novo assembled reptilian transcripts. In addition to 57,502 microsatellite loci, we identify thousands of SNP and indel polymorphisms for population genetic and linkage analyses. We also build very large multiple alignments for Sauropsida and mammals (two million residues per species) and perform extensive phylogenetic analyses suggesting that turtles are not basal living reptiles but are rather associated with Archosaurians, hence, potentially answering a long-standing question in the phylogeny of Amniotes. Conclusions The reptilian transcriptome (freely available at http://www.reptilian-transcriptomes.org) should prove a useful new resource as reptiles are becoming important new models for comparative genomics, ecology, and evolutionary developmental genetics. PMID:21943375

Global transcriptomic analysis suggests carbon dioxide as an environmental stressor in spaceflight: A systems biology GeneLab case study.

PubMed

Beheshti, Afshin; Cekanaviciute, Egle; Smith, David J; Costes, Sylvain V

2018-03-08

Spaceflight introduces a combination of environmental stressors, including microgravity, ionizing radiation, changes in diet and altered atmospheric gas composition. In order to understand the impact of each environmental component on astronauts it is important to investigate potential influences in isolation. Rodent spaceflight experiments involve both standard vivarium cages and animal enclosure modules (AEMs), which are cages used to house rodents in spaceflight. Ground control AEMs are engineered to match the spaceflight environment. There are limited studies examining the biological response invariably due to the configuration of AEM and vivarium housing. To investigate the innate global transcriptomic patterns of rodents housed in spaceflight-matched AEM compared to standard vivarium cages we utilized publicly available data from the NASA GeneLab repository. Using a systems biology approach, we observed that AEM housing was associated with significant transcriptomic differences, including reduced metabolism, altered immune responses, and activation of possible tumorigenic pathways. Although we did not perform any functional studies, our findings revealed a mild hypoxic phenotype in AEM, possibly due to atmospheric carbon dioxide that was increased to match conditions in spaceflight. Our investigation illustrates the process of generating new hypotheses and informing future experimental research by repurposing multiple space-flown datasets.

Fungal proteomics: from identification to function.

PubMed

Doyle, Sean

2011-08-01

Some fungi cause disease in humans and plants, while others have demonstrable potential for the control of insect pests. In addition, fungi are also a rich reservoir of therapeutic metabolites and industrially useful enzymes. Detailed analysis of fungal biochemistry is now enabled by multiple technologies including protein mass spectrometry, genome and transcriptome sequencing and advances in bioinformatics. Yet, the assignment of function to fungal proteins, encoded either by in silico annotated, or unannotated genes, remains problematic. The purpose of this review is to describe the strategies used by many researchers to reveal protein function in fungi, and more importantly, to consolidate the nomenclature of 'unknown function protein' as opposed to 'hypothetical protein' - once any protein has been identified by protein mass spectrometry. A combination of approaches including comparative proteomics, pathogen-induced protein expression and immunoproteomics are outlined, which, when used in combination with a variety of other techniques (e.g. functional genomics, microarray analysis, immunochemical and infection model systems), appear to yield comprehensive and definitive information on protein function in fungi. The relative advantages of proteomic, as opposed to transcriptomic-only, analyses are also described. In the future, combined high-throughput, quantitative proteomics, allied to transcriptomic sequencing, are set to reveal much about protein function in fungi. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

Transcriptomic analysis of grain amaranth (Amaranthus hypochondriacus) using 454 pyrosequencing: comparison with A. tuberculatus, expression profiling in stems and in response to biotic and abiotic stress

PubMed Central

2011-01-01

Background Amaranthus hypochondriacus, a grain amaranth, is a C4 plant noted by its ability to tolerate stressful conditions and produce highly nutritious seeds. These possess an optimal amino acid balance and constitute a rich source of health-promoting peptides. Although several recent studies, mostly involving subtractive hybridization strategies, have contributed to increase the relatively low number of grain amaranth expressed sequence tags (ESTs), transcriptomic information of this species remains limited, particularly regarding tissue-specific and biotic stress-related genes. Thus, a large scale transcriptome analysis was performed to generate stem- and (a)biotic stress-responsive gene expression profiles in grain amaranth. Results A total of 2,700,168 raw reads were obtained from six 454 pyrosequencing runs, which were assembled into 21,207 high quality sequences (20,408 isotigs + 799 contigs). The average sequence length was 1,064 bp and 930 bp for isotigs and contigs, respectively. Only 5,113 singletons were recovered after quality control. Contigs/isotigs were further incorporated into 15,667 isogroups. All unique sequences were queried against the nr, TAIR, UniRef100, UniRef50 and Amaranthaceae EST databases for annotation. Functional GO annotation was performed with all contigs/isotigs that produced significant hits with the TAIR database. Only 8,260 sequences were found to be homologous when the transcriptomes of A. tuberculatus and A. hypochondriacus were compared, most of which were associated with basic house-keeping processes. Digital expression analysis identified 1,971 differentially expressed genes in response to at least one of four stress treatments tested. These included several multiple-stress-inducible genes that could represent potential candidates for use in the engineering of stress-resistant plants. The transcriptomic data generated from pigmented stems shared similarity with findings reported in developing stems of Arabidopsis and black cottonwood (Populus trichocarpa). Conclusions This study represents the first large-scale transcriptomic analysis of A. hypochondriacus, considered to be a highly nutritious and stress-tolerant crop. Numerous genes were found to be induced in response to (a)biotic stress, many of which could further the understanding of the mechanisms that contribute to multiple stress-resistance in plants, a trait that has potential biotechnological applications in agriculture. PMID:21752295

Profiling the venom gland transcriptomes of Costa Rican snakes by 454 pyrosequencing

PubMed Central

2011-01-01

Background A long term research goal of venomics, of applied importance for improving current antivenom therapy, but also for drug discovery, is to understand the pharmacological potential of venoms. Individually or combined, proteomic and transcriptomic studies have demonstrated their feasibility to explore in depth the molecular diversity of venoms. In the absence of genome sequence, transcriptomes represent also valuable searchable databases for proteomic projects. Results The venom gland transcriptomes of 8 Costa Rican taxa from 5 genera (Crotalus, Bothrops, Atropoides, Cerrophidion, and Bothriechis) of pitvipers were investigated using high-throughput 454 pyrosequencing. 100,394 out of 330,010 masked reads produced significant hits in the available databases. 5.165,220 nucleotides (8.27%) were masked by RepeatMasker, the vast majority of which corresponding to class I (retroelements) and class II (DNA transposons) mobile elements. BLAST hits included 79,991 matches to entries of the taxonomic suborder Serpentes, of which 62,433 displayed similarity to documented venom proteins. Strong discrepancies between the transcriptome-computed and the proteome-gathered toxin compositions were obvious at first sight. Although the reasons underlaying this discrepancy are elusive, since no clear trend within or between species is apparent, the data indicate that individual mRNA species may be translationally controlled in a species-dependent manner. The minimum number of genes from each toxin family transcribed into the venom gland transcriptome of each species was calculated from multiple alignments of reads matched to a full-length reference sequence of each toxin family. Reads encoding ORF regions of Kazal-type inhibitor-like proteins were uniquely found in Bothriechis schlegelii and B. lateralis transcriptomes, suggesting a genus-specific recruitment event during the early-Middle Miocene. A transcriptome-based cladogram supports the large divergence between A. mexicanus and A. picadoi, and a closer kinship between A. mexicanus and C. godmani. Conclusions Our comparative next-generation sequencing (NGS) analysis reveals taxon-specific trends governing the formulation of the venom arsenal. Knowledge of the venom proteome provides hints on the translation efficiency of toxin-coding transcripts, contributing thereby to a more accurate interpretation of the transcriptome. The application of NGS to the analysis of snake venom transcriptomes, may represent the tool for opening the door to systems venomics. PMID:21605378

The Need for Integrated Approaches in Metabolic Engineering

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

Lechner, Anna; Brunk, Elizabeth; Keasling, Jay D.

This review highlights state-of-the-art procedures for heterologous small-molecule biosynthesis, the associated bottlenecks, and new strategies that have the potential to accelerate future accomplishments in metabolic engineering. We emphasize that a combination of different approaches over multiple time and size scales must b e considered for successful pathway engineering in a heterologous host. We have classified these optimization procedures based on the "system" that is being manipulated: transcriptome, translatome, proteome, or reactome. By bridging multiple disciplines, including molecular biology, biochemistry, biophysics, and computational sciences, we can create an integral framework for the discovery and implementation of novel biosynthetic production routes.

Development of genome- and transcriptome-derived microsatellites in related species of snapping shrimps with highly duplicated genomes.

PubMed

Gaynor, Kaitlyn M; Solomon, Joseph W; Siller, Stefanie; Jessell, Linnet; Duffy, J Emmett; Rubenstein, Dustin R

2017-11-01

Molecular markers are powerful tools for studying patterns of relatedness and parentage within populations and for making inferences about social evolution. However, the development of molecular markers for simultaneous study of multiple species presents challenges, particularly when species exhibit genome duplication or polyploidy. We developed microsatellite markers for Synalpheus shrimp, a genus in which species exhibit not only great variation in social organization, but also interspecific variation in genome size and partial genome duplication. From the four primary clades within Synalpheus, we identified microsatellites in the genomes of four species and in the consensus transcriptome of two species. Ultimately, we designed and tested primers for 143 microsatellite markers across 25 species. Although the majority of markers were disomic, many markers were polysomic for certain species. Surprisingly, we found no relationship between genome size and the number of polysomic markers. As expected, markers developed for a given species amplified better for closely related species than for more distant relatives. Finally, the markers developed from the transcriptome were more likely to work successfully and to be disomic than those developed from the genome, suggesting that consensus transcriptomes are likely to be conserved across species. Our findings suggest that the transcriptome, particularly consensus sequences from multiple species, can be a valuable source of molecular markers for taxa with complex, duplicated genomes. © 2017 John Wiley & Sons Ltd.

Divergence in Morris Water Maze-Based Cognitive Performance under Chronic Stress Is Associated with the Hippocampal Whole Transcriptomic Modification in Mice

PubMed Central

Jung, Seung H.; Brownlow, Milene L.; Pellegrini, Matteo; Jankord, Ryan

2017-01-01

Individual susceptibility determines the magnitude of stress effects on cognitive function. The hippocampus, a brain region of memory consolidation, is vulnerable to stressful environments, and the impact of stress on hippocampus may determine individual variability in cognitive performance. Therefore, the purpose of this study was to define the relationship between the divergence in spatial memory performance under chronically unpredictable stress and an associated transcriptomic alternation in hippocampus, the brain region of spatial memory consolidation. Multiple strains of BXD (B6 × D2) recombinant inbred mice went through a 4-week chronic variable stress (CVS) paradigm, and the Morris water maze (MWM) test was conducted during the last week of CVS to assess hippocampal-dependent spatial memory performance and grouped animals into low and high performing groups based on the cognitive performance. Using hippocampal whole transcriptome RNA-sequencing data, differential expression, PANTHER analysis, WGCNA, Ingenuity's upstream regulator analysis in the Ingenuity Pathway Analysis® and phenotype association analysis were conducted. Our data identified multiple genes and pathways that were significantly associated with chronic stress-associated cognitive modification and the divergence in hippocampal dependent memory performance under chronic stress. Biological pathways associated with memory performance following chronic stress included metabolism, neurotransmitter and receptor regulation, immune response and cellular process. The Ingenuity's upstream regulator analysis identified 247 upstream transcriptional regulators from 16 different molecule types. Transcripts predictive of cognitive performance under high stress included genes that are associated with a high occurrence of Alzheimer's and cognitive impairments (e.g., Ncl, Eno1, Scn9a, Slc19a3, Ncstn, Fos, Eif4h, Copa, etc.). Our results show that the variable effects of chronic stress on the hippocampal transcriptome are related to the ability to complete the MWM task and that the modulations of specific pathways are indicative of hippocampal dependent memory performance. Thus, the divergence in spatial memory performance following chronic stress is related to the unique pattern of gene expression within the hippocampus. PMID:28912681

A draft of the genome and four transcriptomes of a medicinal and pesticidal angiosperm Azadirachta indica

PubMed Central

2012-01-01

Background The Azadirachta indica (neem) tree is a source of a wide number of natural products, including the potent biopesticide azadirachtin. In spite of its widespread applications in agriculture and medicine, the molecular aspects of the biosynthesis of neem terpenoids remain largely unexplored. The current report describes the draft genome and four transcriptomes of A. indica and attempts to contextualise the sequence information in terms of its molecular phylogeny, transcript expression and terpenoid biosynthesis pathways. A. indica is the first member of the family Meliaceae to be sequenced using next generation sequencing approach. Results The genome and transcriptomes of A. indica were sequenced using multiple sequencing platforms and libraries. The A. indica genome is AT-rich, bears few repetitive DNA elements and comprises about 20,000 genes. The molecular phylogenetic analyses grouped A. indica together with Citrus sinensis from the Rutaceae family validating its conventional taxonomic classification. Comparative transcript expression analysis showed either exclusive or enhanced expression of known genes involved in neem terpenoid biosynthesis pathways compared to other sequenced angiosperms. Genome and transcriptome analyses in A. indica led to the identification of repeat elements, nucleotide composition and expression profiles of genes in various organs. Conclusions This study on A. indica genome and transcriptomes will provide a model for characterization of metabolic pathways involved in synthesis of bioactive compounds, comparative evolutionary studies among various Meliaceae family members and help annotate their genomes. A better understanding of molecular pathways involved in the azadirachtin synthesis in A. indica will pave ways for bulk production of environment friendly biopesticides. PMID:22958331

International Standards for Genomes, Transcriptomes, and Metagenomes

PubMed Central

Mason, Christopher E.; Afshinnekoo, Ebrahim; Tighe, Scott; Wu, Shixiu; Levy, Shawn

2017-01-01

Challenges and biases in preparing, characterizing, and sequencing DNA and RNA can have significant impacts on research in genomics across all kingdoms of life, including experiments in single-cells, RNA profiling, and metagenomics (across multiple genomes). Technical artifacts and contamination can arise at each point of sample manipulation, extraction, sequencing, and analysis. Thus, the measurement and benchmarking of these potential sources of error are of paramount importance as next-generation sequencing (NGS) projects become more global and ubiquitous. Fortunately, a variety of methods, standards, and technologies have recently emerged that improve measurements in genomics and sequencing, from the initial input material to the computational pipelines that process and annotate the data. Here we review current standards and their applications in genomics, including whole genomes, transcriptomes, mixed genomic samples (metagenomes), and the modified bases within each (epigenomes and epitranscriptomes). These standards, tools, and metrics are critical for quantifying the accuracy of NGS methods, which will be essential for robust approaches in clinical genomics and precision medicine. PMID:28337071

Transcriptomic analysis of Portunus trituberculatus reveals a critical role for WNT4 and WNT signalling in limb regeneration.

PubMed

Liu, Lei; Fu, Yuanyuan; Zhu, Fang; Mu, Changkao; Li, Ronghua; Song, Weiwei; Shi, Ce; Ye, Yangfang; Wang, Chunlin

2018-06-05

The swimming crab (Portunus trituberculatus) is among the most economically important seawater crustacean species in Asia. Despite its commercial importance and being well-studied status, genomic and transcriptomic data are scarce for this crab species. In the present study, limb bud tissue was collected at different developmental stages post amputation for transcriptomic analysis. Illumina RNA-sequencing was applied to characterise the limb regeneration transcriptome and identify the most characteristic genes. A total of 289,018 transcripts were obtained by clustering and assembly of clean reads, producing 150,869 unigenes with an average length of 956 bp. Subsequent analysis revealed WNT signalling as the key pathway involved in limb regeneration, with WNT4 a key mediator. Overall, limb regeneration appears to be regulated by multiple signalling pathways, with numerous cell differentiation, muscle growth, moult, metabolism, and immune-related genes upregulated, including WNT4, LAMA, FIP2, FSTL5, TNC, HUS1, SWI5, NCGL, SLC22, PLA2, Tdc2, SMOX, GDH, and SMPD4. This is the first experimental study done on regenerating claws of P. trituberculatus. These findings expand existing sequence resources for crab species, and will likely accelerate research into regeneration and development in crustaceans, particularly functional studies on genes involved in limb regeneration. Copyright © 2018 Elsevier B.V. All rights reserved.

Draft De Novo Transcriptome of the Rat Kangaroo Potorous tridactylus as a Tool for Cell Biology

PubMed Central

Udy, Dylan B.; Voorhies, Mark; Chan, Patricia P.; Lowe, Todd M.; Dumont, Sophie

2015-01-01

The rat kangaroo (long-nosed potoroo, Potorous tridactylus) is a marsupial native to Australia. Cultured rat kangaroo kidney epithelial cells (PtK) are commonly used to study cell biological processes. These mammalian cells are large, adherent, and flat, and contain large and few chromosomes—and are thus ideal for imaging intra-cellular dynamics such as those of mitosis. Despite this, neither the rat kangaroo genome nor transcriptome have been sequenced, creating a challenge for probing the molecular basis of these cellular dynamics. Here, we present the sequencing, assembly and annotation of the draft rat kangaroo de novo transcriptome. We sequenced 679 million reads that mapped to 347,323 Trinity transcripts and 20,079 Unigenes. We present statistics emerging from transcriptome-wide analyses, and analyses suggesting that the transcriptome covers full-length sequences of most genes, many with multiple isoforms. We also validate our findings with a proof-of-concept gene knockdown experiment. We expect that this high quality transcriptome will make rat kangaroo cells a more tractable system for linking molecular-scale function and cellular-scale dynamics. PMID:26252667

Draft De Novo Transcriptome of the Rat Kangaroo Potorous tridactylus as a Tool for Cell Biology.

PubMed

Udy, Dylan B; Voorhies, Mark; Chan, Patricia P; Lowe, Todd M; Dumont, Sophie

2015-01-01

The rat kangaroo (long-nosed potoroo, Potorous tridactylus) is a marsupial native to Australia. Cultured rat kangaroo kidney epithelial cells (PtK) are commonly used to study cell biological processes. These mammalian cells are large, adherent, and flat, and contain large and few chromosomes-and are thus ideal for imaging intra-cellular dynamics such as those of mitosis. Despite this, neither the rat kangaroo genome nor transcriptome have been sequenced, creating a challenge for probing the molecular basis of these cellular dynamics. Here, we present the sequencing, assembly and annotation of the draft rat kangaroo de novo transcriptome. We sequenced 679 million reads that mapped to 347,323 Trinity transcripts and 20,079 Unigenes. We present statistics emerging from transcriptome-wide analyses, and analyses suggesting that the transcriptome covers full-length sequences of most genes, many with multiple isoforms. We also validate our findings with a proof-of-concept gene knockdown experiment. We expect that this high quality transcriptome will make rat kangaroo cells a more tractable system for linking molecular-scale function and cellular-scale dynamics.

Delayed response to cold stress is characterized by successive metabolic shifts culminating in apple fruit peel necrosis.

PubMed

Gapper, Nigel E; Hertog, Maarten L A T M; Lee, Jinwook; Buchanan, David A; Leisso, Rachel S; Fei, Zhangjun; Qu, Guiqin; Giovannoni, James J; Johnston, Jason W; Schaffer, Robert J; Nicolaï, Bart M; Mattheis, James P; Watkins, Christopher B; Rudell, David R

2017-04-21

Superficial scald is a physiological disorder of apple fruit characterized by sunken, necrotic lesions appearing after prolonged cold storage, although initial injury occurs much earlier in the storage period. To determine the degree to which the transition to cell death is an active process and specific metabolism involved, untargeted metabolic and transcriptomic profiling was used to follow metabolism of peel tissue over 180 d of cold storage. The metabolome and transcriptome of peel destined to develop scald began to diverge from peel where scald was controlled using antioxidant (diphenylamine; DPA) or rendered insensitive to ethylene using 1-methylcyclopropene (1-MCP) beginning between 30 and 60 days of storage. Overall metabolic and transcriptomic shifts, representing multiple pathways and processes, occurred alongside α-farnesene oxidation and, later, methanol production alongside symptom development. Results indicate this form of peel necrosis is a product of an active metabolic transition involving multiple pathways triggered by chilling temperatures at cold storage inception rather than physical injury. Among multiple other pathways, enhanced methanol and methyl ester levels alongside upregulated pectin methylesterases are unique to peel that is developing scald symptoms similar to injury resulting from mechanical stress and herbivory in other plants.

Nutrigenomics, the Microbiome, and Gene-Environment Interactions: New Directions in Cardiovascular Disease Research, Prevention, and Treatment: A Scientific Statement From the American Heart Association.

PubMed

Ferguson, Jane F; Allayee, Hooman; Gerszten, Robert E; Ideraabdullah, Folami; Kris-Etherton, Penny M; Ordovás, José M; Rimm, Eric B; Wang, Thomas J; Bennett, Brian J

2016-06-01

Cardiometabolic diseases are the leading cause of death worldwide and are strongly linked to both genetic and nutritional factors. The field of nutrigenomics encompasses multiple approaches aimed at understanding the effects of diet on health or disease development, including nutrigenetic studies investigating the relationship between genetic variants and diet in modulating cardiometabolic risk, as well as the effects of dietary components on multiple "omic" measures, including transcriptomics, metabolomics, proteomics, lipidomics, epigenetic modifications, and the microbiome. Here, we describe the current state of the field of nutrigenomics with respect to cardiometabolic disease research and outline a direction for the integration of multiple omics techniques in future nutrigenomic studies aimed at understanding mechanisms and developing new therapeutic options for cardiometabolic disease treatment and prevention. © 2016 American Heart Association, Inc.

Transcriptome analysis of the honey bee fungal pathogen, Ascosphaera apis: implications for host pathogenesis

PubMed Central

2012-01-01

Background We present a comprehensive transcriptome analysis of the fungus Ascosphaera apis, an economically important pathogen of the Western honey bee (Apis mellifera) that causes chalkbrood disease. Our goals were to further annotate the A. apis reference genome and to identify genes that are candidates for being differentially expressed during host infection versus axenic culture. Results We compared A. apis transcriptome sequence from mycelia grown on liquid or solid media with that dissected from host-infected tissue. 454 pyrosequencing provided 252 Mb of filtered sequence reads from both culture types that were assembled into 10,087 contigs. Transcript contigs, protein sequences from multiple fungal species, and ab initio gene predictions were included as evidence sources in the Maker gene prediction pipeline, resulting in 6,992 consensus gene models. A phylogeny based on 12 of these protein-coding loci further supported the taxonomic placement of Ascosphaera as sister to the core Onygenales. Several common protein domains were less abundant in A. apis compared with related ascomycete genomes, particularly cytochrome p450 and protein kinase domains. A novel gene family was identified that has expanded in some ascomycete lineages, but not others. We manually annotated genes with homologs in other fungal genomes that have known relevance to fungal virulence and life history. Functional categories of interest included genes involved in mating-type specification, intracellular signal transduction, and stress response. Computational and manual annotations have been made publicly available on the Bee Pests and Pathogens website. Conclusions This comprehensive transcriptome analysis substantially enhances our understanding of the A. apis genome and its expression during infection of honey bee larvae. It also provides resources for future molecular studies of chalkbrood disease and ultimately improved disease management. PMID:22747707

A Transcriptomic Analysis of Cave, Surface, and Hybrid Isopod Crustaceans of the Species Asellus aquaticus

PubMed Central

Stahl, Bethany A.; Gross, Joshua B.; Speiser, Daniel I.; Oakley, Todd H.; Patel, Nipam H.; Gould, Douglas B.; Protas, Meredith E.

2015-01-01

Cave animals, compared to surface-dwelling relatives, tend to have reduced eyes and pigment, longer appendages, and enhanced mechanosensory structures. Pressing questions include how certain cave-related traits are gained and lost, and if they originate through the same or different genetic programs in independent lineages. An excellent system for exploring these questions is the isopod, Asellus aquaticus. This species includes multiple cave and surface populations that have numerous morphological differences between them. A key feature is that hybrids between cave and surface individuals are viable, which enables genetic crosses and linkage analyses. Here, we advance this system by analyzing single animal transcriptomes of Asellus aquaticus. We use high throughput sequencing of non-normalized cDNA derived from the head of a surface-dwelling male, the head of a cave-dwelling male, the head of a hybrid male (produced by crossing a surface individual with a cave individual), and a pooled sample of surface embryos and hatchlings. Assembling reads from surface and cave head RNA pools yielded an integrated transcriptome comprised of 23,984 contigs. Using this integrated assembly as a reference transcriptome, we aligned reads from surface-, cave- and hybrid- head tissue and pooled surface embryos and hatchlings. Our approach identified 742 SNPs and placed four new candidate genes to an existing linkage map for A. aquaticus. In addition, we examined SNPs for allele-specific expression differences in the hybrid individual. All of these resources will facilitate identification of genes and associated changes responsible for cave adaptation in A. aquaticus and, in concert with analyses of other species, will inform our understanding of the evolutionary processes accompanying adaptation to the subterranean environment. PMID:26462237

Whole transcriptome analysis using next-generation sequencing of model species Setaria viridis to support C4 photosynthesis research.

PubMed

Xu, Jiajia; Li, Yuanyuan; Ma, Xiuling; Ding, Jianfeng; Wang, Kai; Wang, Sisi; Tian, Ye; Zhang, Hui; Zhu, Xin-Guang

2013-09-01

Setaria viridis is an emerging model species for genetic studies of C4 photosynthesis. Many basic molecular resources need to be developed to support for this species. In this paper, we performed a comprehensive transcriptome analysis from multiple developmental stages and tissues of S. viridis using next-generation sequencing technologies. Sequencing of the transcriptome from multiple tissues across three developmental stages (seed germination, vegetative growth, and reproduction) yielded a total of 71 million single end 100 bp long reads. Reference-based assembly using Setaria italica genome as a reference generated 42,754 transcripts. De novo assembly generated 60,751 transcripts. In addition, 9,576 and 7,056 potential simple sequence repeats (SSRs) covering S. viridis genome were identified when using the reference based assembled transcripts and the de novo assembled transcripts, respectively. This identified transcripts and SSR provided by this study can be used for both reverse and forward genetic studies based on S. viridis.

UniVIO: A Multiple Omics Database with Hormonome and Transcriptome Data from Rice

PubMed Central

Sakurai, Tetsuya; Sakakibara, Hitoshi

2013-01-01

Plant hormones play important roles as signaling molecules in the regulation of growth and development by controlling the expression of downstream genes. Since the hormone signaling system represents a complex network involving functional cross-talk through the mutual regulation of signaling and metabolism, a comprehensive and integrative analysis of plant hormone concentrations and gene expression is important for a deeper understanding of hormone actions. We have developed a database named Uniformed Viewer for Integrated Omics (UniVIO: http://univio.psc.riken.jp/), which displays hormone-metabolome (hormonome) and transcriptome data in a single formatted (uniformed) heat map. At the present time, hormonome and transcriptome data obtained from 14 organ parts of rice plants at the reproductive stage and seedling shoots of three gibberellin signaling mutants are included in the database. The hormone concentration and gene expression data can be searched by substance name, probe ID, gene locus ID or gene description. A correlation search function has been implemented to enable users to obtain information of correlated substance accumulation and gene expression. In the correlation search, calculation method, range of correlation coefficient and plant samples can be selected freely. PMID:23314752

Salicylic acid is an indispensable component of the Ny-1 resistance-gene-mediated response against Potato virus Y infection in potato

PubMed Central

Baebler, Š.; Witek, K.; Gruden, K.; Hennig, J.

2014-01-01

The purpose of the study was to investigate the role of salicylic acid (SA) signalling in Ny-1-mediated hypersensitive resistance (HR) of potato (Solanum tuberosum L.) to Potato virus Y (PVY). The responses of the Ny-1 allele in the Rywal potato cultivar and transgenic NahG-Rywal potato plants that do not accumulate SA were characterized at the cytological, biochemical, transcriptome, and proteome levels. Analysis of noninoculated and inoculated leaves revealed that HR lesions started to develop from 3 d post inoculation and completely restricted the virus spread. At the cytological level, features of programmed cell death in combination with reactive oxygen species burst were observed. In response to PVY infection, SA was synthesized de novo. The lack of SA accumulation in the NahG plants led to the disease phenotype due to unrestricted viral spreading. Grafting experiments show that SA has a critical role in the inhibition of PVY spreading in parenchymal tissue, but not in vascular veins. The whole transcriptome analysis confirmed the central role of SA in orchestrating Ny-1-mediated responses and showed that the absence of SA leads to significant changes at the transcriptome level, including a delay in activation of expression of genes known to participate in defence responses. Moreover, perturbations in the expression of hormonal signalling genes were detected, shown as a switch from SA to jasmonic acid/ethylene signalling. Viral multiplication in the NahG plants was accompanied by downregulation of photosynthesis genes and activation of multiple energy-producing pathways. PMID:24420577

Salicylic acid is an indispensable component of the Ny-1 resistance-gene-mediated response against Potato virus Y infection in potato.

PubMed

Baebler, Š; Witek, K; Petek, M; Stare, K; Tušek-Žnidarič, M; Pompe-Novak, M; Renaut, J; Szajko, K; Strzelczyk-Żyta, D; Marczewski, W; Morgiewicz, K; Gruden, K; Hennig, J

2014-03-01

The purpose of the study was to investigate the role of salicylic acid (SA) signalling in Ny-1-mediated hypersensitive resistance (HR) of potato (Solanum tuberosum L.) to Potato virus Y (PVY). The responses of the Ny-1 allele in the Rywal potato cultivar and transgenic NahG-Rywal potato plants that do not accumulate SA were characterized at the cytological, biochemical, transcriptome, and proteome levels. Analysis of noninoculated and inoculated leaves revealed that HR lesions started to develop from 3 d post inoculation and completely restricted the virus spread. At the cytological level, features of programmed cell death in combination with reactive oxygen species burst were observed. In response to PVY infection, SA was synthesized de novo. The lack of SA accumulation in the NahG plants led to the disease phenotype due to unrestricted viral spreading. Grafting experiments show that SA has a critical role in the inhibition of PVY spreading in parenchymal tissue, but not in vascular veins. The whole transcriptome analysis confirmed the central role of SA in orchestrating Ny-1-mediated responses and showed that the absence of SA leads to significant changes at the transcriptome level, including a delay in activation of expression of genes known to participate in defence responses. Moreover, perturbations in the expression of hormonal signalling genes were detected, shown as a switch from SA to jasmonic acid/ethylene signalling. Viral multiplication in the NahG plants was accompanied by downregulation of photosynthesis genes and activation of multiple energy-producing pathways.

Environmental Interactions and Epistasis Are Revealed in the Proteomic Responses to Complex Stimuli

PubMed Central

Samir, Parimal; Rahul; Slaughter, James C.; Link, Andrew J.

2015-01-01

Ultimately, the genotype of a cell and its interaction with the environment determine the cell’s biochemical state. While the cell’s response to a single stimulus has been studied extensively, a conceptual framework to model the effect of multiple environmental stimuli applied concurrently is not as well developed. In this study, we developed the concepts of environmental interactions and epistasis to explain the responses of the S. cerevisiae proteome to simultaneous environmental stimuli. We hypothesize that, as an abstraction, environmental stimuli can be treated as analogous to genetic elements. This would allow modeling of the effects of multiple stimuli using the concepts and tools developed for studying gene interactions. Mirroring gene interactions, our results show that environmental interactions play a critical role in determining the state of the proteome. We show that individual and complex environmental stimuli behave similarly to genetic elements in regulating the cellular responses to stimuli, including the phenomena of dominance and suppression. Interestingly, we observed that the effect of a stimulus on a protein is dominant over other stimuli if the response to the stimulus involves the protein. Using publicly available transcriptomic data, we find that environmental interactions and epistasis regulate transcriptomic responses as well. PMID:26247773

Transcriptome Profiling of a Multiple Recurrent Muscle-Invasive Urothelial Carcinoma of the Bladder by Deep Sequencing

PubMed Central

Zhang, Shufang; Liu, Yanxuan; Liu, Zhenxiang; Zhang, Chong; Cao, Hui; Ye, Yongqing; Wang, Shunlan; Zhang, Ying'ai; Xiao, Sifang; Yang, Peng; Li, Jindong; Bai, Zhiming

2014-01-01

Urothelial carcinoma of the bladder (UCB) is one of the commonly diagnosed cancers in the world. The UCB has the highest rate of recurrence of any malignancy. A genome-wide screening of transcriptome dysregulation between cancer and normal tissue would provide insight into the molecular basis of UCB recurrence and is a key step to discovering biomarkers for diagnosis and therapeutic targets. Compared with microarray technology, which is commonly used to identify expression level changes, the recently developed RNA-seq technique has the ability to detect other abnormal regulations in the cancer transcriptome, such as alternative splicing. In this study, we performed high-throughput transcriptome sequencing at ∼50× coverage on a recurrent muscle-invasive cisplatin-resistance UCB tissue and the adjacent non-tumor tissue. The results revealed cancer-specific differentially expressed genes between the tumor and non-tumor tissue enriched in the cell adhesion molecules, focal adhesion and ECM-receptor interaction pathway. Five dysregulated genes, including CDH1, VEGFA, PTPRF, CLDN7, and MMP2 were confirmed by Real time qPCR in the sequencing samples and the additional eleven samples. Our data revealed that more than three hundred genes showed differential splicing patterns between tumor tissue and non-tumor tissue. Among these genes, we filtered 24 cancer-associated alternative splicing genes with differential exon usage. The findings from RNA-Seq were validated by Real time qPCR for CD44, PDGFA, NUMB, and LPHN2. This study provides a comprehensive survey of the UCB transcriptome, which provides better insight into the complexity of regulatory changes during recurrence and metastasis. PMID:24622401

Genome and Transcriptome Sequencing of the Ostreid herpesvirus 1 From Tomales Bay, California

NASA Astrophysics Data System (ADS)

Burge, C. A.; Langevin, S.; Closek, C. J.; Roberts, S. B.; Friedman, C. S.

2016-02-01

Mass mortalities of larval and seed bivalve molluscs attributed to the Ostreid herpesvirus 1 (OsHV-1) occur globally. OsHV-1 was fully sequenced and characterized as a member of the Family Malacoherpesviridae. Multiple strains of OsHV-1 exist and may vary in virulence, i.e. OsHV-1 µvar. For most global variants of OsHV-1, sequence data is limited to PCR-based sequencing of segments, including two recent genomes. In the United States, OsHV-1 is limited to detection in adjacent embayments in California, Tomales and Drakes bays. Limited DNA sequence data of OsHV-1 infecting oysters in Tomales Bay indicates the virus detected in Tomales Bay is similar but not identical to any one global variant of OsHV-1. In order to better understand both strain variation and virulence of OsHV-1 infecting oysters in Tomales Bay, we used genomic and transcriptomic sequencing. Meta-genomic sequencing (Illumina MiSeq) was conducted from infected oysters (n=4 per year) collected in 2003, 2007, and 2014, where full OsHV-1 genome sequences and low overall microbial diversity were achieved from highly infected oysters. Increased microbial diversity was detected in three of four samples sequenced from 2003, where qPCR based genome copy numbers of OsHV-1 were lower. Expression analysis (SOLiD RNA sequencing) of OsHV-1 genes expressed in oyster larvae at 24 hours post exposure revealed a nearly complete transcriptome, with several highly expressed genes, which are similar to recent transcriptomic analyses of other OsHV-1 variants. Taken together, our results indicate that genome and transcriptome sequencing may be powerful tools in understanding both strain variation and virulence of non-culturable marine viruses.

A Single Transcriptome of a Green Toad (Bufo viridis) Yields Candidate Genes for Sex Determination and -Differentiation and Non-Anonymous Population Genetic Markers

PubMed Central

Gerchen, Jörn F.; Reichert, Samuel J.; Röhr, Johannes T.; Dieterich, Christoph; Kloas, Werner

2016-01-01

Large genome size, including immense repetitive and non-coding fractions, still present challenges for capacity, bioinformatics and thus affordability of whole genome sequencing in most amphibians. Here, we test the performance of a single transcriptome to understand whether it can provide a cost-efficient resource for species with large unknown genomes. Using RNA from six different tissues from a single Palearctic green toad (Bufo viridis) specimen and Hiseq2000, we obtained 22,5 Mio reads and publish >100,000 unigene sequences. To evaluate efficacy and quality, we first use this data to identify green toad specific candidate genes, known from other vertebrates for their role in sex determination and differentiation. Of a list of 37 genes, the transcriptome yielded 32 (87%), many of which providing the first such data for this non-model anuran species. However, for many of these genes, only fragments could be retrieved. In order to allow also applications to population genetics, we further used the transcriptome for the targeted development of 21 non-anonymous microsatellites and tested them in genetic families and backcrosses. Eleven markers were specifically developed to be located on the B. viridis sex chromosomes; for eight markers we can indeed demonstrate sex-specific transmission in genetic families. Depending on phylogenetic distance, several markers, which are sex-linked in green toads, show high cross-amplification success across the anuran phylogeny, involving nine systematic anuran families. Our data support the view that single transcriptome sequencing (based on multiple tissues) provides a reliable genomic resource and cost-efficient method for non-model amphibian species with large genome size and, despite limitations, should be considered as long as genome sequencing remains unaffordable for most species. PMID:27232626

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

PubMed Central

Neymotin, Benjamin; Ettorre, Victoria; Gresham, David

2016-01-01

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

Transcriptome analysis of woodland strawberry (Fragaria vesca) response to the infection by Strawberry vein banding virus (SVBV).

PubMed

Chen, Jing; Zhang, Hanping; Feng, Mingfeng; Zuo, Dengpan; Hu, Yahui; Jiang, Tong

2016-07-13

Woodland strawberry (Fragaria vesca) infected with Strawberry vein banding virus (SVBV) exhibits chlorotic symptoms along the leaf veins. However, little is known about the molecular mechanism of strawberry disease caused by SVBV. We performed the next-generation sequencing (RNA-Seq) study to identify gene expression changes induced by SVBV in woodland strawberry using mock-inoculated plants as a control. Using RNA-Seq, we have identified 36,850 unigenes, of which 517 were differentially expressed in the virus-infected plants (DEGs). The unigenes were annotated and classified with Gene Ontology (GO), Clusters of Orthologous Group (COG) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. The KEGG pathway analysis of these genes suggested that strawberry disease caused by SVBV may affect multiple processes including pigment metabolism, photosynthesis and plant-pathogen interactions. Our research provides comprehensive transcriptome information regarding SVBV infection in strawberry.

Tissue-Specific Transcriptome Profiling of Plutella Xylostella Third Instar Larval Midgut

PubMed Central

Xie, Wen; Lei, Yanyuan; Fu, Wei; Yang, Zhongxia; Zhu, Xun; Guo, Zhaojiang; Wu, Qingjun; Wang, Shaoli; Xu, Baoyun; Zhou, Xuguo; Zhang, Youjun

2012-01-01

The larval midgut of diamondback moth, Plutella xylostella, is a dynamic tissue that interfaces with a diverse array of physiological and toxicological processes, including nutrient digestion and allocation, xenobiotic detoxification, innate and adaptive immune response, and pathogen defense. Despite its enormous agricultural importance, the genomic resources for P. xylostella are surprisingly scarce. In this study, a Bt resistant P. xylostella strain was subjected to the in-depth transcriptome analysis to identify genes and gene networks putatively involved in various physiological and toxicological processes in the P. xylostella larval midgut. Using Illumina deep sequencing, we obtained roughly 40 million reads containing approximately 3.6 gigabases of sequence data. De novo assembly generated 63,312 ESTs with an average read length of 416bp, and approximately half of the P. xylostella sequences (45.4%, 28,768) showed similarity to the non-redundant database in GenBank with a cut-off E-value below 10-5. Among them, 11,092 unigenes were assigned to one or multiple GO terms and 16,732 unigenes were assigned to 226 specific pathways. In-depth analysis indentified genes putatively involved in insecticide resistance, nutrient digestion, and innate immune defense. Besides conventional detoxification enzymes and insecticide targets, novel genes, including 28 chymotrypsins and 53 ABC transporters, have been uncovered in the P. xylostella larval midgut transcriptome; which are potentially linked to the Bt toxicity and resistance. Furthermore, an unexpectedly high number of ESTs, including 46 serpins and 7 lysozymes, were predicted to be involved in the immune defense. As the first tissue-specific transcriptome analysis of P. xylostella, this study sheds light on the molecular understanding of insecticide resistance, especially Bt resistance in an agriculturally important insect pest, and lays the foundation for future functional genomics research. In addition, current sequencing effort greatly enriched the existing P. xylostella EST database, and makes RNAseq a viable option in the future genomic analysis. PMID:23091412

Tissue-specific transcriptome profiling of Plutella xylostella third instar larval midgut.

PubMed

Xie, Wen; Lei, Yanyuan; Fu, Wei; Yang, Zhongxia; Zhu, Xun; Guo, Zhaojiang; Wu, Qingjun; Wang, Shaoli; Xu, Baoyun; Zhou, Xuguo; Zhang, Youjun

2012-01-01

The larval midgut of diamondback moth, Plutella xylostella, is a dynamic tissue that interfaces with a diverse array of physiological and toxicological processes, including nutrient digestion and allocation, xenobiotic detoxification, innate and adaptive immune response, and pathogen defense. Despite its enormous agricultural importance, the genomic resources for P. xylostella are surprisingly scarce. In this study, a Bt resistant P. xylostella strain was subjected to the in-depth transcriptome analysis to identify genes and gene networks putatively involved in various physiological and toxicological processes in the P. xylostella larval midgut. Using Illumina deep sequencing, we obtained roughly 40 million reads containing approximately 3.6 gigabases of sequence data. De novo assembly generated 63,312 ESTs with an average read length of 416 bp, and approximately half of the P. xylostella sequences (45.4%, 28,768) showed similarity to the non-redundant database in GenBank with a cut-off E-value below 10(-5). Among them, 11,092 unigenes were assigned to one or multiple GO terms and 16,732 unigenes were assigned to 226 specific pathways. In-depth analysis identified genes putatively involved in insecticide resistance, nutrient digestion, and innate immune defense. Besides conventional detoxification enzymes and insecticide targets, novel genes, including 28 chymotrypsins and 53 ABC transporters, have been uncovered in the P. xylostella larval midgut transcriptome; which are potentially linked to the Bt toxicity and resistance. Furthermore, an unexpectedly high number of ESTs, including 46 serpins and 7 lysozymes, were predicted to be involved in the immune defense.As the first tissue-specific transcriptome analysis of P. xylostella, this study sheds light on the molecular understanding of insecticide resistance, especially Bt resistance in an agriculturally important insect pest, and lays the foundation for future functional genomics research. In addition, current sequencing effort greatly enriched the existing P. xylostella EST database, and makes RNAseq a viable option in the future genomic analysis.

Comparative transcriptomics of elasmobranchs and teleosts highlight important processes in adaptive immunity and regional endothermy.

PubMed

Marra, Nicholas J; Richards, Vincent P; Early, Angela; Bogdanowicz, Steve M; Pavinski Bitar, Paulina D; Stanhope, Michael J; Shivji, Mahmood S

2017-01-30

Comparative genomic and/or transcriptomic analyses involving elasmobranchs remain limited, with genome level comparisons of the elasmobranch immune system to that of higher vertebrates, non-existent. This paper reports a comparative RNA-seq analysis of heart tissue from seven species, including four elasmobranchs and three teleosts, focusing on immunity, but concomitantly seeking to identify genetic similarities shared by the two lamnid sharks and the single billfish in our study, which could be linked to convergent evolution of regional endothermy. Across seven species, we identified an average of 10,877 Swiss-Prot annotated genes from an average of 32,474 open reading frames within each species' heart transcriptome. About half of these genes were shared between all species while the remainder included functional differences between our groups of interest (elasmobranch vs. teleost and endotherms vs. ectotherms) as revealed by Gene Ontology (GO) and selection analyses. A repeatedly represented functional category, in both the uniquely expressed elasmobranch genes (total of 259) and the elasmobranch GO enrichment results, involved antibody-mediated immunity, either in the recruitment of immune cells (Fc receptors) or in antigen presentation, including such terms as "antigen processing and presentation of exogenous peptide antigen via MHC class II", and such genes as MHC class II, HLA-DPB1. Molecular adaptation analyses identified three genes in elasmobranchs with a history of positive selection, including legumain (LGMN), a gene with roles in both innate and adaptive immunity including producing antigens for presentation by MHC class II. Comparisons between the endothermic and ectothermic species revealed an enrichment of GO terms associated with cardiac muscle contraction in endotherms, with 19 genes expressed solely in endotherms, several of which have significant roles in lipid and fat metabolism. This collective comparative evidence provides the first multi-taxa transcriptomic-based perspective on differences between elasmobranchs and teleosts, and suggests various unique features associated with the adaptive immune system of elasmobranchs, pointing in particular to the potential importance of MHC Class II. This in turn suggests that expanded comparative work involving additional tissues, as well as genome sequencing of multiple elasmobranch species would be productive in elucidating the regulatory and genome architectural hallmarks of elasmobranchs.

SC3 - consensus clustering of single-cell RNA-Seq data

PubMed Central

Kiselev, Vladimir Yu.; Kirschner, Kristina; Schaub, Michael T.; Andrews, Tallulah; Yiu, Andrew; Chandra, Tamir; Natarajan, Kedar N; Reik, Wolf; Barahona, Mauricio; Green, Anthony R; Hemberg, Martin

2017-01-01

Single-cell RNA-seq (scRNA-seq) enables a quantitative cell-type characterisation based on global transcriptome profiles. We present Single-Cell Consensus Clustering (SC3), a user-friendly tool for unsupervised clustering which achieves high accuracy and robustness by combining multiple clustering solutions through a consensus approach. We demonstrate that SC3 is capable of identifying subclones based on the transcriptomes from neoplastic cells collected from patients. PMID:28346451

Trinity | Informatics Technology for Cancer Research (ITCR)

Cancer.gov

Trinity Cancer Transcriptome Analysis Toolkit (CTAT) including de novo transcriptome assembly with downstream support for expression analysis and focused analyses on cancer transcriptomes, incorporating mutation and fusion transcript discovery, and single cell analysis.

Single-cell transcriptomics using spliced leader PCR: Evidence for multiple losses of photosynthesis in polykrikoid dinoflagellates.

PubMed

Gavelis, Gregory S; White, Richard A; Suttle, Curtis A; Keeling, Patrick J; Leander, Brian S

2015-07-17

Most microbial eukaryotes are uncultivated and thus poorly suited to standard genomic techniques. This is the case for Polykrikos lebouriae, a dinoflagellate with ultrastructurally aberrant plastids. It has been suggested that these plastids stem from a novel symbiosis with either a diatom or haptophyte, but this hypothesis has been difficult to test as P. lebouriae dwells in marine sand rife with potential genetic contaminants. We applied spliced-leader targeted PCR (SLPCR) to obtain dinoflagellate-specific transcriptomes on single-cell isolates of P. lebouriae from marine sediments. Polykrikos lebouriae expressed nuclear-encoded photosynthetic genes that were characteristic of the peridinin-plastids of dinoflagellates, rather than those from a diatom of haptophyte. We confirmed these findings at the genomic level using multiple displacement amplification (MDA) to obtain a partial plastome of P. lebouriae. From these data, we infer that P. lebouriae has retained the peridinin plastids ancestral for dinoflagellates as a whole, while its closest relatives have lost photosynthesis multiple times independently. We discuss these losses with reference to mixotrophy in polykrikoid dinoflagellates. Our findings demonstrate new levels of variation associated with the peridinin plastids of dinoflagellates and the usefulness of SLPCR approaches on single cell isolates. Unlike other transcriptomic methods, SLPCR has taxonomic specificity, and can in principle be adapted to different splice-leader bearing groups.

Ethyl carbamate induces cell death through its effects on multiple metabolic pathways.

PubMed

Liu, Huichang; Cui, Bo; Xu, Yi; Hu, Chaoyang; Liu, Ying; Qu, Guorun; Li, Dawei; Wu, Yongning; Zhang, Dabing; Quan, Sheng; Shi, Jianxin

2017-11-01

Ethyl carbamate (EC), a multisite carcinogenic chemical causing tumors in various animal species, is probably carcinogenic to humans. However, information about the possible carcinogenic and toxicological effects of EC in humans is quite limited. Because EC is found in many dietary foods (such as fermented foods) and tobacco and its products, and exposure of humans to EC often occurs inevitably, its toxicological effects in humans need to be studied. This study was conducted to understand the metabolomic and transcriptomic changes in human hepatocellular carcinoma cells (HepG2) exposed to 100 mM EC for short term (4 h) and long term (12 h) period, respectively. The results revealed multiple influences of EC on the metabolome and transcriptome of HepG2 cells, which was exposure time-dependent and well correlated with the kinetic changes of cell viability and mortality. EC treatment affected multiple metabolic pathways, inducing oxidative stress, reducing detoxification capacity, depleting energy, decreasing reducing power, disrupting membrane integrity, and damaging DNA and protein. These metabolomic and transcriptomic biomarkers of EC on human cell metabolism identified in this study would facilitate further studies on the risk assessment and the mitigation of dietary EC. Copyright © 2017 Elsevier B.V. All rights reserved.

-A curated transcriptomic dataset collection relevant to embryonic development associated with in vitro fertilization in healthy individuals and patients with polycystic ovary syndrome.

PubMed

Mackeh, Rafah; Boughorbel, Sabri; Chaussabel, Damien; Kino, Tomoshige

2017-01-01

The collection of large-scale datasets available in public repositories is rapidly growing and providing opportunities to identify and fill gaps in different fields of biomedical research. However, users of these datasets should be able to selectively browse datasets related to their field of interest. Here we made available a collection of transcriptome datasets related to human follicular cells from normal individuals or patients with polycystic ovary syndrome, in the process of their development, during in vitro fertilization. After RNA-seq dataset exclusion and careful selection based on study description and sample information, 12 datasets, encompassing a total of 85 unique transcriptome profiles, were identified in NCBI Gene Expression Omnibus and uploaded to the Gene Expression Browser (GXB), a web application specifically designed for interactive query and visualization of integrated large-scale data. Once annotated in GXB, multiple sample grouping has been made in order to create rank lists to allow easy data interpretation and comparison. The GXB tool also allows the users to browse a single gene across multiple projects to evaluate its expression profiles in multiple biological systems/conditions in a web-based customized graphical views. The curated dataset is accessible at the following link: http://ivf.gxbsidra.org/dm3/landing.gsp.

­A curated transcriptomic dataset collection relevant to embryonic development associated with in vitro fertilization in healthy individuals and patients with polycystic ovary syndrome

PubMed Central

Mackeh, Rafah; Boughorbel, Sabri; Chaussabel, Damien; Kino, Tomoshige

2017-01-01

The collection of large-scale datasets available in public repositories is rapidly growing and providing opportunities to identify and fill gaps in different fields of biomedical research. However, users of these datasets should be able to selectively browse datasets related to their field of interest. Here we made available a collection of transcriptome datasets related to human follicular cells from normal individuals or patients with polycystic ovary syndrome, in the process of their development, during in vitro fertilization. After RNA-seq dataset exclusion and careful selection based on study description and sample information, 12 datasets, encompassing a total of 85 unique transcriptome profiles, were identified in NCBI Gene Expression Omnibus and uploaded to the Gene Expression Browser (GXB), a web application specifically designed for interactive query and visualization of integrated large-scale data. Once annotated in GXB, multiple sample grouping has been made in order to create rank lists to allow easy data interpretation and comparison. The GXB tool also allows the users to browse a single gene across multiple projects to evaluate its expression profiles in multiple biological systems/conditions in a web-based customized graphical views. The curated dataset is accessible at the following link: http://ivf.gxbsidra.org/dm3/landing.gsp. PMID:28413616

The Need for Integrated Approaches in Metabolic Engineering.

PubMed

Lechner, Anna; Brunk, Elizabeth; Keasling, Jay D

2016-11-01

This review highlights state-of-the-art procedures for heterologous small-molecule biosynthesis, the associated bottlenecks, and new strategies that have the potential to accelerate future accomplishments in metabolic engineering. We emphasize that a combination of different approaches over multiple time and size scales must be considered for successful pathway engineering in a heterologous host. We have classified these optimization procedures based on the "system" that is being manipulated: transcriptome, translatome, proteome, or reactome. By bridging multiple disciplines, including molecular biology, biochemistry, biophysics, and computational sciences, we can create an integral framework for the discovery and implementation of novel biosynthetic production routes. Copyright © 2016 Cold Spring Harbor Laboratory Press; all rights reserved.

Transcriptome profiles in sarcoidosis and their potential role in disease prediction.

PubMed

Schupp, Jonas C; Vukmirovic, Milica; Kaminski, Naftali; Prasse, Antje

2017-09-01

Sarcoidosis is a systemic disease defined by the presence of nonnecrotizing granuloma in the absence of any known cause. Although the heterogeneity of sarcoidosis is well characterized clinically, the transcriptome of sarcoidosis and underlying molecular mechanisms are not. The signal of all transcripts, small and long noncoding RNAs, can be detected using microarrays or RNA-Sequencing. Analyzing the transcriptome of tissues that are directly affected by granulomas is of great importance to understand biology of the disease and may be predictive of disease and treatment outcome. Multiple genome wide expression studies performed on sarcoidosis affected tissues were published in the last 11 years. Published studies focused on differences in gene expression between sarcoidosis vs. control tissues, stable vs. progressive sarcoidosis, as well as sarcoidosis vs. other diseases. Strikingly, all these transcriptomics data confirm the key role of TH1 immune response in sarcoidosis and particularly of interferon-γ (IFN-γ) and type I IFN-driven signaling pathways. The steps toward transcriptomics of sarcoidosis in precision medicine highlight the potentials of this approach. Large prospective follow-up studies are required to identify signatures predictive of disease progression and outcome.

Global exosome transcriptome profiling reveals biomarkers for multiple sclerosis.

PubMed

Selmaj, Igor; Cichalewska, Maria; Namiecinska, Magdalena; Galazka, Grazyna; Horzelski, Wojciech; Selmaj, Krzysztof W; Mycko, Marcin P

2017-05-01

Accumulating evidence supports a role for exosomes in immune regulation. In this study, we investigated the total circulating exosome transcriptome in relapsing-remitting multiple sclerosis (RRMS) patients and healthy controls (HC). Next generation sequencing (NGS) was used to define the global RNA profile of serum exosomes in 19 RRMS patients (9 in relapse, 10 in remission) and 10 HC. We analyzed 5 million reads and >50,000 transcripts per sample, including a detailed analysis of microRNAs (miRNAs) differentially expressed in RRMS. The discovery set data were validated by quantification using digital quantitative polymerase chain reaction with an independent cohort of 63 RRMS patients (33 in relapse, 30 in remission) and 32 HC. Exosomal RNA NGS revealed that of 15 different classes of transcripts detected, 4 circulating exosomal sequences within the miRNA category were differentially expressed in RRMS patients versus HC: hsa-miR-122-5p, hsa-miR-196b-5p, hsa-miR-301a-3p, and hsa-miR-532-5p. Serum exosomal expression of these miRNAs was significantly decreased during relapse in RRMS. These miRNAs were also decreased in patients with a gadolinium enhancement on brain magnetic resonance imaging. In vitro secretion of these miRNAs by peripheral blood mononuclear cells was also significantly impaired in RRMS. These data show that circulating exosomes have a distinct RNA profile in RRMS. Because putative targets for these miRNAs include the signal transducer and activator of transcription 3 and the cell cycle regulator aryl hydrocarbon receptor, the data suggest a disturbed cell-to-cell communication in this disease. Thus, exosomal miRNAs might represent a useful biomarker to distinguish multiple sclerosis relapse. Ann Neurol 2017;81:703-717. © 2017 American Neurological Association.

Allopatric integrations selectively change host transcriptomes, leading to varied expression efficiencies of exotic genes in Myxococcus xanthus.

PubMed

Zhu, Li-Ping; Yue, Xin-Jing; Han, Kui; Li, Zhi-Feng; Zheng, Lian-Shuai; Yi, Xiu-Nan; Wang, Hai-Long; Zhang, You-Ming; Li, Yue-Zhong

2015-07-22

Exotic genes, especially clustered multiple-genes for a complex pathway, are normally integrated into chromosome for heterologous expression. The influences of insertion sites on heterologous expression and allotropic expressions of exotic genes on host remain mostly unclear. We compared the integration and expression efficiencies of single and multiple exotic genes that were inserted into Myxococcus xanthus genome by transposition and attB-site-directed recombination. While the site-directed integration had a rather stable chloramphenicol acetyl transferase (CAT) activity, the transposition produced varied CAT enzyme activities. We attempted to integrate the 56-kb gene cluster for the biosynthesis of antitumor polyketides epothilones into M. xanthus genome by site-direction but failed, which was determined to be due to the insertion size limitation at the attB site. The transposition technique produced many recombinants with varied production capabilities of epothilones, which, however, were not paralleled to the transcriptional characteristics of the local sites where the genes were integrated. Comparative transcriptomics analysis demonstrated that the allopatric integrations caused selective changes of host transcriptomes, leading to varied expressions of epothilone genes in different mutants. With the increase of insertion fragment size, transposition is a more practicable integration method for the expression of exotic genes. Allopatric integrations selectively change host transcriptomes, which lead to varied expression efficiencies of exotic genes.

Deep Learning Applications for Predicting Pharmacological Properties of Drugs and Drug Repurposing Using Transcriptomic Data.

PubMed

Aliper, Alexander; Plis, Sergey; Artemov, Artem; Ulloa, Alvaro; Mamoshina, Polina; Zhavoronkov, Alex

2016-07-05

Deep learning is rapidly advancing many areas of science and technology with multiple success stories in image, text, voice and video recognition, robotics, and autonomous driving. In this paper we demonstrate how deep neural networks (DNN) trained on large transcriptional response data sets can classify various drugs to therapeutic categories solely based on their transcriptional profiles. We used the perturbation samples of 678 drugs across A549, MCF-7, and PC-3 cell lines from the LINCS Project and linked those to 12 therapeutic use categories derived from MeSH. To train the DNN, we utilized both gene level transcriptomic data and transcriptomic data processed using a pathway activation scoring algorithm, for a pooled data set of samples perturbed with different concentrations of the drug for 6 and 24 hours. In both pathway and gene level classification, DNN achieved high classification accuracy and convincingly outperformed the support vector machine (SVM) model on every multiclass classification problem, however, models based on pathway level data performed significantly better. For the first time we demonstrate a deep learning neural net trained on transcriptomic data to recognize pharmacological properties of multiple drugs across different biological systems and conditions. We also propose using deep neural net confusion matrices for drug repositioning. This work is a proof of principle for applying deep learning to drug discovery and development.

Deep learning applications for predicting pharmacological properties of drugs and drug repurposing using transcriptomic data

PubMed Central

Aliper, Alexander; Plis, Sergey; Artemov, Artem; Ulloa, Alvaro; Mamoshina, Polina; Zhavoronkov, Alex

2016-01-01

Deep learning is rapidly advancing many areas of science and technology with multiple success stories in image, text, voice and video recognition, robotics and autonomous driving. In this paper we demonstrate how deep neural networks (DNN) trained on large transcriptional response data sets can classify various drugs to therapeutic categories solely based on their transcriptional profiles. We used the perturbation samples of 678 drugs across A549, MCF‐7 and PC‐3 cell lines from the LINCS project and linked those to 12 therapeutic use categories derived from MeSH. To train the DNN, we utilized both gene level transcriptomic data and transcriptomic data processed using a pathway activation scoring algorithm, for a pooled dataset of samples perturbed with different concentrations of the drug for 6 and 24 hours. In both gene and pathway level classification, DNN convincingly outperformed support vector machine (SVM) model on every multiclass classification problem, however, models based on a pathway level classification perform better. For the first time we demonstrate a deep learning neural net trained on transcriptomic data to recognize pharmacological properties of multiple drugs across different biological systems and conditions. We also propose using deep neural net confusion matrices for drug repositioning. This work is a proof of principle for applying deep learning to drug discovery and development. PMID:27200455

Multimodal RNA-seq using single-strand, double-strand, and CircLigase-based capture yields a refined and extended description of the C. elegans transcriptome.

PubMed

Lamm, Ayelet T; Stadler, Michael R; Zhang, Huibin; Gent, Jonathan I; Fire, Andrew Z

2011-02-01

We have used a combination of three high-throughput RNA capture and sequencing methods to refine and augment the transcriptome map of a well-studied genetic model, Caenorhabditis elegans. The three methods include a standard (non-directional) library preparation protocol relying on cDNA priming and foldback that has been used in several previous studies for transcriptome characterization in this species, and two directional protocols, one involving direct capture of single-stranded RNA fragments and one involving circular-template PCR (CircLigase). We find that each RNA-seq approach shows specific limitations and biases, with the application of multiple methods providing a more complete map than was obtained from any single method. Of particular note in the analysis were substantial advantages of CircLigase-based and ssRNA-based capture for defining sequences and structures of the precise 5' ends (which were lost using the double-strand cDNA capture method). Of the three methods, ssRNA capture was most effective in defining sequences to the poly(A) junction. Using data sets from a spectrum of C. elegans strains and stages and the UCSC Genome Browser, we provide a series of tools, which facilitate rapid visualization and assignment of gene structures.

Transcriptomic insight into pathogenicity-associated factors of Conidiobolus obscurus, an obligate aphid-pathogenic fungus belonging to Entomopthoromycota.

PubMed

Wang, Jianghong; Zhou, Xiang; Guo, Kai; Zhang, Xinqi; Lin, Haiping; Montalva, Cristian

2018-01-16

Conidiobolus obscurus is a widespread fungal entomopathogen with aphid biocontrol potential. This study focused on a de novo transcriptomic analysis of C. obscurus. A number of pathogenicity-associated factors were annotated for the first time from the assembled 17 231 fungal unigenes, including those encoding subtilisin-like proteolytic enzymes (Pr1s), trypsin-like proteases, metalloproteases, carboxypeptidases and endochitinases. Many of these genes were transcriptionally up-regulated by at least twofold in mycotized cadavers compared with the in vitro fungal cultures. The resultant transcriptomic database was validated by the transcript levels of three selected pathogenicity-related genes quantified from different in vivo and in vitro material in real-time quantitative polymerase chain reaction (PCR). The involvement of multiple Pr1 proteases in the first stage of fungal infection was also suggested. Interestingly, a unique cytolytic (Cyt)-like δ-endotoxin gene was highly expressed in both mycotized cadavers and fungal cultures, and was more or less distinct from its homologues in bacteria and other fungi. Our findings provide the first global insight into various pathogenicity-related genes in this obligate aphid pathogen and may help to develop novel biocontrol strategy against aphid pests. © 2018 Society of Chemical Industry. © 2018 Society of Chemical Industry.

From cacti to carnivores: Improved phylotranscriptomic sampling and hierarchical homology inference provide further insight into the evolution of Caryophyllales.

PubMed

Walker, Joseph F; Yang, Ya; Feng, Tao; Timoneda, Alfonso; Mikenas, Jessica; Hutchison, Vera; Edwards, Caroline; Wang, Ning; Ahluwalia, Sonia; Olivieri, Julia; Walker-Hale, Nathanael; Majure, Lucas C; Puente, Raúl; Kadereit, Gudrun; Lauterbach, Maximilian; Eggli, Urs; Flores-Olvera, Hilda; Ochoterena, Helga; Brockington, Samuel F; Moore, Michael J; Smith, Stephen A

2018-03-01

The Caryophyllales contain ~12,500 species and are known for their cosmopolitan distribution, convergence of trait evolution, and extreme adaptations. Some relationships within the Caryophyllales, like those of many large plant clades, remain unclear, and phylogenetic studies often recover alternative hypotheses. We explore the utility of broad and dense transcriptome sampling across the order for resolving evolutionary relationships in Caryophyllales. We generated 84 transcriptomes and combined these with 224 publicly available transcriptomes to perform a phylogenomic analysis of Caryophyllales. To overcome the computational challenge of ortholog detection in such a large data set, we developed an approach for clustering gene families that allowed us to analyze >300 transcriptomes and genomes. We then inferred the species relationships using multiple methods and performed gene-tree conflict analyses. Our phylogenetic analyses resolved many clades with strong support, but also showed significant gene-tree discordance. This discordance is not only a common feature of phylogenomic studies, but also represents an opportunity to understand processes that have structured phylogenies. We also found taxon sampling influences species-tree inference, highlighting the importance of more focused studies with additional taxon sampling. Transcriptomes are useful both for species-tree inference and for uncovering evolutionary complexity within lineages. Through analyses of gene-tree conflict and multiple methods of species-tree inference, we demonstrate that phylogenomic data can provide unparalleled insight into the evolutionary history of Caryophyllales. We also discuss a method for overcoming computational challenges associated with homolog clustering in large data sets. © 2018 The Authors. American Journal of Botany is published by Wiley Periodicals, Inc. on behalf of the Botanical Society of America.

Genomics of Adaptation to Multiple Concurrent Stresses: Insights from Comparative Transcriptomics of a Cichlid Fish from One of Earth's Most Extreme Environments, the Hypersaline Soda Lake Magadi in Kenya, East Africa.

PubMed

Kavembe, Geraldine D; Franchini, Paolo; Irisarri, Iker; Machado-Schiaffino, Gonzalo; Meyer, Axel

2015-10-01

The Magadi tilapia (Alcolapia grahami) is a cichlid fish that inhabits one of the Earth's most extreme aquatic environments, with high pH (~10), salinity (~60% of seawater), high temperatures (~40 °C), and fluctuating oxygen regimes. The Magadi tilapia evolved several unique behavioral, physiological, and anatomical adaptations, some of which are constituent and thus retained in freshwater conditions. We conducted a transcriptomic analysis on A. grahami to study the evolutionary basis of tolerance to multiple stressors. To identify the adaptive regulatory changes associated with stress responses, we massively sequenced gill transcriptomes (RNAseq) from wild and freshwater-acclimated specimens of A. grahami. As a control, corresponding transcriptome data from Oreochromis leucostictus, a closely related freshwater species, were generated. We found expression differences in a large number of genes with known functions related to osmoregulation, energy metabolism, ion transport, and chemical detoxification. Over-representation of metabolism-related gene ontology terms in wild individuals compared to laboratory-acclimated specimens suggested that freshwater conditions greatly decrease the metabolic requirements of this species. Twenty-five genes with diverse physiological functions related to responses to water stress showed signs of divergent natural selection between the Magadi tilapia and its freshwater relative, which shared a most recent common ancestor only about four million years ago. The complete set of genes responsible for urea excretion was identified in the gill transcriptome of A. grahami, making it the only fish species to have a functional ornithine-urea cycle pathway in the gills--a major innovation for increasing nitrogenous waste efficiency.

Integrating Transcriptomic and Proteomic Data Using Predictive Regulatory Network Models of Host Response to Pathogens

PubMed Central

Chasman, Deborah; Walters, Kevin B.; Lopes, Tiago J. S.; Eisfeld, Amie J.; Kawaoka, Yoshihiro; Roy, Sushmita

2016-01-01

Mammalian host response to pathogenic infections is controlled by a complex regulatory network connecting regulatory proteins such as transcription factors and signaling proteins to target genes. An important challenge in infectious disease research is to understand molecular similarities and differences in mammalian host response to diverse sets of pathogens. Recently, systems biology studies have produced rich collections of omic profiles measuring host response to infectious agents such as influenza viruses at multiple levels. To gain a comprehensive understanding of the regulatory network driving host response to multiple infectious agents, we integrated host transcriptomes and proteomes using a network-based approach. Our approach combines expression-based regulatory network inference, structured-sparsity based regression, and network information flow to infer putative physical regulatory programs for expression modules. We applied our approach to identify regulatory networks, modules and subnetworks that drive host response to multiple influenza infections. The inferred regulatory network and modules are significantly enriched for known pathways of immune response and implicate apoptosis, splicing, and interferon signaling processes in the differential response of viral infections of different pathogenicities. We used the learned network to prioritize regulators and study virus and time-point specific networks. RNAi-based knockdown of predicted regulators had significant impact on viral replication and include several previously unknown regulators. Taken together, our integrated analysis identified novel module level patterns that capture strain and pathogenicity-specific patterns of expression and helped identify important regulators of host response to influenza infection. PMID:27403523

Revealing stable processing products from ribosome-associated small RNAs by deep-sequencing data analysis.

PubMed

Zywicki, Marek; Bakowska-Zywicka, Kamilla; Polacek, Norbert

2012-05-01

The exploration of the non-protein-coding RNA (ncRNA) transcriptome is currently focused on profiling of microRNA expression and detection of novel ncRNA transcription units. However, recent studies suggest that RNA processing can be a multi-layer process leading to the generation of ncRNAs of diverse functions from a single primary transcript. Up to date no methodology has been presented to distinguish stable functional RNA species from rapidly degraded side products of nucleases. Thus the correct assessment of widespread RNA processing events is one of the major obstacles in transcriptome research. Here, we present a novel automated computational pipeline, named APART, providing a complete workflow for the reliable detection of RNA processing products from next-generation-sequencing data. The major features include efficient handling of non-unique reads, detection of novel stable ncRNA transcripts and processing products and annotation of known transcripts based on multiple sources of information. To disclose the potential of APART, we have analyzed a cDNA library derived from small ribosome-associated RNAs in Saccharomyces cerevisiae. By employing the APART pipeline, we were able to detect and confirm by independent experimental methods multiple novel stable RNA molecules differentially processed from well known ncRNAs, like rRNAs, tRNAs or snoRNAs, in a stress-dependent manner.

Meta-transcriptomics indicates biotic cross-tolerance in willow trees cultivated on petroleum hydrocarbon contaminated soil.

PubMed

Gonzalez, Emmanuel; Brereton, Nicholas J B; Marleau, Julie; Guidi Nissim, Werther; Labrecque, Michel; Pitre, Frederic E; Joly, Simon

2015-10-12

High concentrations of petroleum hydrocarbon (PHC) pollution can be hazardous to human health and leave soils incapable of supporting agricultural crops. A cheap solution, which can help restore biodiversity and bring land back to productivity, is cultivation of high biomass yielding willow trees. However, the genetic mechanisms which allow these fast-growing trees to tolerate PHCs are as yet unclear. Salix purpurea 'Fish Creek' trees were pot-grown in soil from a former petroleum refinery, either lacking or enriched with C10-C50 PHCs. De novo assembled transcriptomes were compared between tree organs and impartially annotated without a priori constraint to any organism. Over 45% of differentially expressed genes originated from foreign organisms, the majority from the two-spotted spidermite, Tetranychus urticae. Over 99% of T. urticae transcripts were differentially expressed with greater abundance in non-contaminated trees. Plant transcripts involved in the polypropanoid pathway, including phenylalanine ammonia-lyase (PAL), had greater expression in contaminated trees whereas most resistance genes showed higher expression in non-contaminated trees. The impartial approach to annotation of the de novo transcriptomes, allowing for the possibility for multiple species identification, was essential for interpretation of the crop's response treatment. The meta-transcriptomic pattern of expression suggests a cross-tolerance mechanism whereby abiotic stress resistance systems provide improved biotic resistance. These findings highlight a valuable but complex biotic and abiotic stress response to real-world, multidimensional contamination which could, in part, help explain why crops such as willow can produce uniquely high biomass yields on challenging marginal land.

Insulin immuno-neutralization in fed chickens: effects on liver and muscle transcriptome.

PubMed

Simon, Jean; Milenkovic, Dragan; Godet, Estelle; Cabau, Cedric; Collin, Anne; Métayer-Coustard, Sonia; Rideau, Nicole; Tesseraud, Sophie; Derouet, Michel; Crochet, Sabine; Cailleau-Audouin, Estelle; Hennequet-Antier, Christelle; Gespach, Christian; Porter, Tom E; Duclos, Michel J; Dupont, Joëlle; Cogburn, Larry A

2012-03-01

Chickens mimic an insulin-resistance state by exhibiting several peculiarities with regard to plasma glucose level and its control by insulin. To gain insight into the role of insulin in the control of chicken transcriptome, liver and leg muscle transcriptomes were compared in fed controls and "diabetic" chickens, at 5 h after insulin immuno-neutralization, using 20.7K-chicken oligo-microarrays. At a level of false discovery rate <0.01, 1,573 and 1,225 signals were significantly modified by insulin privation in liver and muscle, respectively. Microarray data agreed reasonably well with qRT-PCR and some protein level measurements. Differentially expressed mRNAs with human ID were classified using Biorag analysis and Ingenuity Pathway Analysis. Multiple metabolic pathways, structural proteins, transporters and proteins of intracellular trafficking, major signaling pathways, and elements of the transcriptional control machinery were largely represented in both tissues. At least 42 mRNAs have already been associated with diabetes, insulin resistance, obesity, energy expenditure, or identified as sensors of metabolism in mice or humans. The contribution of the pathways presently identified to chicken physiology (particularly those not yet related to insulin) needs to be evaluated in future studies. Other challenges include the characterization of "unknown" mRNAs and the identification of the steps or networks, which disturbed tissue transcriptome so extensively, quickly after the turning off of the insulin signal. In conclusion, pleiotropic effects of insulin in chickens are further evidenced; major pathways controlled by insulin in mammals have been conserved despite the presence of unique features of insulin signaling in chicken muscle.

Analysis of the Transcriptomes Downstream of Eyeless and the Hedgehog, Decapentaplegic and Notch Signaling Pathways in Drosophila melanogaster

PubMed Central

Nfonsam, Landry E.; Cano, Carlos; Mudge, Joann; Schilkey, Faye D.; Curtiss, Jennifer

2012-01-01

Tissue-specific transcription factors are thought to cooperate with signaling pathways to promote patterned tissue specification, in part by co-regulating transcription. The Drosophila melanogaster Pax6 homolog Eyeless forms a complex, incompletely understood regulatory network with the Hedgehog, Decapentaplegic and Notch signaling pathways to control eye-specific gene expression. We report a combinatorial approach, including mRNAseq and microarray analyses, to identify targets co-regulated by Eyeless and Hedgehog, Decapentaplegic or Notch. Multiple analyses suggest that the transcriptomes resulting from co-misexpression of Eyeless+signaling factors provide a more complete picture of eye development compared to previous efforts involving Eyeless alone: (1) Principal components analysis and two-way hierarchical clustering revealed that the Eyeless+signaling factor transcriptomes are closer to the eye control transcriptome than when Eyeless is misexpressed alone; (2) more genes are upregulated at least three-fold in response to Eyeless+signaling factors compared to Eyeless alone; (3) based on gene ontology analysis, the genes upregulated in response to Eyeless+signaling factors had a greater diversity of functions compared to Eyeless alone. Through a secondary screen that utilized RNA interference, we show that the predicted gene CG4721 has a role in eye development. CG4721 encodes a neprilysin family metalloprotease that is highly up-regulated in response to Eyeless+Notch, confirming the validity of our approach. Given the similarity between D. melanogaster and vertebrate eye development, the large number of novel genes identified as potential targets of Ey+signaling factors will provide novel insights to our understanding of eye development in D. melanogaster and humans. PMID:22952997

Integrating multiple fitting regression and Bayes decision for cancer diagnosis with transcriptomic data from tumor-educated blood platelets.

PubMed

Huang, Guangzao; Yuan, Mingshun; Chen, Moliang; Li, Lei; You, Wenjie; Li, Hanjie; Cai, James J; Ji, Guoli

2017-10-07

The application of machine learning in cancer diagnostics has shown great promise and is of importance in clinic settings. Here we consider applying machine learning methods to transcriptomic data derived from tumor-educated platelets (TEPs) from individuals with different types of cancer. We aim to define a reliability measure for diagnostic purposes to increase the potential for facilitating personalized treatments. To this end, we present a novel classification method called MFRB (for Multiple Fitting Regression and Bayes decision), which integrates the process of multiple fitting regression (MFR) with Bayes decision theory. MFR is first used to map multidimensional features of the transcriptomic data into a one-dimensional feature. The probability density function of each class in the mapped space is then adjusted using the Gaussian probability density function. Finally, the Bayes decision theory is used to build a probabilistic classifier with the estimated probability density functions. The output of MFRB can be used to determine which class a sample belongs to, as well as to assign a reliability measure for a given class. The classical support vector machine (SVM) and probabilistic SVM (PSVM) are used to evaluate the performance of the proposed method with simulated and real TEP datasets. Our results indicate that the proposed MFRB method achieves the best performance compared to SVM and PSVM, mainly due to its strong generalization ability for limited, imbalanced, and noisy data.

Rapid transcriptome sequencing of an invasive pest, the brown marmorated stink bug Halyomorpha halys.

PubMed

Ioannidis, Panagiotis; Lu, Yong; Kumar, Nikhil; Creasy, Todd; Daugherty, Sean; Chibucos, Marcus C; Orvis, Joshua; Shetty, Amol; Ott, Sandra; Flowers, Melissa; Sengamalay, Naomi; Tallon, Luke J; Pick, Leslie; Dunning Hotopp, Julie C

2014-08-29

Halyomorpha halys (Stål) (Insecta:Hemiptera;Pentatomidae), commonly known as the Brown Marmorated Stink Bug (BMSB), is an invasive pest of the mid-Atlantic region of the United States, causing economically important damage to a wide range of crops. Native to Asia, BMSB was first observed in Allentown, PA, USA, in 1996, and this pest is now well-established throughout the US mid-Atlantic region and beyond. In addition to the serious threat BMSB poses to agriculture, BMSB has become a nuisance to homeowners, invading home gardens and congregating in large numbers in human-made structures, including homes, to overwinter. Despite its significance as an agricultural pest with limited control options, only 100 bp of BMSB sequence data was available in public databases when this project began. Transcriptome sequencing was undertaken to provide a molecular resource to the research community to inform the development of pest control strategies and to provide molecular data for population genetics studies of BMSB. Using normalized, strand-specific libraries, we sequenced pools of all BMSB life stages on the Illumina HiSeq. Trinity was used to assemble 200,000 putative transcripts in >100,000 components. A novel bioinformatic method that analyzed the strand-specificity of the data reduced this to 53,071 putative transcripts from 18,573 components. By integrating multiple other data types, we narrowed this further to 13,211 representative transcripts. Bacterial endosymbiont genes were identified in this dataset, some of which have a copy number consistent with being lateral gene transfers between endosymbiont genomes and Hemiptera, including ankyrin-repeat related proteins, lysozyme, and mannanase. Such genes and endosymbionts may provide novel targets for BMSB-specific biocontrol. This study demonstrates the utility of strand-specific sequencing in generating shotgun transcriptomes and that rapid sequencing shotgun transcriptomes is possible without the need for extensive inbreeding to generate homozygous lines. Such sequencing can provide a rapid response to pest invasions similar to that already described for disease epidemiology.

Saccular Transcriptome Profiles of the Seasonal Breeding Plainfin Midshipman Fish (Porichthys notatus), a Teleost with Divergent Sexual Phenotypes.

PubMed

Faber-Hammond, Joshua; Samanta, Manoj P; Whitchurch, Elizabeth A; Manning, Dustin; Sisneros, Joseph A; Coffin, Allison B

2015-01-01

Acoustic communication is essential for the reproductive success of the plainfin midshipman fish (Porichthys notatus). During the breeding season, type I males use acoustic cues to advertise nest location to potential mates, creating an audible signal that attracts reproductive females. Type II (sneaker) males also likely use this social acoustic signal to find breeding pairs from which to steal fertilizations. Estrogen-induced changes in the auditory system of breeding females are thought to enhance neural encoding of the advertisement call, and recent anatomical data suggest the saccule (the main auditory end organ) as one possible target for this seasonal modulation. Here we describe saccular transcriptomes from all three sexual phenotypes (females, type I and II males) collected during the breeding season as a first step in understanding the mechanisms underlying sexual phenotype-specific and seasonal differences in auditory function. We used RNA-Seq on the Ion Torrent platform to create a combined transcriptome dataset containing over 79,000 assembled transcripts representing almost 9,000 unique annotated genes. These identified genes include several with known inner ear function and multiple steroid hormone receptors. Transcripts most closely matched to published genomes of nile tilapia and large yellow croaker, inconsistent with the phylogenetic relationship between these species but consistent with the importance of acoustic communication in their life-history strategies. We then compared the RNA-Seq results from the saccules of reproductive females with a separate transcriptome from the non-reproductive female phenotype and found over 700 differentially expressed transcripts, including members of the Wnt and Notch signaling pathways that mediate cell proliferation and hair cell addition in the inner ear. These data constitute a valuable resource for furthering our understanding of the molecular basis for peripheral auditory function as well as a range of future midshipman and cross-species comparative studies of the auditory periphery.

De novo Assembly of the Burying Beetle Nicrophorus orbicollis (Coleoptera: Silphidae) Transcriptome Across Developmental Stages with Identification of Key Immune Transcripts

PubMed Central

Won, Harim I.; Schulze, Thomas T.; Clement, Emalie J.; Watson, Gabrielle F.; Watson, Sean M.; Warner, Rosalie C.; Ramler, Elizabeth A. M.; Witte, Elias J.; Schoenbeck, Mark A.; Rauter, Claudia M.; Davis, Paul H.

2018-01-01

Burying beetles (Nicrophorus spp.) are among the relatively few insects that provide parental care while not belonging to the eusocial insects such as ants or bees. This behavior incurs energy costs as evidenced by immune deficits and shorter life-spans in reproducing beetles. In the absence of an assembled transcriptome, relatively little is known concerning the molecular biology of these beetles. This work details the assembly and analysis of the Nicrophorus orbicollis transcriptome at multiple developmental stages. RNA-Seq reads were obtained by next-generation sequencing and the transcriptome was assembled using the Trinity assembler. Validation of the assembly was performed by functional characterization using Gene Ontology (GO), Eukaryotic Orthologous Groups (KOG), and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. Differential expression analysis highlights developmental stage-specific expression patterns, and immunity-related transcripts are discussed. The data presented provides a valuable molecular resource to aid further investigation into immunocompetence throughout this organism's sexual development. PMID:29707046

Multiple genes contribute to anhydrobiosis (tolerance to extreme desiccation) in the nematode Panagrolaimus superbus

PubMed Central

Evangelista, Cláudia Carolina Silva; Guidelli, Giovanna Vieira; Borges, Gustavo; Araujo, Thais Fenz; de Souza, Tiago Alves Jorge; Neves, Ubiraci Pereira da Costa; Tunnacliffe, Alan; Pereira, Tiago Campos

2017-01-01

Abstract The molecular basis of anhydrobiosis, the state of suspended animation entered by some species during extreme desiccation, is still poorly understood despite a number of transcriptome and proteome studies. We therefore conducted functional screening by RNA interference (RNAi) for genes involved in anhydrobiosis in the holo-anhydrobiotic nematode Panagrolaimus superbus. A new method of survival analysis, based on staining, and proof-of-principle RNAi experiments confirmed a role for genes involved in oxidative stress tolerance, while a novel medium-scale RNAi workflow identified a further 40 anhydrobiosis-associated genes, including several involved in proteostasis, DNA repair and signal transduction pathways. This suggests that multiple genes contribute to anhydrobiosis in P. superbus. PMID:29111563

Systemic lupus erythematosus diagnostics in the ‘omics’ era

PubMed Central

Arriens, Cristina; Mohan, Chandra

2014-01-01

Systemic lupus erythematosus is a complex autoimmune disease affecting multiple organ systems. Currently, diagnosis relies upon meeting at least four out of eleven criteria outlined by the ACR. The scientific community actively pursues discovery of novel diagnostics in the hope of better identifying susceptible individuals in early stages of disease. Comprehensive studies have been conducted at multiple biological levels including: DNA (or genomics), mRNA (or transcriptomics), protein (or proteomics) and metabolites (or metabolomics). The ‘omics’ platforms allow us to re-examine systemic lupus erythematosus at a greater degree of molecular resolution. More importantly, one is hopeful that these ‘omics’ platforms may yield newer biomarkers for systemic lupus erythematosus that can help clinicians track the disease course with greater sensitivity and specificity. PMID:24860621

Developing single nucleotide polymorphism (SNP) markers from transcriptome sequences for identification of longan (Dimocarpus longan) germplasm

PubMed Central

Wang, Boyi; Tan, Hua-Wei; Fang, Wanping; Meinhardt, Lyndel W; Mischke, Sue; Matsumoto, Tracie; Zhang, Dapeng

2015-01-01

Longan (Dimocarpus longan Lour.) is an important tropical fruit tree crop. Accurate varietal identification is essential for germplasm management and breeding. Using longan transcriptome sequences from public databases, we developed single nucleotide polymorphism (SNP) markers; validated 60 SNPs in 50 longan germplasm accessions, including cultivated varieties and wild germplasm; and designated 25 SNP markers that unambiguously identified all tested longan varieties with high statistical rigor (P<0.0001). Multiple trees from the same clone were verified and off-type trees were identified. Diversity analysis revealed genetic relationships among analyzed accessions. Cultivated varieties differed significantly from wild populations (Fst=0.300; P<0.001), demonstrating untapped genetic diversity for germplasm conservation and utilization. Within cultivated varieties, apparent differences between varieties from China and those from Thailand and Hawaii indicated geographic patterns of genetic differentiation. These SNP markers provide a powerful tool to manage longan genetic resources and breeding, with accurate and efficient genotype identification. PMID:26504559

Joint-specific DNA methylation and transcriptome signatures in rheumatoid arthritis identify distinct pathogenic processes

PubMed Central

Ai, Rizi; Hammaker, Deepa; Boyle, David L.; Morgan, Rachel; Walsh, Alice M.; Fan, Shicai; Firestein, Gary S.; Wang, Wei

2016-01-01

Stratifying patients on the basis of molecular signatures could facilitate development of therapeutics that target pathways specific to a particular disease or tissue location. Previous studies suggest that pathogenesis of rheumatoid arthritis (RA) is similar in all affected joints. Here we show that distinct DNA methylation and transcriptome signatures not only discriminate RA fibroblast-like synoviocytes (FLS) from osteoarthritis FLS, but also distinguish RA FLS isolated from knees and hips. Using genome-wide methods, we show differences between RA knee and hip FLS in the methylation of genes encoding biological pathways, such as IL-6 signalling via JAK-STAT pathway. Furthermore, differentially expressed genes are identified between knee and hip FLS using RNA-sequencing. Double-evidenced genes that are both differentially methylated and expressed include multiple HOX genes. Joint-specific DNA signatures suggest that RA disease mechanisms might vary from joint to joint, thus potentially explaining some of the diversity of drug responses in RA patients. PMID:27282753

Global Analysis of Transcriptome Responses and Gene Expression Profiles to Cold Stress of Jatropha curcas L.

PubMed Central

Wang, Haibo; Zou, Zhurong; Wang, Shasha; Gong, Ming

2013-01-01

Background Jatropha curcas L., also called the Physic nut, is an oil-rich shrub with multiple uses, including biodiesel production, and is currently exploited as a renewable energy resource in many countries. Nevertheless, because of its origin from the tropical MidAmerican zone, J. curcas confers an inherent but undesirable characteristic (low cold resistance) that may seriously restrict its large-scale popularization. This adaptive flaw can be genetically improved by elucidating the mechanisms underlying plant tolerance to cold temperatures. The newly developed Illumina Hiseq™ 2000 RNA-seq and Digital Gene Expression (DGE) are deep high-throughput approaches for gene expression analysis at the transcriptome level, using which we carefully investigated the gene expression profiles in response to cold stress to gain insight into the molecular mechanisms of cold response in J. curcas. Results In total, 45,251 unigenes were obtained by assembly of clean data generated by RNA-seq analysis of the J. curcas transcriptome. A total of 33,363 and 912 complete or partial coding sequences (CDSs) were determined by protein database alignments and ESTScan prediction, respectively. Among these unigenes, more than 41.52% were involved in approximately 128 known metabolic or signaling pathways, and 4,185 were possibly associated with cold resistance. DGE analysis was used to assess the changes in gene expression when exposed to cold condition (12°C) for 12, 24, and 48 h. The results showed that 3,178 genes were significantly upregulated and 1,244 were downregulated under cold stress. These genes were then functionally annotated based on the transcriptome data from RNA-seq analysis. Conclusions This study provides a global view of transcriptome response and gene expression profiling of J. curcas in response to cold stress. The results can help improve our current understanding of the mechanisms underlying plant cold resistance and favor the screening of crucial genes for genetically enhancing cold resistance in J. curcas. PMID:24349370

Global analysis of transcriptome responses and gene expression profiles to cold stress of Jatropha curcas L.

PubMed

Wang, Haibo; Zou, Zhurong; Wang, Shasha; Gong, Ming

2013-01-01

Jatropha curcas L., also called the Physic nut, is an oil-rich shrub with multiple uses, including biodiesel production, and is currently exploited as a renewable energy resource in many countries. Nevertheless, because of its origin from the tropical MidAmerican zone, J. curcas confers an inherent but undesirable characteristic (low cold resistance) that may seriously restrict its large-scale popularization. This adaptive flaw can be genetically improved by elucidating the mechanisms underlying plant tolerance to cold temperatures. The newly developed Illumina Hiseq™ 2000 RNA-seq and Digital Gene Expression (DGE) are deep high-throughput approaches for gene expression analysis at the transcriptome level, using which we carefully investigated the gene expression profiles in response to cold stress to gain insight into the molecular mechanisms of cold response in J. curcas. In total, 45,251 unigenes were obtained by assembly of clean data generated by RNA-seq analysis of the J. curcas transcriptome. A total of 33,363 and 912 complete or partial coding sequences (CDSs) were determined by protein database alignments and ESTScan prediction, respectively. Among these unigenes, more than 41.52% were involved in approximately 128 known metabolic or signaling pathways, and 4,185 were possibly associated with cold resistance. DGE analysis was used to assess the changes in gene expression when exposed to cold condition (12°C) for 12, 24, and 48 h. The results showed that 3,178 genes were significantly upregulated and 1,244 were downregulated under cold stress. These genes were then functionally annotated based on the transcriptome data from RNA-seq analysis. This study provides a global view of transcriptome response and gene expression profiling of J. curcas in response to cold stress. The results can help improve our current understanding of the mechanisms underlying plant cold resistance and favor the screening of crucial genes for genetically enhancing cold resistance in J. curcas.

Roel Verhaak, Ph.D., Presents the Somatic Genomic Landscape of Glioblastoma - TCGA

Cancer.gov

Diffuse lower grade gliomas (LGGs) are infiltrative neoplasms of the central nervous system that include astrocytoma, oligodendroglioma and oligo-astrocytoma histologies of grades II and III. Roel G.W. Verhaak, Ph.D., presents a comprehensive analysis of 293 LGGs using multiple advanced genomic, transcriptomic and proteomic platforms from The Cancer Genome Atlas to provide a deeper understanding of the molecular features of this group of neoplasms, to classify them in a clinically-relevant manner, and to provide a public resource that identifies potential targets for emerging therapies.

Improving amphibian genomic resources: a multitissue reference transcriptome of an iconic invader.

PubMed

Richardson, Mark F; Sequeira, Fernando; Selechnik, Daniel; Carneiro, Miguel; Vallinoto, Marcelo; Reid, Jack G; West, Andrea J; Crossland, Michael R; Shine, Richard; Rollins, Lee A

2018-01-01

Cane toads (Rhinella marina) are an iconic invasive species introduced to 4 continents and well utilized for studies of rapid evolution in introduced environments. Despite the long introduction history of this species, its profound ecological impacts, and its utility for demonstrating evolutionary principles, genetic information is sparse. Here we produce a de novo transcriptome spanning multiple tissues and life stages to enable investigation of the genetic basis of previously identified rapid phenotypic change over the introduced range. Using approximately 1.9 billion reads from developing tadpoles and 6 adult tissue-specific cDNA libraries, as well as a transcriptome assembly pipeline encompassing 100 separate de novo assemblies, we constructed 62 202 transcripts, of which we functionally annotated ∼50%. Our transcriptome assembly exhibits 90% full-length completeness of the Benchmarking Universal Single-Copy Orthologs data set. Robust assembly metrics and comparisons with several available anuran transcriptomes and genomes indicate that our cane toad assembly is one of the most complete anuran genomic resources available. This comprehensive anuran transcriptome will provide a valuable resource for investigation of genes under selection during invasion in cane toads, but will also greatly expand our general knowledge of anuran genomes, which are underrepresented in the literature. The data set is publically available in NCBI and GigaDB to serve as a resource for other researchers. © The Authors 2017. Published by Oxford University Press.

Improving amphibian genomic resources: a multitissue reference transcriptome of an iconic invader

PubMed Central

Reid, Jack G; Crossland, Michael R

2018-01-01

Abstract Background Cane toads (Rhinella marina) are an iconic invasive species introduced to 4 continents and well utilized for studies of rapid evolution in introduced environments. Despite the long introduction history of this species, its profound ecological impacts, and its utility for demonstrating evolutionary principles, genetic information is sparse. Here we produce a de novo transcriptome spanning multiple tissues and life stages to enable investigation of the genetic basis of previously identified rapid phenotypic change over the introduced range. Findings Using approximately 1.9 billion reads from developing tadpoles and 6 adult tissue-specific cDNA libraries, as well as a transcriptome assembly pipeline encompassing 100 separate de novo assemblies, we constructed 62 202 transcripts, of which we functionally annotated ∼50%. Our transcriptome assembly exhibits 90% full-length completeness of the Benchmarking Universal Single-Copy Orthologs data set. Robust assembly metrics and comparisons with several available anuran transcriptomes and genomes indicate that our cane toad assembly is one of the most complete anuran genomic resources available. Conclusions This comprehensive anuran transcriptome will provide a valuable resource for investigation of genes under selection during invasion in cane toads, but will also greatly expand our general knowledge of anuran genomes, which are underrepresented in the literature. The data set is publically available in NCBI and GigaDB to serve as a resource for other researchers. PMID:29186423

Transcriptomes of Eight Arabidopsis thaliana Accessions Reveal Core Conserved, Genotype- and Organ-Specific Responses to Flooding Stress1[OPEN

PubMed Central

van Veen, Hans; Vashisht, Divya; Akman, Melis; Girke, Thomas; Mustroph, Angelika; Reinen, Emilie; Kooiker, Maarten; van Tienderen, Peter; Voesenek, Laurentius A.C.J.

2016-01-01

Climate change has increased the frequency and severity of flooding events, with significant negative impact on agricultural productivity. These events often submerge plant aerial organs and roots, limiting growth and survival due to a severe reduction in light reactions and gas exchange necessary for photosynthesis and respiration, respectively. To distinguish molecular responses to the compound stress imposed by submergence, we investigated transcriptomic adjustments to darkness in air and under submerged conditions using eight Arabidopsis (Arabidopsis thaliana) accessions differing significantly in sensitivity to submergence. Evaluation of root and rosette transcriptomes revealed an early transcriptional and posttranscriptional response signature that was conserved primarily across genotypes, although flooding susceptibility-associated and genotype-specific responses also were uncovered. Posttranscriptional regulation encompassed darkness- and submergence-induced alternative splicing of transcripts from pathways involved in the alternative mobilization of energy reserves. The organ-specific transcriptome adjustments reflected the distinct physiological status of roots and shoots. Root-specific transcriptome changes included marked up-regulation of chloroplast-encoded photosynthesis and redox-related genes, whereas those of the rosette were related to the regulation of development and growth processes. We identified a novel set of tolerance genes, recognized mainly by quantitative differences. These included a transcriptome signature of more pronounced gluconeogenesis in tolerant accessions, a response that included stress-induced alternative splicing. This study provides organ-specific molecular resolution of genetic variation in submergence responses involving interactions between darkness and low-oxygen constraints of flooding stress and demonstrates that early transcriptome plasticity, including alternative splicing, is associated with the ability to cope with a compound environmental stress. PMID:27208254

Unprecedented high-resolution view of bacterial operon architecture revealed by RNA sequencing.

PubMed

Conway, Tyrrell; Creecy, James P; Maddox, Scott M; Grissom, Joe E; Conkle, Trevor L; Shadid, Tyler M; Teramoto, Jun; San Miguel, Phillip; Shimada, Tomohiro; Ishihama, Akira; Mori, Hirotada; Wanner, Barry L

2014-07-08

We analyzed the transcriptome of Escherichia coli K-12 by strand-specific RNA sequencing at single-nucleotide resolution during steady-state (logarithmic-phase) growth and upon entry into stationary phase in glucose minimal medium. To generate high-resolution transcriptome maps, we developed an organizational schema which showed that in practice only three features are required to define operon architecture: the promoter, terminator, and deep RNA sequence read coverage. We precisely annotated 2,122 promoters and 1,774 terminators, defining 1,510 operons with an average of 1.98 genes per operon. Our analyses revealed an unprecedented view of E. coli operon architecture. A large proportion (36%) of operons are complex with internal promoters or terminators that generate multiple transcription units. For 43% of operons, we observed differential expression of polycistronic genes, despite being in the same operons, indicating that E. coli operon architecture allows fine-tuning of gene expression. We found that 276 of 370 convergent operons terminate inefficiently, generating complementary 3' transcript ends which overlap on average by 286 nucleotides, and 136 of 388 divergent operons have promoters arranged such that their 5' ends overlap on average by 168 nucleotides. We found 89 antisense transcripts of 397-nucleotide average length, 7 unannotated transcripts within intergenic regions, and 18 sense transcripts that completely overlap operons on the opposite strand. Of 519 overlapping transcripts, 75% correspond to sequences that are highly conserved in E. coli (>50 genomes). Our data extend recent studies showing unexpected transcriptome complexity in several bacteria and suggest that antisense RNA regulation is widespread. Importance: We precisely mapped the 5' and 3' ends of RNA transcripts across the E. coli K-12 genome by using a single-nucleotide analytical approach. Our resulting high-resolution transcriptome maps show that ca. one-third of E. coli operons are complex, with internal promoters and terminators generating multiple transcription units and allowing differential gene expression within these operons. We discovered extensive antisense transcription that results from more than 500 operons, which fully overlap or extensively overlap adjacent divergent or convergent operons. The genomic regions corresponding to these antisense transcripts are highly conserved in E. coli (including Shigella species), although it remains to be proven whether or not they are functional. Our observations of features unearthed by single-nucleotide transcriptome mapping suggest that deeper layers of transcriptional regulation in bacteria are likely to be revealed in the future. Copyright © 2014 Conway et al.

Functional similarity and molecular divergence of a novel reproductive transcriptome in two male-pregnant Syngnathus pipefish species

PubMed Central

Small, Clayton M; Harlin-Cognato, April D; Jones, Adam G

2013-01-01

Evolutionary studies have revealed that reproductive proteins in animals and plants often evolve more rapidly than the genome-wide average. The causes of this pattern, which may include relaxed purifying selection, sexual selection, sexual conflict, pathogen resistance, reinforcement, or gene duplication, remain elusive. Investigative expansions to additional taxa and reproductive tissues have the potential to shed new light on this unresolved problem. Here, we embark on such an expansion, in a comparison of the brood-pouch transcriptome between two male-pregnant species of the pipefish genus Syngnathus. Male brooding tissues in syngnathid fishes represent a novel, nonurogenital reproductive trait, heretofore mostly uncharacterized from a molecular perspective. We leveraged next-generation sequencing (Roche 454 pyrosequencing) to compare transcript abundance in the male brooding tissues of pregnant with nonpregnant samples from Gulf (S. scovelli) and dusky (S. floridae) pipefish. A core set of protein-coding genes, including multiple members of astacin metalloprotease and c-type lectin gene families, is consistent between species in both the direction and magnitude of expression bias. As predicted, coding DNA sequence analysis of these putative “male pregnancy proteins” suggests rapid evolution relative to nondifferentially expressed genes and reflects signatures of adaptation similar in magnitude to those reported from Drosophila male accessory gland proteins. Although the precise drivers of male pregnancy protein divergence remain unknown, we argue that the male pregnancy transcriptome in syngnathid fishes, a clade diverse with respect to brooding morphology and mating system, represents a unique and promising object of study for understanding the perplexing evolutionary nature of reproductive molecules. PMID:24324861

Transcriptome and proteomic analyses reveal multiple differences associated with chloroplast development in the spaceflight-induced wheat albino mutant mta.

PubMed

Shi, Kui; Gu, Jiayu; Guo, Huijun; Zhao, Linshu; Xie, Yongdun; Xiong, Hongchun; Li, Junhui; Zhao, Shirong; Song, Xiyun; Liu, Luxiang

2017-01-01

Chloroplast development is an integral part of plant survival and growth, and occurs in parallel with chlorophyll biosynthesis. However, little is known about the mechanisms underlying chloroplast development in hexaploid wheat. Here, we obtained a spaceflight-induced wheat albino mutant mta. Chloroplast ultra-structural observation showed that chloroplasts of mta exhibit abnormal morphology and distribution compared to wild type. Photosynthetic pigments content was also significantly decreased in mta. Transcriptome and chloroplast proteome profiling of mta and wild type were done to identify differentially expressed genes (DEGs) and proteins (DEPs), respectively. In total 4,588 DEGs including 1,980 up- and 2,608 down-regulated, and 48 chloroplast DEPs including 15 up- and 33 down-regulated were identified in mta. Classification of DEGs revealed that most were involved in chloroplast development, chlorophyll biosynthesis, or photosynthesis. Besides, transcription factors such as PIF3, GLK and MYB which might participate in those pathways were also identified. The correlation analysis between DEGs and DEPs revealed that the transcript-to-protein in abundance was functioned into photosynthesis and chloroplast relevant groups. Real time qPCR analysis validated that the expression level of genes encoding photosynthetic proteins was significantly decreased in mta. Together, our results suggest that the molecular mechanism for albino leaf color formation in mta is a thoroughly regulated and complicated process. The combined analysis of transcriptome and proteome afford comprehensive information for further research on chloroplast development mechanism in wheat. And spaceflight provides a potential means for mutagenesis in crop breeding.

Transcriptomic analysis illuminates genes involved in chlorophyll synthesis after nitrogen starvation in Acaryochloris sp. CCMEE 5410.

PubMed

Yoneda, Aki; Wittmann, Bruce J; King, Jeremy D; Blankenship, Robert E; Dantas, Gautam

2016-08-01

Acaryochloris species are a genus of cyanobacteria that utilize chlorophyll (chl) d as their primary chlorophyll molecule during oxygenic photosynthesis. Chl d allows Acaryochloris to harvest red-shifted light, which gives them the ability to live in filtered light environments that are depleted in visible light. Although genomes of multiple Acaryochloris species have been sequenced, their analysis has not revealed how chl d is synthesized. Here, we demonstrate that Acaryochloris sp. CCMEE 5410 cells undergo chlorosis by nitrogen depletion and exhibit robust regeneration of chl d by nitrogen repletion. We performed a time course RNA-Seq experiment to quantify global transcriptomic changes during chlorophyll recovery. We observed upregulation of numerous known chl biosynthesis genes and also identified an oxygenase gene with a similar transcriptional profile as these chl biosynthesis genes, suggesting its possible involvement in chl d biosynthesis. Moreover, our data suggest that multiple prochlorophyte chlorophyll-binding homologs are important during chlorophyll recovery, and light-independent chl synthesis genes are more dominant than the light-dependent gene at the transcription level. Transcriptomic characterization of this organism provides crucial clues toward mechanistic elucidation of chl d biosynthesis.

Network Analysis of Rodent Transcriptomes in Spaceflight

NASA Technical Reports Server (NTRS)

Ramachandran, Maya; Fogle, Homer; Costes, Sylvain

2017-01-01

Network analysis methods leverage prior knowledge of cellular systems and the statistical and conceptual relationships between analyte measurements to determine gene connectivity. Correlation and conditional metrics are used to infer a network topology and provide a systems-level context for cellular responses. Integration across multiple experimental conditions and omics domains can reveal the regulatory mechanisms that underlie gene expression. GeneLab has assembled rich multi-omic (transcriptomics, proteomics, epigenomics, and epitranscriptomics) datasets for multiple murine tissues from the Rodent Research 1 (RR-1) experiment. RR-1 assesses the impact of 37 days of spaceflight on gene expression across a variety of tissue types, such as adrenal glands, quadriceps, gastrocnemius, tibalius anterior, extensor digitorum longus, soleus, eye, and kidney. Network analysis is particularly useful for RR-1 -omics datasets because it reinforces subtle relationships that may be overlooked in isolated analyses and subdues confounding factors. Our objective is to use network analysis to determine potential target nodes for therapeutic intervention and identify similarities with existing disease models. Multiple network algorithms are used for a higher confidence consensus.

Global Control of GacA in Secondary Metabolism, Primary Metabolism, Secretion Systems, and Motility in the Rhizobacterium Pseudomonas aeruginosa M18

PubMed Central

Wei, Xue; Tang, Lulu; Wu, Daqiang

2013-01-01

The rhizobacterium Pseudomonas aeruginosa M18 can produce a broad spectrum of secondary metabolites, including the antibiotics pyoluteorin (Plt) and phenazine-1-carboxylic acid (PCA), hydrogen cyanide, and the siderophores pyoverdine and pyochelin. The antibiotic biosynthesis of M18 is coordinately controlled by multiple distinct regulatory pathways, of which the GacS/GacA system activates Plt biosynthesis but strongly downregulates PCA biosynthesis. Here, we investigated the global influence of a gacA mutation on the M18 transcriptome and related metabolic and physiological processes. Transcriptome profiling revealed that the transcript levels of 839 genes, which account for approximately 15% of the annotated genes in the M18 genome, were significantly influenced by the gacA mutation during the early stationary growth phase of M18. Most secondary metabolic gene clusters, such as pvd, pch, plt, amb, and hcn, were activated by GacA. The GacA regulon also included genes encoding extracellular enzymes and cytochrome oxidases. Interestingly, the primary metabolism involved in the assimilation and metabolism of phosphorus, sulfur, and nitrogen sources was also notably regulated by GacA. Another important category of the GacA regulon was secretion systems, including H1, H2, and H3 (type VI secretion systems [T6SSs]), Hxc (T2SS), and Has and Apr (T1SSs), and CupE and Tad pili. More remarkably, GacA inhibited swimming, swarming, and twitching motilities. Taken together, the Gac-initiated global regulation, which was mostly mediated through multiple regulatory systems or factors, was mainly involved in secondary and primary metabolism, secretion systems, motility, etc., contributing to ecological or nutritional competence, ion homeostasis, and biocontrol in M18. PMID:23708134

Roles of the Sodium-Translocating NADH:Quinone Oxidoreductase (Na+-NQR) on Vibrio cholerae Metabolism, Motility and Osmotic Stress Resistance

PubMed Central

Minato, Yusuke; Halang, Petra; Quinn, Matthew J.; Faulkner, Wyatt J.; Aagesen, Alisha M.; Steuber, Julia; Stevens, Jan F.; Häse, Claudia C.

2014-01-01

The Na+ translocating NADH:quinone oxidoreductase (Na+-NQR) is a unique respiratory enzyme catalyzing the electron transfer from NADH to quinone coupled with the translocation of sodium ions across the membrane. Typically, Vibrio spp., including Vibrio cholerae, have this enzyme but lack the proton-pumping NADH:ubiquinone oxidoreductase (Complex I). Thus, Na+-NQR should significantly contribute to multiple aspects of V. cholerae physiology; however, no detailed characterization of this aspect has been reported so far. In this study, we broadly investigated the effects of loss of Na+-NQR on V. cholerae physiology by using Phenotype Microarray (Biolog), transcriptome and metabolomics analyses. We found that the V. cholerae ΔnqrA-F mutant showed multiple defects in metabolism detected by Phenotype Microarray. Transcriptome analysis revealed that the V. cholerae ΔnqrA-F mutant up-regulates 31 genes and down-regulates 55 genes in both early and mid-growth phases. The most up-regulated genes included the cadA and cadB genes, encoding a lysine decarboxylase and a lysine/cadaverine antiporter, respectively. Increased CadAB activity was further suggested by the metabolomics analysis. The down-regulated genes include sialic acid catabolism genes. Metabolomic analysis also suggested increased reductive pathway of TCA cycle and decreased purine metabolism in the V. cholerae ΔnqrA-F mutant. Lack of Na+-NQR did not affect any of the Na+ pumping-related phenotypes of V. cholerae suggesting that other secondary Na+ pump(s) can compensate for Na+ pumping activity of Na+-NQR. Overall, our study provides important insights into the contribution of Na+-NQR to V. cholerae physiology. PMID:24811312

AMPKα Modulation in Cancer Progression: Multilayer Integrative Analysis of the Whole Transcriptome in Asian Gastric Cancer

PubMed Central

Cho, Jae Yong; Cheong, Jae-Ho; Kim, Hoguen; Li, Min; Downey, Thomas J.; Dyer, Matthew D.; Sun, Yongming; Sun, Jingtao; Beasley, Ellen M.; Chung, Hyun Cheol; Noh, Sung Hoon; Weinstein, John N.; Liu, Chang-Gong; Powis, Garth

2013-01-01

Gastric cancer is the most common cancer in Asia and most developing countries. Despite the use of multimodality therapeutics, it remains the second leading cause of cancer death in the world. To identify the molecular underpinnings of gastric cancer in the Asian population, we applied an RNA-sequencing approach to gastric tumor and noncancerous specimens, generating 680 million informative short reads to quantitatively characterize the entire transcriptome of gastric cancer (including mRNAs and microRNAs). A multi-layer analysis was then developed to identify multiple types of transcriptional aberrations associated with different stages of gastric cancer, including differentially expressed mRNAs, recurrent somatic mutations and key differentially expressed microRNAs. Through this approach, we identified the central metabolic regulator AMPK-α as a potential functional target in Asian gastric cancer. Further, we experimentally demonstrated the translational relevance of this gene as a potential therapeutic target for early-stage gastric cancer in Asian patients. Together, our findings not only provide a valuable information resource for identifying and elucidating the molecular mechanisms of Asian gastric cancer, but also represent a general integrative framework to develop more effective therapeutic targets. PMID:22434430

Nutrigenomics: implications for breast and colon cancer prevention.

PubMed

Riscuta, Gabriela; Dumitrescu, Ramona G

2012-01-01

Nutrigenomics refers to the interaction between one's diet and his/her genes. These interactions can markedly influence digestion, absorption, and the elimination of bioactive food components, as well as influence their site of actions/molecular targets. Nutrigenomics comprises nutrigenetics, epigenetics, and transcriptomics, coupled with other "omic," such as proteomics and metabolomics, that apparently account for the wide variability in cancer risk among individuals with similar dietary habits. Multiple food components including essential nutrients, phytochemical, zoochemicals, fungochemical, and bacterochemicals have been implicated in cancer risk and tumor behavior, admittedly with mixed results. Such findings suggest that not all individuals respond identically to a diet. This chapter highlights the influence of single-nucleotide polymorphism, copy number, epigenetic events, and transcriptomic homeostasis as factors influencing the response to food components and ultimately health, including cancer risk. Both breast and colorectal cancers are reviewed as examples about how nutrigenomics may influence the response to dietary intakes. As the concept that "one size fits all" comes to an end and personalized approaches surface, additional research data will be required to identify those who will benefit most from dietary change and any who might be placed at risk because of an adjustment.

The aquatic animals' transcriptome resource for comparative functional analysis.

PubMed

Chou, Chih-Hung; Huang, Hsi-Yuan; Huang, Wei-Chih; Hsu, Sheng-Da; Hsiao, Chung-Der; Liu, Chia-Yu; Chen, Yu-Hung; Liu, Yu-Chen; Huang, Wei-Yun; Lee, Meng-Lin; Chen, Yi-Chang; Huang, Hsien-Da

2018-05-09

Aquatic animals have great economic and ecological importance. Among them, non-model organisms have been studied regarding eco-toxicity, stress biology, and environmental adaptation. Due to recent advances in next-generation sequencing techniques, large amounts of RNA-seq data for aquatic animals are publicly available. However, currently there is no comprehensive resource exist for the analysis, unification, and integration of these datasets. This study utilizes computational approaches to build a new resource of transcriptomic maps for aquatic animals. This aquatic animal transcriptome map database dbATM provides de novo assembly of transcriptome, gene annotation and comparative analysis of more than twenty aquatic organisms without draft genome. To improve the assembly quality, three computational tools (Trinity, Oases and SOAPdenovo-Trans) were employed to enhance individual transcriptome assembly, and CAP3 and CD-HIT-EST software were then used to merge these three assembled transcriptomes. In addition, functional annotation analysis provides valuable clues to gene characteristics, including full-length transcript coding regions, conserved domains, gene ontology and KEGG pathways. Furthermore, all aquatic animal genes are essential for comparative genomics tasks such as constructing homologous gene groups and blast databases and phylogenetic analysis. In conclusion, we establish a resource for non model organism aquatic animals, which is great economic and ecological importance and provide transcriptomic information including functional annotation and comparative transcriptome analysis. The database is now publically accessible through the URL http://dbATM.mbc.nctu.edu.tw/ .

VESPA: Software to Facilitate Genomic Annotation of Prokaryotic Organisms Through Integration of Proteomic and Transcriptomic Data

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

Peterson, Elena S.; McCue, Lee Ann; Rutledge, Alexandra C.

2012-04-25

Visual Exploration and Statistics to Promote Annotation (VESPA) is an interactive visual analysis software tool that facilitates the discovery of structural mis-annotations in prokaryotic genomes. VESPA integrates high-throughput peptide-centric proteomics data and oligo-centric or RNA-Seq transcriptomics data into a genomic context. The data may be interrogated via visual analysis across multiple levels of genomic resolution, linked searches, exports and interaction with BLAST to rapidly identify location of interest within the genome and evaluate potential mis-annotations.

Transcriptome Analysis of Mycobacteria-Specific CD4+ T Cells Identified by Activation-Induced Expression of CD154.

PubMed

Kunnath-Velayudhan, Shajo; Goldberg, Michael F; Saini, Neeraj K; Johndrow, Christopher T; Ng, Tony W; Johnson, Alison J; Xu, Jiayong; Chan, John; Jacobs, William R; Porcelli, Steven A

2017-10-01

Analysis of Ag-specific CD4 + T cells in mycobacterial infections at the transcriptome level is informative but technically challenging. Although several methods exist for identifying Ag-specific T cells, including intracellular cytokine staining, cell surface cytokine-capture assays, and staining with peptide:MHC class II multimers, all of these have significant technical constraints that limit their usefulness. Measurement of activation-induced expression of CD154 has been reported to detect live Ag-specific CD4 + T cells, but this approach remains underexplored and, to our knowledge, has not previously been applied in mycobacteria-infected animals. In this article, we show that CD154 expression identifies adoptively transferred or endogenous Ag-specific CD4 + T cells induced by Mycobacterium bovis bacillus Calmette-Guérin vaccination. We confirmed that Ag-specific cytokine production was positively correlated with CD154 expression by CD4 + T cells from bacillus Calmette-Guérin-vaccinated mice and show that high-quality microarrays can be performed from RNA isolated from CD154 + cells purified by cell sorting. Analysis of microarray data demonstrated that the transcriptome of CD4 + CD154 + cells was distinct from that of CD154 - cells and showed major enrichment of transcripts encoding multiple cytokines and pathways of cellular activation. One notable finding was the identification of a previously unrecognized subset of mycobacteria-specific CD4 + T cells that is characterized by the production of IL-3. Our results support the use of CD154 expression as a practical and reliable method to isolate live Ag-specific CD4 + T cells for transcriptomic analysis and potentially for a range of other studies in infected or previously immunized hosts. Copyright © 2017 by The American Association of Immunologists, Inc.

Optimizing and benchmarking de novo transcriptome sequencing: from library preparation to assembly evaluation.

PubMed

Hara, Yuichiro; Tatsumi, Kaori; Yoshida, Michio; Kajikawa, Eriko; Kiyonari, Hiroshi; Kuraku, Shigehiro

2015-11-18

RNA-seq enables gene expression profiling in selected spatiotemporal windows and yields massive sequence information with relatively low cost and time investment, even for non-model species. However, there remains a large room for optimizing its workflow, in order to take full advantage of continuously developing sequencing capacity. Transcriptome sequencing for three embryonic stages of Madagascar ground gecko (Paroedura picta) was performed with the Illumina platform. The output reads were assembled de novo for reconstructing transcript sequences. In order to evaluate the completeness of transcriptome assemblies, we prepared a reference gene set consisting of vertebrate one-to-one orthologs. To take advantage of increased read length of >150 nt, we demonstrated shortened RNA fragmentation time, which resulted in a dramatic shift of insert size distribution. To evaluate products of multiple de novo assembly runs incorporating reads with different RNA sources, read lengths, and insert sizes, we introduce a new reference gene set, core vertebrate genes (CVG), consisting of 233 genes that are shared as one-to-one orthologs by all vertebrate genomes examined (29 species)., The completeness assessment performed by the computational pipelines CEGMA and BUSCO referring to CVG, demonstrated higher accuracy and resolution than with the gene set previously established for this purpose. As a result of the assessment with CVG, we have derived the most comprehensive transcript sequence set of the Madagascar ground gecko by means of assembling individual libraries followed by clustering the assembled sequences based on their overall similarities. Our results provide several insights into optimizing de novo RNA-seq workflow, including the coordination between library insert size and read length, which manifested in improved connectivity of assemblies. The approach and assembly assessment with CVG demonstrated here would be applicable to transcriptome analysis of other species as well as whole genome analyses.

AmpuBase: a transcriptome database for eight species of apple snails (Gastropoda: Ampullariidae).

PubMed

Ip, Jack C H; Mu, Huawei; Chen, Qian; Sun, Jin; Ituarte, Santiago; Heras, Horacio; Van Bocxlaer, Bert; Ganmanee, Monthon; Huang, Xin; Qiu, Jian-Wen

2018-03-05

Gastropoda, with approximately 80,000 living species, is the largest class of Mollusca. Among gastropods, apple snails (family Ampullariidae) are globally distributed in tropical and subtropical freshwater ecosystems and many species are ecologically and economically important. Ampullariids exhibit various morphological and physiological adaptations to their respective habitats, which make them ideal candidates for studying adaptation, population divergence, speciation, and larger-scale patterns of diversity, including the biogeography of native and invasive populations. The limited availability of genomic data, however, hinders in-depth ecological and evolutionary studies of these non-model organisms. Using Illumina Hiseq platforms, we sequenced 1220 million reads for seven species of apple snails. Together with the previously published RNA-Seq data of two apple snails, we conducted de novo transcriptome assembly of eight species that belong to five genera of Ampullariidae, two of which represent Old World lineages and the other three New World lineages. There were 20,730 to 35,828 unigenes with predicted open reading frames for the eight species, with N50 (shortest sequence length at 50% of the unigenes) ranging from 1320 to 1803 bp. 69.7% to 80.2% of these unigenes were functionally annotated by searching against NCBI's non-redundant, Gene Ontology database and the Kyoto Encyclopaedia of Genes and Genomes. With these data we developed AmpuBase, a relational database that features online BLAST functionality for DNA/protein sequences, keyword searching for unigenes/functional terms, and download functions for sequences and whole transcriptomes. In summary, we have generated comprehensive transcriptome data for multiple ampullariid genera and species, and created a publicly accessible database with a user-friendly interface to facilitate future basic and applied studies on ampullariids, and comparative molecular studies with other invertebrates.

Transcriptome-Wide Analysis of Hepatitis B Virus-Mediated Changes to Normal Hepatocyte Gene Expression.

PubMed

Lamontagne, Jason; Mell, Joshua C; Bouchard, Michael J

2016-02-01

Globally, a chronic hepatitis B virus (HBV) infection remains the leading cause of primary liver cancer. The mechanisms leading to the development of HBV-associated liver cancer remain incompletely understood. In part, this is because studies have been limited by the lack of effective model systems that are both readily available and mimic the cellular environment of a normal hepatocyte. Additionally, many studies have focused on single, specific factors or pathways that may be affected by HBV, without addressing cell physiology as a whole. Here, we apply RNA-seq technology to investigate transcriptome-wide, HBV-mediated changes in gene expression to identify single factors and pathways as well as networks of genes and pathways that are affected in the context of HBV replication. Importantly, these studies were conducted in an ex vivo model of cultured primary hepatocytes, allowing for the transcriptomic characterization of this model system and an investigation of early HBV-mediated effects in a biologically relevant context. We analyzed differential gene expression within the context of time-mediated gene-expression changes and show that in the context of HBV replication a number of genes and cellular pathways are altered, including those associated with metabolism, cell cycle regulation, and lipid biosynthesis. Multiple analysis pipelines, as well as qRT-PCR and an independent, replicate RNA-seq analysis, were used to identify and confirm differentially expressed genes. HBV-mediated alterations to the transcriptome that we identified likely represent early changes to hepatocytes following an HBV infection, suggesting potential targets for early therapeutic intervention. Overall, these studies have produced a valuable resource that can be used to expand our understanding of the complex network of host-virus interactions and the impact of HBV-mediated changes to normal hepatocyte physiology on viral replication.

Transcriptomic and innate immune responses to Yersinia pestis in the lymph node during bubonic plague.

PubMed

Comer, Jason E; Sturdevant, Daniel E; Carmody, Aaron B; Virtaneva, Kimmo; Gardner, Donald; Long, Dan; Rosenke, Rebecca; Porcella, Stephen F; Hinnebusch, B Joseph

2010-12-01

A delayed inflammatory response is a prominent feature of infection with Yersinia pestis, the agent of bubonic and pneumonic plague. Using a rat model of bubonic plague, we examined lymph node histopathology, transcriptome, and extracellular cytokine levels to broadly characterize the kinetics and extent of the host response to Y. pestis and how it is influenced by the Yersinia virulence plasmid (pYV). Remarkably, dissemination and multiplication of wild-type Y. pestis during the bubonic stage of disease did not induce any detectable gene expression or cytokine response by host lymph node cells in the developing bubo. Only after systemic spread had led to terminal septicemic plague was a transcriptomic response detected, which included upregulation of several cytokine, chemokine, and other immune response genes. Although an initial intracellular phase of Y. pestis infection has been postulated, a Th1-type cytokine response associated with classical activation of macrophages was not observed during the bubonic stage of disease. However, elevated levels of interleukin-17 (IL-17) were present in infected lymph nodes. In the absence of pYV, sustained recruitment to the lymph node of polymorphonuclear leukocytes (PMN, or neutrophils), the major IL-17 effector cells, correlated with clearance of infection. Thus, the ability to counteract a PMN response in the lymph node appears to be a major in vivo function of the Y. pestis virulence plasmid.

Modifying Effects of Vitamin E on Chlorpyrifos Toxicity in Atlantic Salmon

PubMed Central

Olsvik, Pål A.; Berntssen, Marc H. G.; Søfteland, Liv

2015-01-01

The aim of this study was to elucidate how vitamin E (alpha tocopherol) may ameliorate the toxicity of the pesticide chlorpyrifos in Atlantic salmon. Freshly isolated hepatocytes were exposed to vitamin E, chlorpyrifos or a combination of vitamin E and chlorpyrifos (all 100 μM). Transcriptomics (RNA-seq) and metabolomics were used to screen for effects of vitamin E and chlorpyrifos. By introducing vitamin E, the number of upregulated transcripts induced by chlorpyrifos exposure was reduced from 941 to 626, while the number of downregulated transcripts was reduced from 901 to 742 compared to the control. Adding only vitamin E had no effect on the transcriptome. Jak-STAT signaling was the most significantly affected pathway by chlorpyrifos treatment according to the transcriptomics data. The metabolomics data showed that accumulation of multiple long chain fatty acids and dipeptides and amino acids in chlorpyrifos treated cells was partially alleviated by vitamin E treatment. Significant interaction effects between chlorpyrifos and vitamin E were seen for 15 metabolites, including 12 dipeptides. The antioxidant had relatively modest effects on chlorpyrifos-induced oxidative stress. By combining the two data sets, the study suggests that vitamin E supplementation prevents uptake and accumulation of fatty acids, and counteracts inhibited carbohydrate metabolism. Overall, this study shows that vitamin E only to a moderate degree modifies chlorpyrifos toxicity in Atlantic salmon liver cells. PMID:25774794

Integrated network analysis identifies fight-club nodes as a class of hubs encompassing key putative switch genes that induce major transcriptome reprogramming during grapevine development.

PubMed

Palumbo, Maria Concetta; Zenoni, Sara; Fasoli, Marianna; Massonnet, Mélanie; Farina, Lorenzo; Castiglione, Filippo; Pezzotti, Mario; Paci, Paola

2014-12-01

We developed an approach that integrates different network-based methods to analyze the correlation network arising from large-scale gene expression data. By studying grapevine (Vitis vinifera) and tomato (Solanum lycopersicum) gene expression atlases and a grapevine berry transcriptomic data set during the transition from immature to mature growth, we identified a category named "fight-club hubs" characterized by a marked negative correlation with the expression profiles of neighboring genes in the network. A special subset named "switch genes" was identified, with the additional property of many significant negative correlations outside their own group in the network. Switch genes are involved in multiple processes and include transcription factors that may be considered master regulators of the previously reported transcriptome remodeling that marks the developmental shift from immature to mature growth. All switch genes, expressed at low levels in vegetative/green tissues, showed a significant increase in mature/woody organs, suggesting a potential regulatory role during the developmental transition. Finally, our analysis of tomato gene expression data sets showed that wild-type switch genes are downregulated in ripening-deficient mutants. The identification of known master regulators of tomato fruit maturation suggests our method is suitable for the detection of key regulators of organ development in different fleshy fruit crops. © 2014 American Society of Plant Biologists. All rights reserved.

Integrated Network Analysis Identifies Fight-Club Nodes as a Class of Hubs Encompassing Key Putative Switch Genes That Induce Major Transcriptome Reprogramming during Grapevine Development[W][OPEN

PubMed Central

Palumbo, Maria Concetta; Zenoni, Sara; Fasoli, Marianna; Massonnet, Mélanie; Farina, Lorenzo; Castiglione, Filippo; Pezzotti, Mario; Paci, Paola

2014-01-01

We developed an approach that integrates different network-based methods to analyze the correlation network arising from large-scale gene expression data. By studying grapevine (Vitis vinifera) and tomato (Solanum lycopersicum) gene expression atlases and a grapevine berry transcriptomic data set during the transition from immature to mature growth, we identified a category named “fight-club hubs” characterized by a marked negative correlation with the expression profiles of neighboring genes in the network. A special subset named “switch genes” was identified, with the additional property of many significant negative correlations outside their own group in the network. Switch genes are involved in multiple processes and include transcription factors that may be considered master regulators of the previously reported transcriptome remodeling that marks the developmental shift from immature to mature growth. All switch genes, expressed at low levels in vegetative/green tissues, showed a significant increase in mature/woody organs, suggesting a potential regulatory role during the developmental transition. Finally, our analysis of tomato gene expression data sets showed that wild-type switch genes are downregulated in ripening-deficient mutants. The identification of known master regulators of tomato fruit maturation suggests our method is suitable for the detection of key regulators of organ development in different fleshy fruit crops. PMID:25490918

Transcriptome-wide comparison of the impact of Atoh1 and miR-183 family on pluripotent stem cells and multipotent otic progenitor cells.

PubMed

Ebeid, Michael; Sripal, Prashanth; Pecka, Jason; Beisel, Kirk W; Kwan, Kelvin; Soukup, Garrett A

2017-01-01

Over 5% of the global population suffers from disabling hearing loss caused by multiple factors including aging, noise exposure, genetic predisposition, or use of ototoxic drugs. Sensorineural hearing loss is often caused by the loss of sensory hair cells (HCs) of the inner ear. A barrier to hearing restoration after HC loss is the limited ability of mammalian auditory HCs to spontaneously regenerate. Understanding the molecular mechanisms orchestrating HC development is expected to facilitate cell replacement therapies. Multiple events are known to be essential for proper HC development including the expression of Atoh1 transcription factor and the miR-183 family. We have developed a series of vectors expressing the miR-183 family and/or Atoh1 that was used to transfect two different developmental cell models: pluripotent mouse embryonic stem cells (mESCs) and immortalized multipotent otic progenitor (iMOP) cells representing an advanced developmental stage. Transcriptome profiling of transfected cells show that the impact of Atoh1 is contextually dependent with more HC-specific effects on iMOP cells. miR-183 family expression in combination with Atoh1 not only appears to fine tune gene expression in favor of HC fate, but is also required for the expression of some HC-specific genes. Overall, the work provides novel insight into the combined role of Atoh1 and the miR-183 family during HC development that may ultimately inform strategies to promote HC regeneration or maintenance.

Understanding the response to endurance exercise using a systems biology approach: combining blood metabolomics, transcriptomics and miRNomics in horses.

PubMed

Mach, Núria; Ramayo-Caldas, Yuliaxis; Clark, Allison; Moroldo, Marco; Robert, Céline; Barrey, Eric; López, Jesús Maria; Le Moyec, Laurence

2017-02-17

Endurance exercise in horses requires adaptive processes involving physiological, biochemical, and cognitive-behavioral responses in an attempt to regain homeostasis. We hypothesized that the identification of the relationships between blood metabolome, transcriptome, and miRNome during endurance exercise in horses could provide significant insights into the molecular response to endurance exercise. For this reason, the serum metabolome and whole-blood transcriptome and miRNome data were obtained from ten horses before and after a 160 km endurance competition. We obtained a global regulatory network based on 11 unique metabolites, 263 metabolic genes and 5 miRNAs whose expression was significantly altered at T1 (post- endurance competition) relative to T0 (baseline, pre-endurance competition). This network provided new insights into the cross talk between the distinct molecular pathways (e.g. energy and oxygen sensing, oxidative stress, and inflammation) that were not detectable when analyzing single metabolites or transcripts alone. Single metabolites and transcripts were carrying out multiple roles and thus sharing several biochemical pathways. Using a regulatory impact factor metric analysis, this regulatory network was further confirmed at the transcription factor and miRNA levels. In an extended cohort of 31 independent animals, multiple factor analysis confirmed the strong associations between lactate, methylene derivatives, miR-21-5p, miR-16-5p, let-7 family and genes that coded proteins involved in metabolic reactions primarily related to energy, ubiquitin proteasome and lipopolysaccharide immune responses after the endurance competition. Multiple factor analysis also identified potential biomarkers at T0 for an increased likelihood for failure to finish an endurance competition. To the best of our knowledge, the present study is the first to provide a comprehensive and integrated overview of the metabolome, transcriptome, and miRNome co-regulatory networks that may have a key role in regulating the metabolic and immune response to endurance exercise in horses.

Comparative transcriptomics of early dipteran development

PubMed Central

2013-01-01

Background Modern sequencing technologies have massively increased the amount of data available for comparative genomics. Whole-transcriptome shotgun sequencing (RNA-seq) provides a powerful basis for comparative studies. In particular, this approach holds great promise for emerging model species in fields such as evolutionary developmental biology (evo-devo). Results We have sequenced early embryonic transcriptomes of two non-drosophilid dipteran species: the moth midge Clogmia albipunctata, and the scuttle fly Megaselia abdita. Our analysis includes a third, published, transcriptome for the hoverfly Episyrphus balteatus. These emerging models for comparative developmental studies close an important phylogenetic gap between Drosophila melanogaster and other insect model systems. In this paper, we provide a comparative analysis of early embryonic transcriptomes across species, and use our data for a phylogenomic re-evaluation of dipteran phylogenetic relationships. Conclusions We show how comparative transcriptomics can be used to create useful resources for evo-devo, and to investigate phylogenetic relationships. Our results demonstrate that de novo assembly of short (Illumina) reads yields high-quality, high-coverage transcriptomic data sets. We use these data to investigate deep dipteran phylogenetic relationships. Our results, based on a concatenation of 160 orthologous genes, provide support for the traditional view of Clogmia being the sister group of Brachycera (Megaselia, Episyrphus, Drosophila), rather than that of Culicomorpha (which includes mosquitoes and blackflies). PMID:23432914

The Plasmodium falciparum transcriptome in severe malaria reveals altered expression of genes involved in important processes including surface antigen–encoding var genes

PubMed Central

Tonkin-Hill, Gerry Q.; Trianty, Leily; Noviyanti, Rintis; Nguyen, Hanh H. T.; Sebayang, Boni F.; Lampah, Daniel A.; Marfurt, Jutta; Cobbold, Simon A.; Rambhatla, Janavi S.; McConville, Malcolm J.; Rogerson, Stephen J.; Brown, Graham V.; Day, Karen P.; Price, Ric N.; Anstey, Nicholas M.

2018-01-01

Within the human host, the malaria parasite Plasmodium falciparum is exposed to multiple selection pressures. The host environment changes dramatically in severe malaria, but the extent to which the parasite responds to—or is selected by—this environment remains unclear. From previous studies, the parasites that cause severe malaria appear to increase expression of a restricted but poorly defined subset of the PfEMP1 variant, surface antigens. PfEMP1s are major targets of protective immunity. Here, we used RNA sequencing (RNAseq) to analyse gene expression in 44 parasite isolates that caused severe and uncomplicated malaria in Papuan patients. The transcriptomes of 19 parasite isolates associated with severe malaria indicated that these parasites had decreased glycolysis without activation of compensatory pathways; altered chromatin structure and probably transcriptional regulation through decreased histone methylation; reduced surface expression of PfEMP1; and down-regulated expression of multiple chaperone proteins. Our RNAseq also identified novel associations between disease severity and PfEMP1 transcripts, domains, and smaller sequence segments and also confirmed all previously reported associations between expressed PfEMP1 sequences and severe disease. These findings will inform efforts to identify vaccine targets for severe malaria and also indicate how parasites adapt to—or are selected by—the host environment in severe malaria. PMID:29529020

Multiple Roles of Integrin-Linked Kinase in Epidermal Development, Maturation and Pigmentation Revealed by Molecular Profiling

PubMed Central

Judah, David; Rudkouskaya, Alena; Wilson, Ryan; Carter, David E.; Dagnino, Lina

2012-01-01

Integrin-linked kinase (ILK) is an important scaffold protein that mediates a variety of cellular responses to integrin stimulation by extracellular matrix proteins. Mice with epidermis-restricted inactivation of the Ilk gene exhibit pleiotropic phenotypic defects, including impaired hair follicle morphogenesis, reduced epidermal adhesion to the basement membrane, compromised epidermal integrity, as well as wasting and failure to thrive leading to perinatal death. To better understand the underlying molecular mechanisms that cause such a broad range of alterations, we investigated the impact of Ilk gene inactivation on the epidermis transcriptome. Microarray analysis showed over 700 differentially regulated mRNAs encoding proteins involved in multiple aspects of epidermal function, including keratinocyte differentiation and barrier formation, inflammation, regeneration after injury, and fundamental epidermal developmental pathways. These studies also revealed potential effects on genes not previously implicated in ILK functions, including those important for melanocyte and melanoblast development and function, regulation of cytoskeletal dynamics, and homeobox genes. This study shows that ILK is a critical regulator of multiple aspects of epidermal function and homeostasis, and reveals the previously unreported involvement of ILK not only in epidermal differentiation and barrier formation, but also in melanocyte genesis and function. PMID:22574216

Multiple roles of integrin-linked kinase in epidermal development, maturation and pigmentation revealed by molecular profiling.

PubMed

Judah, David; Rudkouskaya, Alena; Wilson, Ryan; Carter, David E; Dagnino, Lina

2012-01-01

Integrin-linked kinase (ILK) is an important scaffold protein that mediates a variety of cellular responses to integrin stimulation by extracellular matrix proteins. Mice with epidermis-restricted inactivation of the Ilk gene exhibit pleiotropic phenotypic defects, including impaired hair follicle morphogenesis, reduced epidermal adhesion to the basement membrane, compromised epidermal integrity, as well as wasting and failure to thrive leading to perinatal death. To better understand the underlying molecular mechanisms that cause such a broad range of alterations, we investigated the impact of Ilk gene inactivation on the epidermis transcriptome. Microarray analysis showed over 700 differentially regulated mRNAs encoding proteins involved in multiple aspects of epidermal function, including keratinocyte differentiation and barrier formation, inflammation, regeneration after injury, and fundamental epidermal developmental pathways. These studies also revealed potential effects on genes not previously implicated in ILK functions, including those important for melanocyte and melanoblast development and function, regulation of cytoskeletal dynamics, and homeobox genes. This study shows that ILK is a critical regulator of multiple aspects of epidermal function and homeostasis, and reveals the previously unreported involvement of ILK not only in epidermal differentiation and barrier formation, but also in melanocyte genesis and function.

Genetic variation and gene expression across multiple tissues and developmental stages in a nonhuman primate.

PubMed

Jasinska, Anna J; Zelaya, Ivette; Service, Susan K; Peterson, Christine B; Cantor, Rita M; Choi, Oi-Wa; DeYoung, Joseph; Eskin, Eleazar; Fairbanks, Lynn A; Fears, Scott; Furterer, Allison E; Huang, Yu S; Ramensky, Vasily; Schmitt, Christopher A; Svardal, Hannes; Jorgensen, Matthew J; Kaplan, Jay R; Villar, Diego; Aken, Bronwen L; Flicek, Paul; Nag, Rishi; Wong, Emily S; Blangero, John; Dyer, Thomas D; Bogomolov, Marina; Benjamini, Yoav; Weinstock, George M; Dewar, Ken; Sabatti, Chiara; Wilson, Richard K; Jentsch, J David; Warren, Wesley; Coppola, Giovanni; Woods, Roger P; Freimer, Nelson B

2017-12-01

By analyzing multitissue gene expression and genome-wide genetic variation data in samples from a vervet monkey pedigree, we generated a transcriptome resource and produced the first catalog of expression quantitative trait loci (eQTLs) in a nonhuman primate model. This catalog contains more genome-wide significant eQTLs per sample than comparable human resources and identifies sex- and age-related expression patterns. Findings include a master regulatory locus that likely has a role in immune function and a locus regulating hippocampal long noncoding RNAs (lncRNAs), whose expression correlates with hippocampal volume. This resource will facilitate genetic investigation of quantitative traits, including brain and behavioral phenotypes relevant to neuropsychiatric disorders.

Heterogeneous data fusion for brain tumor classification.

PubMed

Metsis, Vangelis; Huang, Heng; Andronesi, Ovidiu C; Makedon, Fillia; Tzika, Aria

2012-10-01

Current research in biomedical informatics involves analysis of multiple heterogeneous data sets. This includes patient demographics, clinical and pathology data, treatment history, patient outcomes as well as gene expression, DNA sequences and other information sources such as gene ontology. Analysis of these data sets could lead to better disease diagnosis, prognosis, treatment and drug discovery. In this report, we present a novel machine learning framework for brain tumor classification based on heterogeneous data fusion of metabolic and molecular datasets, including state-of-the-art high-resolution magic angle spinning (HRMAS) proton (1H) magnetic resonance spectroscopy and gene transcriptome profiling, obtained from intact brain tumor biopsies. Our experimental results show that our novel framework outperforms any analysis using individual dataset.

De Novo Assembly of a Transcriptome for Calanus finmarchicus (Crustacea, Copepoda) – The Dominant Zooplankter of the North Atlantic Ocean

PubMed Central

Lenz, Petra H.; Roncalli, Vittoria; Hassett, R. Patrick; Wu, Le-Shin; Cieslak, Matthew C.; Hartline, Daniel K.; Christie, Andrew E.

2014-01-01

Assessing the impact of global warming on the food web of the North Atlantic will require difficult-to-obtain physiological data on a key copepod crustacean, Calanus finmarchicus. The de novo transcriptome presented here represents a new resource for acquiring such data. It was produced from multiplexed gene libraries using RNA collected from six developmental stages: embryo, early nauplius (NI-II), late nauplius (NV-VI), early copepodite (CI-II), late copepodite (CV) and adult (CVI) female. Over 400,000,000 paired-end reads (100 base-pairs long) were sequenced on an Illumina instrument, and assembled into 206,041 contigs using Trinity software. Coverage was estimated to be at least 65%. A reference transcriptome comprising 96,090 unique components (“comps”) was annotated using Blast2GO. 40% of the comps had significant blast hits. 11% of the comps were successfully annotated with gene ontology (GO) terms. Expression of many comps was found to be near zero in one or more developmental stages suggesting that 35 to 48% of the transcriptome is “silent” at any given life stage. Transcripts involved in lipid biosynthesis pathways, critical for the C. finmarchicus life cycle, were identified and their expression pattern during development was examined. Relative expression of three transcripts suggests wax ester biosynthesis in late copepodites, but triacylglyceride biosynthesis in adult females. Two of these transcripts may be involved in the preparatory phase of diapause. A key environmental challenge for C. finmarchicus is the seasonal exposure to the dinoflagellate Alexandrium fundyense with high concentrations of saxitoxins, neurotoxins that block voltage-gated sodium channels. Multiple contigs encoding putative voltage-gated sodium channels were identified. They appeared to be the result of both alternate splicing and gene duplication. This is the first report of multiple NaV1 genes in a protostome. These data provide new insights into the transcriptome and physiology of this environmentally important zooplankter. PMID:24586345

Transcriptomic Dose-Response Analysis for Mode of Action ...

EPA Pesticide Factsheets

Microarray and RNA-seq technologies can play an important role in assessing the health risks associated with environmental exposures. The utility of gene expression data to predict hazard has been well documented. Early toxicogenomics studies used relatively high, single doses with minimal replication. Thus, they were not useful in understanding health risks at environmentally-relevant doses. Until the past decade, application of toxicogenomics in dose response assessment and determination of chemical mode of action has been limited. New transcriptomic biomarkers have evolved to detect chemical hazards in multiple tissues together with pathway methods to study biological effects across the full dose response range and critical time course. Comprehensive low dose datasets are now available and with the use of transcriptomic benchmark dose estimation techniques within a mode of action framework, the ability to incorporate informative genomic data into human health risk assessment has substantially improved. The key advantage to applying transcriptomic technology to risk assessment is both the sensitivity and comprehensive examination of direct and indirect molecular changes that lead to adverse outcomes. Book Chapter with topic on future application of toxicogenomics technologies for MoA and risk assessment

Adult Mouse Cortical Cell Taxonomy by Single Cell Transcriptomics

PubMed Central

Tasic, Bosiljka; Menon, Vilas; Nguyen, Thuc Nghi; Kim, Tae Kyung; Jarsky, Tim; Yao, Zizhen; Levi, Boaz; Gray, Lucas T.; Sorensen, Staci A.; Dolbeare, Tim; Bertagnolli, Darren; Goldy, Jeff; Shapovalova, Nadiya; Parry, Sheana; Lee, Changkyu; Smith, Kimberly; Bernard, Amy; Madisen, Linda; Sunkin, Susan M.; Hawrylycz, Michael; Koch, Christof; Zeng, Hongkui

2016-01-01

Nervous systems are composed of various cell types, but the extent of cell type diversity is poorly understood. Here, we construct a cellular taxonomy of one cortical region, primary visual cortex, in adult mice based on single cell RNA-sequencing. We identify 49 transcriptomic cell types including 23 GABAergic, 19 glutamatergic and seven non-neuronal types. We also analyze cell-type specific mRNA processing and characterize genetic access to these transcriptomic types by many transgenic Cre lines. Finally, we show that some of our transcriptomic cell types display specific and differential electrophysiological and axon projection properties, thereby confirming that the single cell transcriptomic signatures can be associated with specific cellular properties. PMID:26727548

Construction of Pará rubber tree genome and multi-transcriptome database accelerates rubber researches.

PubMed

Makita, Yuko; Kawashima, Mika; Lau, Nyok Sean; Othman, Ahmad Sofiman; Matsui, Minami

2018-01-19

Natural rubber is an economically important material. Currently the Pará rubber tree, Hevea brasiliensis is the main commercial source. Little is known about rubber biosynthesis at the molecular level. Next-generation sequencing (NGS) technologies brought draft genomes of three rubber cultivars and a variety of RNA sequencing (RNA-seq) data. However, no current genome or transcriptome databases (DB) are organized by gene. A gene-oriented database is a valuable support for rubber research. Based on our original draft genome sequence of H. brasiliensis RRIM600, we constructed a rubber tree genome and transcriptome DB. Our DB provides genome information including gene functional annotations and multi-transcriptome data of RNA-seq, full-length cDNAs including PacBio Isoform sequencing (Iso-Seq), ESTs and genome wide transcription start sites (TSSs) derived from CAGE technology. Using our original and publically available RNA-seq data, we calculated co-expressed genes for identifying functionally related gene sets and/or genes regulated by the same transcription factor (TF). Users can access multi-transcriptome data through both a gene-oriented web page and a genome browser. For the gene searching system, we provide keyword search, sequence homology search and gene expression search; users can also select their expression threshold easily. The rubber genome and transcriptome DB provides rubber tree genome sequence and multi-transcriptomics data. This DB is useful for comprehensive understanding of the rubber transcriptome. This will assist both industrial and academic researchers for rubber and economically important close relatives such as R. communis, M. esculenta and J. curcas. The Rubber Transcriptome DB release 2017.03 is accessible at http://matsui-lab.riken.jp/rubber/ .

Lactobacillus rhamnosus CNCMI-4317 Modulates Fiaf/Angptl4 in Intestinal Epithelial Cells and Circulating Level in Mice

PubMed Central

Jacouton, Elsa; Mach, Núria; Cadiou, Julie; Lapaque, Nicolas; Clément, Karine; Doré, Joël; van Hylckama Vlieg, Johan E. T.; Smokvina, Tamara; Blottière, Hervé M

2015-01-01

Background and Objectives Identification of new targets for metabolic diseases treatment or prevention is required. In this context, FIAF/ANGPTL4 appears as a crucial regulator of energy homeostasis. Lactobacilli are often considered to display beneficial effect for their hosts, acting on different regulatory pathways. The aim of the present work was to study the effect of several lactobacilli strains on Fiaf gene expression in human intestinal epithelial cells (IECs) and on mice tissues to decipher the underlying mechanisms. Subjects and Methods Nineteen lactobacilli strains have been tested on HT–29 human intestinal epithelial cells for their ability to regulate Fiaf gene expression by RT-qPCR. In order to determine regulated pathways, we analysed the whole genome transcriptome of IECs. We then validated in vivo bacterial effects using C57BL/6 mono-colonized mice fed with normal chow. Results We identified one strain (Lactobacillus rhamnosus CNCMI–4317) that modulated Fiaf expression in IECs. This regulation relied potentially on bacterial surface-exposed molecules and seemed to be PPAR-γ independent but PPAR-α dependent. Transcriptome functional analysis revealed that multiple pathways including cellular function and maintenance, lymphoid tissue structure and development, as well as lipid metabolism were regulated by this strain. The regulation of immune system and lipid and carbohydrate metabolism was also confirmed by overrepresentation of Gene Ontology terms analysis. In vivo, circulating FIAF protein was increased by the strain but this phenomenon was not correlated with modulation Fiaf expression in tissues (except a trend in distal small intestine). Conclusion We showed that Lactobacillus rhamnosus CNCMI–4317 induced Fiaf expression in human IECs, and increased circulating FIAF protein level in mice. Moreover, this effect was accompanied by transcriptome modulation of several pathways including immune response and metabolism in vitro. PMID:26439630

Genome-Wide Mapping of Cystitis Due to Streptococcus agalactiae and Escherichia coli in Mice Identifies a Unique Bladder Transcriptome That Signifies Pathogen-Specific Antimicrobial Defense against Urinary Tract Infection

PubMed Central

Tan, Chee K.; Carey, Alison J.; Cui, Xiangqin; Webb, Richard I.; Ipe, Deepak; Crowley, Michael; Cripps, Allan W.; Benjamin, William H.; Ulett, Kimberly B.; Schembri, Mark A.

2012-01-01

The most common causes of urinary tract infections (UTIs) are Gram-negative pathogens such as Escherichia coli; however, Gram-positive organisms, including Streptococcus agalactiae, or group B streptococcus (GBS), also cause UTI. In GBS infection, UTI progresses to cystitis once the bacteria colonize the bladder, but the host responses triggered in the bladder immediately following infection are largely unknown. Here, we used genome-wide expression profiling to map the bladder transcriptome of GBS UTI in mice infected transurethrally with uropathogenic GBS that was cultured from a 35-year-old women with cystitis. RNA from bladders was applied to Affymetrix Gene-1.0ST microarrays; quantitative reverse transcriptase PCR (qRT-PCR) was used to analyze selected gene responses identified in array data sets. A surprisingly small significant-gene list of 172 genes was identified at 24 h; this compared to 2,507 genes identified in a side-by-side comparison with uropathogenic E. coli (UPEC). No genes exhibited significantly altered expression at 2 h in GBS-infected mice according to arrays despite high bladder bacterial loads at this early time point. The absence of a marked early host response to GBS juxtaposed with broad-based bladder responses activated by UPEC at 2 h. Bioinformatics analyses, including integrative system-level network mapping, revealed multiple activated biological pathways in the GBS bladder transcriptome that regulate leukocyte activation, inflammation, apoptosis, and cytokine-chemokine biosynthesis. These findings define a novel, minimalistic type of bladder host response triggered by GBS UTI, which comprises collective antimicrobial pathways that differ dramatically from those activated by UPEC. Overall, this study emphasizes the unique nature of bladder immune activation mechanisms triggered by distinct uropathogens. PMID:22733575

Genome and transcriptome sequencing in prospective metastatic triple-negative breast cancer uncovers therapeutic vulnerabilities.

PubMed

Craig, David W; O'Shaughnessy, Joyce A; Kiefer, Jeffrey A; Aldrich, Jessica; Sinari, Shripad; Moses, Tracy M; Wong, Shukmei; Dinh, Jennifer; Christoforides, Alexis; Blum, Joanne L; Aitelli, Cristi L; Osborne, Cynthia R; Izatt, Tyler; Kurdoglu, Ahmet; Baker, Angela; Koeman, Julie; Barbacioru, Catalin; Sakarya, Onur; De La Vega, Francisco M; Siddiqui, Asim; Hoang, Linh; Billings, Paul R; Salhia, Bodour; Tolcher, Anthony W; Trent, Jeffrey M; Mousses, Spyro; Von Hoff, Daniel; Carpten, John D

2013-01-01

Triple-negative breast cancer (TNBC) is characterized by the absence of expression of estrogen receptor, progesterone receptor, and HER-2. Thirty percent of patients recur after first-line treatment, and metastatic TNBC (mTNBC) has a poor prognosis with median survival of one year. Here, we present initial analyses of whole genome and transcriptome sequencing data from 14 prospective mTNBC. We have cataloged the collection of somatic genomic alterations in these advanced tumors, particularly those that may inform targeted therapies. Genes mutated in multiple tumors included TP53, LRP1B, HERC1, CDH5, RB1, and NF1. Notable genes involved in focal structural events were CTNNA1, PTEN, FBXW7, BRCA2, WT1, FGFR1, KRAS, HRAS, ARAF, BRAF, and PGCP. Homozygous deletion of CTNNA1 was detected in 2 of 6 African Americans. RNA sequencing revealed consistent overexpression of the FOXM1 gene when tumor gene expression was compared with nonmalignant breast samples. Using an outlier analysis of gene expression comparing one cancer with all the others, we detected expression patterns unique to each patient's tumor. Integrative DNA/RNA analysis provided evidence for deregulation of mutated genes, including the monoallelic expression of TP53 mutations. Finally, molecular alterations in several cancers supported targeted therapeutic intervention on clinical trials with known inhibitors, particularly for alterations in the RAS/RAF/MEK/ERK and PI3K/AKT/mTOR pathways. In conclusion, whole genome and transcriptome profiling of mTNBC have provided insights into somatic events occurring in this difficult to treat cancer. These genomic data have guided patients to investigational treatment trials and provide hypotheses for future trials in this irremediable cancer.

Transcriptomic Assessment of Isozymes in the Biphenyl Pathway of Rhodococcus sp. Strain RHA1†

PubMed Central

Gonçalves, Edmilson R.; Hara, Hirofumi; Miyazawa, Daisuke; Davies, Julian E.; Eltis, Lindsay D.; Mohn, William W.

2006-01-01

Rhodococcus sp. RHA1 grows on a broad range of aromatic compounds and vigorously degrades polychlorinated biphenyls (PCBs). Previous work identified RHA1 genes encoding multiple isozymes for most of the seven steps of the biphenyl (BPH) pathway, provided evidence for coexpression of some of these isozymes, and indicated the involvement of some of these enzymes in the degradation of BPH, ethylbenzene (ETB), and PCBs. To investigate the expression of these isozymes and better understand how they contribute to the robust degradative capacity of RHA1, we comprehensively analyzed the 9.7-Mb genome of RHA1 for BPH pathway genes and characterized the transcriptome of RHA1 growing on benzoate (BEN), BPH, and ETB. Sequence analyses revealed 54 potential BPH pathway genes, including 28 not previously reported. Transcriptomic analysis with a DNA microarray containing 70-mer probes for 8,213 RHA1 genes revealed a suite of 320 genes of diverse functions that were upregulated during growth both on BPH and on ETB, relative to growth on the control substrate, pyruvate. By contrast, only 65 genes were upregulated during growth on BEN. Quantitative PCR assays confirmed microarray results for selected genes and indicated that some of the catabolic genes were upregulated over 10,000-fold. Our analysis suggests that up to 22 enzymes, including 8 newly identified ones, may function in the BPH pathway of RHA1. The relative expression levels of catabolic genes did not differ for BPH and ETB, suggesting a common regulatory mechanism. This study delineated a suite of catabolic enzymes for biphenyl and alkyl-benzenes in RHA1, which is larger than previously recognized and which may serve as a model for catabolism in other environmentally important bacteria having large genomes. PMID:16957245

Single-Cell RNA-Seq Reveals Dynamic Early Embryonic-like Programs during Chemical Reprogramming.

PubMed

Zhao, Ting; Fu, Yao; Zhu, Jialiang; Liu, Yifang; Zhang, Qian; Yi, Zexuan; Chen, Shi; Jiao, Zhonggang; Xu, Xiaochan; Xu, Junquan; Duo, Shuguang; Bai, Yun; Tang, Chao; Li, Cheng; Deng, Hongkui

2018-06-12

Chemical reprogramming provides a powerful platform for exploring the molecular dynamics that lead to pluripotency. Although previous studies have uncovered an intermediate extraembryonic endoderm (XEN)-like state during this process, the molecular underpinnings of pluripotency acquisition remain largely undefined. Here, we profile 36,199 single-cell transcriptomes at multiple time points throughout a highly efficient chemical reprogramming system using RNA-sequencing and reconstruct their progression trajectories. Through identifying sequential molecular events, we reveal that the dynamic early embryonic-like programs are key aspects of successful reprogramming from XEN-like state to pluripotency, including the concomitant transcriptomic signatures of two-cell (2C) embryonic-like and early pluripotency programs and the epigenetic signature of notable genome-wide DNA demethylation. Moreover, via enhancing the 2C-like program by fine-tuning chemical treatment, the reprogramming process is remarkably accelerated. Collectively, our findings offer a high-resolution dissection of cell fate dynamics during chemical reprogramming and shed light on mechanistic insights into the nature of induced pluripotency. Copyright © 2018 Elsevier Inc. All rights reserved.

Variant discovery in the sheepmeat odour and flavour in javanese fat tailed sheep using RNA sequencing

NASA Astrophysics Data System (ADS)

Abuzahra, M. A. M.; Jakaria; Listyarini, K.; Furqon, A.; Sumantri, C.; Uddin, M. J.; Gunawan, A.

2018-05-01

High-throughput RNA sequencing (RNA-Seq) reveals new challenges for the detection of transcriptome variants (SNPs) in different tissues and species. The aims of this study was to characterize a SNP discovery analysis in the sheep meat odour and flavour transcriptome using RNA-Seq. Six liver samples from divergent sheep meat odour and flavour were analyzed using the Illumina Genome Hiseq 2500 Analyzer. The SNP detection analysis revealed 142 SNPs in sheep meat samples, and a large number of those corresponded to differences between high and low sheep meat odour and flavour ovis genome assembly OAR v4.0. Among them, about 90.4% of genes had multiple polymorphisms within 12 genes (JAML, ANGPTL8, LOC101103463, SEPW1, SCN5A, LOC101113036, DOCK6, GTSE1, KIF12, KCTD17, KANK2, CYP2A6). Several of the SNPs (JAML, CYP2A6, SEPW1, and KIF12) found in this study could be included as suitable markers in genotyping platforms to perform association analyses in commercial populations and apply genomic selection protocols in the sheep meat production.

MITIE: Simultaneous RNA-Seq-based transcript identification and quantification in multiple samples.

PubMed

Behr, Jonas; Kahles, André; Zhong, Yi; Sreedharan, Vipin T; Drewe, Philipp; Rätsch, Gunnar

2013-10-15

High-throughput sequencing of mRNA (RNA-Seq) has led to tremendous improvements in the detection of expressed genes and reconstruction of RNA transcripts. However, the extensive dynamic range of gene expression, technical limitations and biases, as well as the observed complexity of the transcriptional landscape, pose profound computational challenges for transcriptome reconstruction. We present the novel framework MITIE (Mixed Integer Transcript IdEntification) for simultaneous transcript reconstruction and quantification. We define a likelihood function based on the negative binomial distribution, use a regularization approach to select a few transcripts collectively explaining the observed read data and show how to find the optimal solution using Mixed Integer Programming. MITIE can (i) take advantage of known transcripts, (ii) reconstruct and quantify transcripts simultaneously in multiple samples, and (iii) resolve the location of multi-mapping reads. It is designed for genome- and assembly-based transcriptome reconstruction. We present an extensive study based on realistic simulated RNA-Seq data. When compared with state-of-the-art approaches, MITIE proves to be significantly more sensitive and overall more accurate. Moreover, MITIE yields substantial performance gains when used with multiple samples. We applied our system to 38 Drosophila melanogaster modENCODE RNA-Seq libraries and estimated the sensitivity of reconstructing omitted transcript annotations and the specificity with respect to annotated transcripts. Our results corroborate that a well-motivated objective paired with appropriate optimization techniques lead to significant improvements over the state-of-the-art in transcriptome reconstruction. MITIE is implemented in C++ and is available from http://bioweb.me/mitie under the GPL license.

Transcriptome dynamics through alternative polyadenylation in developmental and environmental responses in plants revealed by deep sequencing

PubMed Central

Shen, Yingjia; Venu, R.C.; Nobuta, Kan; Wu, Xiaohui; Notibala, Varun; Demirci, Caghan; Meyers, Blake C.; Wang, Guo-Liang; Ji, Guoli; Li, Qingshun Q.

2011-01-01

Polyadenylation sites mark the ends of mRNA transcripts. Alternative polyadenylation (APA) may alter sequence elements and/or the coding capacity of transcripts, a mechanism that has been demonstrated to regulate gene expression and transcriptome diversity. To study the role of APA in transcriptome dynamics, we analyzed a large-scale data set of RNA “tags” that signify poly(A) sites and expression levels of mRNA. These tags were derived from a wide range of tissues and developmental stages that were mutated or exposed to environmental treatments, and generated using digital gene expression (DGE)–based protocols of the massively parallel signature sequencing (MPSS-DGE) and the Illumina sequencing-by-synthesis (SBS-DGE) sequencing platforms. The data offer a global view of APA and how it contributes to transcriptome dynamics. Upon analysis of these data, we found that ∼60% of Arabidopsis genes have multiple poly(A) sites. Likewise, ∼47% and 82% of rice genes use APA, supported by MPSS-DGE and SBS-DGE tags, respectively. In both species, ∼49%–66% of APA events were mapped upstream of annotated stop codons. Interestingly, 10% of the transcriptomes are made up of APA transcripts that are differentially distributed among developmental stages and in tissues responding to environmental stresses, providing an additional level of transcriptome dynamics. Examples of pollen-specific APA switching and salicylic acid treatment-specific APA clearly demonstrated such dynamics. The significance of these APAs is more evident in the 3034 genes that have conserved APA events between rice and Arabidopsis. PMID:21813626

A detailed gene expression study of the Miscanthus genus reveals changes in the transcriptome associated with the rejuvenation of spring rhizomes.

PubMed

Barling, Adam; Swaminathan, Kankshita; Mitros, Therese; James, Brandon T; Morris, Juliette; Ngamboma, Ornella; Hall, Megan C; Kirkpatrick, Jessica; Alabady, Magdy; Spence, Ashley K; Hudson, Matthew E; Rokhsar, Daniel S; Moose, Stephen P

2013-12-09

The Miscanthus genus of perennial C4 grasses contains promising biofuel crops for temperate climates. However, few genomic resources exist for Miscanthus, which limits understanding of its interesting biology and future genetic improvement. A comprehensive catalog of expressed sequences were generated from a variety of Miscanthus species and tissue types, with an emphasis on characterizing gene expression changes in spring compared to fall rhizomes. Illumina short read sequencing technology was used to produce transcriptome sequences from different tissues and organs during distinct developmental stages for multiple Miscanthus species, including Miscanthus sinensis, Miscanthus sacchariflorus, and their interspecific hybrid Miscanthus × giganteus. More than fifty billion base-pairs of Miscanthus transcript sequence were produced. Overall, 26,230 Sorghum gene models (i.e., ~ 96% of predicted Sorghum genes) had at least five Miscanthus reads mapped to them, suggesting that a large portion of the Miscanthus transcriptome is represented in this dataset. The Miscanthus × giganteus data was used to identify genes preferentially expressed in a single tissue, such as the spring rhizome, using Sorghum bicolor as a reference. Quantitative real-time PCR was used to verify examples of preferential expression predicted via RNA-Seq. Contiguous consensus transcript sequences were assembled for each species and annotated using InterProScan. Sequences from the assembled transcriptome were used to amplify genomic segments from a doubled haploid Miscanthus sinensis and from Miscanthus × giganteus to further disentangle the allelic and paralogous variations in genes. This large expressed sequence tag collection creates a valuable resource for the study of Miscanthus biology by providing detailed gene sequence information and tissue preferred expression patterns. We have successfully generated a database of transcriptome assemblies and demonstrated its use in the study of genes of interest. Analysis of gene expression profiles revealed biological pathways that exhibit altered regulation in spring compared to fall rhizomes, which are consistent with their different physiological functions. The expression profiles of the subterranean rhizome provides a better understanding of the biological activities of the underground stem structures that are essentials for perenniality and the storage or remobilization of carbon and nutrient resources.

Insights from the pollination drop proteome and the ovule transcriptome of Cephalotaxus at the time of pollination drop production

PubMed Central

Pirone-Davies, Cary; Prior, Natalie; von Aderkas, Patrick; Smith, Derek; Hardie, Darryl; Friedman, William E.; Mathews, Sarah

2016-01-01

Background and Aims Many gymnosperms produce an ovular secretion, the pollination drop, during reproduction. The drops serve as a landing site for pollen, but also contain a suite of ions and organic compounds, including proteins, that suggests diverse roles for the drop during pollination. Proteins in the drops of species of Chamaecyparis, Juniperus, Taxus, Pseudotsuga, Ephedra and Welwitschia are thought to function in the conversion of sugars, defence against pathogens, and pollen growth and development. To better understand gymnosperm pollination biology, the pollination drop proteomes of pollination drops from two species of Cephalotaxus have been characterized and an ovular transcriptome for C. sinensis has been assembled. Methods Mass spectrometry was used to identify proteins in the pollination drops of Cephalotaxus sinensis and C. koreana. RNA-sequencing (RNA-Seq) was employed to assemble a transcriptome and identify transcripts present in the ovules of C. sinensis at the time of pollination drop production. Key Results About 30 proteins were detected in the pollination drops of both species. Many of these have been detected in the drops of other gymnosperms and probably function in defence, polysaccharide metabolism and pollen tube growth. Other proteins appear to be unique to Cephalotaxus, and their putative functions include starch and callose degradation, among others. Together, the proteins appear either to have been secreted into the drop or to occur there due to breakdown of ovular cells during drop production. Ovular transcripts represent a wide range of gene ontology categories, and some may be involved in drop formation, ovule development and pollen–ovule interactions. Conclusions The proteome of Cephalotaxus pollination drops shares a number of components with those of other conifers and gnetophytes, including proteins for defence such as chitinases and for carbohydrate modification such as β-galactosidase. Proteins likely to be of intracellular origin, however, form a larger component of drops from Cephalotaxus than expected from studies of other conifers. This is consistent with the observation of nucellar breakdown during drop formation in Cephalotaxus. The transcriptome data provide a framework for understanding multiple metabolic processes that occur within the ovule and the pollination drop just before fertilization. They reveal the deep conservation of WUSCHEL expression in ovules and raise questions about whether any of the S-locus transcripts in Cephalotaxus ovules might be involved in pollen–ovule recognition. PMID:27045089

Uncovering the pathways underlying whole body regeneration in a chordate model, Botrylloides leachi using de novo transcriptome analysis.

PubMed

Zondag, Lisa E; Rutherford, Kim; Gemmell, Neil J; Wilson, Megan J

2016-02-16

Regenerative capacity differs greatly between animals. In vertebrates regenerative abilities are highly limited and tissue or organ specific. However the closest related chordate to the vertebrate clade, Botrylloides leachi, can undergo whole body regeneration (WBR). Therefore, research on WBR in B. leachi has focused on pathways known to be important for regeneration in vertebrates. To obtain a comprehensive vision of this unique process we have carried out the first de novo transcriptome sequencing for multiple stages of WBR occurring in B. leachi. The identified changes in gene expression during B. leachi WBR offer novel insights into this remarkable ability to regenerate. The transcriptome of B. leachi tissue undergoing WBR were analysed using differential gene expression, gene ontology and pathway analyses. We observed up-regulation in the expression of genes involved in wound healing and known developmental pathways including WNT, TGF-β and Notch, during the earliest stages of WBR. Later in WBR, the expression patterns in several pathways required for protein synthesis, biogenesis and the organisation of cellular components were up-regulated. While the genes expressed early on are characteristic of a necessary wound healing response to an otherwise lethal injury, the subsequent vast increase in protein synthesis conceivably sustains the reestablishment of the tissue complexity and body axis polarity within the regenerating zooid. We have, for the first time, provided a global overview of the genes and their corresponding pathways that are modulated during WBR in B. leachi.

Genomic and transcriptomic approaches to study immunology in cyprinids: What is next?

PubMed

Petit, Jules; David, Lior; Dirks, Ron; Wiegertjes, Geert F

2017-10-01

Accelerated by the introduction of Next-Generation Sequencing (NGS), a number of genomes of cyprinid fish species have been drafted, leading to a highly valuable collective resource of comparative genome information on cyprinids (Cyprinidae). In addition, NGS-based transcriptome analyses of different developmental stages, organs, or cell types, increasingly contribute to the understanding of complex physiological processes, including immune responses. Cyprinids are a highly interesting family because they comprise one of the most-diversified families of teleosts and because of their variation in ploidy level, with diploid, triploid, tetraploid, hexaploid and sometimes even octoploid species. The wealth of data obtained from NGS technologies provides both challenges and opportunities for immunological research, which will be discussed here. Correct interpretation of ploidy effects on immune responses requires knowledge of the degree of functional divergence between duplicated genes, which can differ even between closely-related cyprinid fish species. We summarize NGS-based progress in analysing immune responses and discuss the importance of respecting the presence of (multiple) duplicated gene sequences when performing transcriptome analyses for detailed understanding of complex physiological processes. Progressively, advances in NGS technology are providing workable methods to further elucidate the implications of gene duplication events and functional divergence of duplicates genes and proteins involved in immune responses in cyprinids. We conclude with discussing how future applications of NGS technologies and analysis methods could enhance immunological research and understanding. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Alterations in the Aedes aegypti Transcriptome during Infection with West Nile, Dengue and Yellow Fever Viruses

PubMed Central

Colpitts, Tonya M.; Cox, Jonathan; Vanlandingham, Dana L.; Feitosa, Fabiana M.; Cheng, Gong; Kurscheid, Sebastian; Wang, Penghua; Krishnan, Manoj N.; Higgs, Stephen; Fikrig, Erol

2011-01-01

West Nile (WNV), dengue (DENV) and yellow fever (YFV) viruses are (re)emerging, mosquito-borne flaviviruses that cause human disease and mortality worldwide. Alterations in mosquito gene expression common and unique to individual flaviviral infections are poorly understood. Here, we present a microarray analysis of the Aedes aegypti transcriptome over time during infection with DENV, WNV or YFV. We identified 203 mosquito genes that were ≥5-fold differentially up-regulated (DUR) and 202 genes that were ≥10-fold differentially down-regulated (DDR) during infection with one of the three flaviviruses. Comparative analysis revealed that the expression profile of 20 DUR genes and 15 DDR genes was quite similar between the three flaviviruses on D1 of infection, indicating a potentially conserved transcriptomic signature of flaviviral infection. Bioinformatics analysis revealed changes in expression of genes from diverse cellular processes, including ion binding, transport, metabolic processes and peptidase activity. We also demonstrate that virally-regulated gene expression is tissue-specific. The overexpression of several virally down-regulated genes decreased WNV infection in mosquito cells and Aedes aegypti mosquitoes. Among these, a pupal cuticle protein was shown to bind WNV envelope protein, leading to inhibition of infection in vitro and the prevention of lethal WNV encephalitis in mice. This work provides an extensive list of targets for controlling flaviviral infection in mosquitoes that may also be used to develop broad preventative and therapeutic measures for multiple flaviviruses. PMID:21909258

Alterations in the Aedes aegypti transcriptome during infection with West Nile, dengue and yellow fever viruses.

PubMed

Colpitts, Tonya M; Cox, Jonathan; Vanlandingham, Dana L; Feitosa, Fabiana M; Cheng, Gong; Kurscheid, Sebastian; Wang, Penghua; Krishnan, Manoj N; Higgs, Stephen; Fikrig, Erol

2011-09-01

West Nile (WNV), dengue (DENV) and yellow fever (YFV) viruses are (re)emerging, mosquito-borne flaviviruses that cause human disease and mortality worldwide. Alterations in mosquito gene expression common and unique to individual flaviviral infections are poorly understood. Here, we present a microarray analysis of the Aedes aegypti transcriptome over time during infection with DENV, WNV or YFV. We identified 203 mosquito genes that were ≥ 5-fold differentially up-regulated (DUR) and 202 genes that were ≥ 10-fold differentially down-regulated (DDR) during infection with one of the three flaviviruses. Comparative analysis revealed that the expression profile of 20 DUR genes and 15 DDR genes was quite similar between the three flaviviruses on D1 of infection, indicating a potentially conserved transcriptomic signature of flaviviral infection. Bioinformatics analysis revealed changes in expression of genes from diverse cellular processes, including ion binding, transport, metabolic processes and peptidase activity. We also demonstrate that virally-regulated gene expression is tissue-specific. The overexpression of several virally down-regulated genes decreased WNV infection in mosquito cells and Aedes aegypti mosquitoes. Among these, a pupal cuticle protein was shown to bind WNV envelope protein, leading to inhibition of infection in vitro and the prevention of lethal WNV encephalitis in mice. This work provides an extensive list of targets for controlling flaviviral infection in mosquitoes that may also be used to develop broad preventative and therapeutic measures for multiple flaviviruses.

Developing a 'personalome' for precision medicine: emerging methods that compute interpretable effect sizes from single-subject transcriptomes.

PubMed

Vitali, Francesca; Li, Qike; Schissler, A Grant; Berghout, Joanne; Kenost, Colleen; Lussier, Yves A

2017-12-18

The development of computational methods capable of analyzing -omics data at the individual level is critical for the success of precision medicine. Although unprecedented opportunities now exist to gather data on an individual's -omics profile ('personalome'), interpreting and extracting meaningful information from single-subject -omics remain underdeveloped, particularly for quantitative non-sequence measurements, including complete transcriptome or proteome expression and metabolite abundance. Conventional bioinformatics approaches have largely been designed for making population-level inferences about 'average' disease processes; thus, they may not adequately capture and describe individual variability. Novel approaches intended to exploit a variety of -omics data are required for identifying individualized signals for meaningful interpretation. In this review-intended for biomedical researchers, computational biologists and bioinformaticians-we survey emerging computational and translational informatics methods capable of constructing a single subject's 'personalome' for predicting clinical outcomes or therapeutic responses, with an emphasis on methods that provide interpretable readouts. (i) the single-subject analytics of the transcriptome shows the greatest development to date and, (ii) the methods were all validated in simulations, cross-validations or independent retrospective data sets. This survey uncovers a growing field that offers numerous opportunities for the development of novel validation methods and opens the door for future studies focusing on the interpretation of comprehensive 'personalomes' through the integration of multiple -omics, providing valuable insights into individual patient outcomes and treatments. © The Author 2017. Published by Oxford University Press.

A deep transcriptomic analysis of pod development in the vanilla orchid (Vanilla planifolia).

PubMed

Rao, Xiaolan; Krom, Nick; Tang, Yuhong; Widiez, Thomas; Havkin-Frenkel, Daphna; Belanger, Faith C; Dixon, Richard A; Chen, Fang

2014-11-07

Pods of the vanilla orchid (Vanilla planifolia) accumulate large amounts of the flavor compound vanillin (3-methoxy, 4-hydroxy-benzaldehyde) as a glucoside during the later stages of their development. At earlier stages, the developing seeds within the pod synthesize a novel lignin polymer, catechyl (C) lignin, in their coats. Genomic resources for determining the biosynthetic routes to these compounds and other flavor components in V. planifolia are currently limited. Using next-generation sequencing technologies, we have generated very large gene sequence datasets from vanilla pods at different times of development, and representing different tissue types, including the seeds, hairs, placental and mesocarp tissues. This developmental series was chosen as being the most informative for interrogation of pathways of vanillin and C-lignin biosynthesis in the pod and seed, respectively. The combined 454/Illumina RNA-seq platforms provide both deep sequence coverage and high quality de novo transcriptome assembly for this non-model crop species. The annotated sequence data provide a foundation for understanding multiple aspects of the biochemistry and development of the vanilla bean, as exemplified by the identification of candidate genes involved in lignin biosynthesis. Our transcriptome data indicate that C-lignin formation in the seed coat involves coordinate expression of monolignol biosynthetic genes with the exception of those encoding the caffeoyl coenzyme A 3-O-methyltransferase for conversion of caffeoyl to feruloyl moieties. This database provides a general resource for further studies on this important flavor species.

Prediction of constitutive A-to-I editing sites from human transcriptomes in the absence of genomic sequences

PubMed Central

2013-01-01

Background Adenosine-to-inosine (A-to-I) RNA editing is recognized as a cellular mechanism for generating both RNA and protein diversity. Inosine base pairs with cytidine during reverse transcription and therefore appears as guanosine during sequencing of cDNA. Current approaches of RNA editing identification largely depend on the comparison between transcriptomes and genomic DNA (gDNA) sequencing datasets from the same individuals, and it has been challenging to identify editing candidates from transcriptomes in the absence of gDNA information. Results We have developed a new strategy to accurately predict constitutive RNA editing sites from publicly available human RNA-seq datasets in the absence of relevant genomic sequences. Our approach establishes new parameters to increase the ability to map mismatches and to minimize sequencing/mapping errors and unreported genome variations. We identified 695 novel constitutive A-to-I editing sites that appear in clusters (named “editing boxes”) in multiple samples and which exhibit spatial and dynamic regulation across human tissues. Some of these editing boxes are enriched in non-repetitive regions lacking inverted repeat structures and contain an extremely high conversion frequency of As to Is. We validated a number of editing boxes in multiple human cell lines and confirmed that ADAR1 is responsible for the observed promiscuous editing events in non-repetitive regions, further expanding our knowledge of the catalytic substrate of A-to-I RNA editing by ADAR enzymes. Conclusions The approach we present here provides a novel way of identifying A-to-I RNA editing events by analyzing only RNA-seq datasets. This method has allowed us to gain new insights into RNA editing and should also aid in the identification of more constitutive A-to-I editing sites from additional transcriptomes. PMID:23537002

Optimizing Hybrid de Novo Transcriptome Assembly and Extending Genomic Resources for Giant Freshwater Prawns (Macrobrachium rosenbergii): The Identification of Genes and Markers Associated with Reproduction.

PubMed

Jung, Hyungtaek; Yoon, Byung-Ha; Kim, Woo-Jin; Kim, Dong-Wook; Hurwood, David A; Lyons, Russell E; Salin, Krishna R; Kim, Heui-Soo; Baek, Ilseon; Chand, Vincent; Mather, Peter B

2016-05-07

The giant freshwater prawn, Macrobrachium rosenbergii, a sexually dimorphic decapod crustacean is currently the world's most economically important cultured freshwater crustacean species. Despite its economic importance, there is currently a lack of genomic resources available for this species, and this has limited exploration of the molecular mechanisms that control the M. rosenbergii sex-differentiation system more widely in freshwater prawns. Here, we present the first hybrid transcriptome from M. rosenbergii applying RNA-Seq technologies directed at identifying genes that have potential functional roles in reproductive-related traits. A total of 13,733,210 combined raw reads (1720 Mbp) were obtained from Ion-Torrent PGM and 454 FLX. Bioinformatic analyses based on three state-of-the-art assemblers, the CLC Genomic Workbench, Trans-ABySS, and Trinity, that use single and multiple k-mer methods respectively, were used to analyse the data. The influence of multiple k-mers on assembly performance was assessed to gain insight into transcriptome assembly from short reads. After optimisation, de novo assembly resulted in 44,407 contigs with a mean length of 437 bp, and the assembled transcripts were further functionally annotated to detect single nucleotide polymorphisms and simple sequence repeat motifs. Gene expression analysis was also used to compare expression patterns from ovary and testis tissue libraries to identify genes with potential roles in reproduction and sex differentiation. The large transcript set assembled here represents the most comprehensive set of transcriptomic resources ever developed for reproduction traits in M. rosenbergii, and the large number of genetic markers predicted should constitute an invaluable resource for future genetic research studies on M. rosenbergii and can be applied more widely on other freshwater prawn species in the genus Macrobrachium.

Optimizing Hybrid de Novo Transcriptome Assembly and Extending Genomic Resources for Giant Freshwater Prawns (Macrobrachium rosenbergii): The Identification of Genes and Markers Associated with Reproduction

PubMed Central

Jung, Hyungtaek; Yoon, Byung-Ha; Kim, Woo-Jin; Kim, Dong-Wook; Hurwood, David A.; Lyons, Russell E.; Salin, Krishna R.; Kim, Heui-Soo; Baek, Ilseon; Chand, Vincent; Mather, Peter B.

2016-01-01

The giant freshwater prawn, Macrobrachium rosenbergii, a sexually dimorphic decapod crustacean is currently the world’s most economically important cultured freshwater crustacean species. Despite its economic importance, there is currently a lack of genomic resources available for this species, and this has limited exploration of the molecular mechanisms that control the M. rosenbergii sex-differentiation system more widely in freshwater prawns. Here, we present the first hybrid transcriptome from M. rosenbergii applying RNA-Seq technologies directed at identifying genes that have potential functional roles in reproductive-related traits. A total of 13,733,210 combined raw reads (1720 Mbp) were obtained from Ion-Torrent PGM and 454 FLX. Bioinformatic analyses based on three state-of-the-art assemblers, the CLC Genomic Workbench, Trans-ABySS, and Trinity, that use single and multiple k-mer methods respectively, were used to analyse the data. The influence of multiple k-mers on assembly performance was assessed to gain insight into transcriptome assembly from short reads. After optimisation, de novo assembly resulted in 44,407 contigs with a mean length of 437 bp, and the assembled transcripts were further functionally annotated to detect single nucleotide polymorphisms and simple sequence repeat motifs. Gene expression analysis was also used to compare expression patterns from ovary and testis tissue libraries to identify genes with potential roles in reproduction and sex differentiation. The large transcript set assembled here represents the most comprehensive set of transcriptomic resources ever developed for reproduction traits in M. rosenbergii, and the large number of genetic markers predicted should constitute an invaluable resource for future genetic research studies on M. rosenbergii and can be applied more widely on other freshwater prawn species in the genus Macrobrachium. PMID:27164098

Next-generation sequencing (NGS) transcriptomes reveal association of multiple genes and pathways contributing to secondary metabolites accumulation in tuberous roots of Aconitum heterophyllum Wall.

PubMed

Pal, Tarun; Malhotra, Nikhil; Chanumolu, Sree Krishna; Chauhan, Rajinder Singh

2015-07-01

The transcriptomes of Aconitum heterophyllum were assembled and characterized for the first time to decipher molecular components contributing to biosynthesis and accumulation of metabolites in tuberous roots. Aconitum heterophyllum Wall., popularly known as Atis, is a high-value medicinal herb of North-Western Himalayas. No information exists as of today on genetic factors contributing to the biosynthesis of secondary metabolites accumulating in tuberous roots, thereby, limiting genetic interventions towards genetic improvement of A. heterophyllum. Illumina paired-end sequencing followed by de novo assembly yielded 75,548 transcripts for root transcriptome and 39,100 transcripts for shoot transcriptome with minimum length of 200 bp. Biological role analysis of root versus shoot transcriptomes assigned 27,596 and 16,604 root transcripts; 12,340 and 9398 shoot transcripts into gene ontology and clusters of orthologous group, respectively. KEGG pathway mapping assigned 37 and 31 transcripts onto starch-sucrose metabolism while 329 and 341 KEGG orthologies associated with transcripts were found to be involved in biosynthesis of various secondary metabolites for root and shoot transcriptomes, respectively. In silico expression profiling of the mevalonate/2-C-methyl-D-erythritol 4-phosphate (non-mevalonate) pathway genes for aconites biosynthesis revealed 4 genes HMGR (3-hydroxy-3-methylglutaryl-CoA reductase), MVK (mevalonate kinase), MVDD (mevalonate diphosphate decarboxylase) and HDS (1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate synthase) with higher expression in root transcriptome compared to shoot transcriptome suggesting their key role in biosynthesis of aconite alkaloids. Five genes, GMPase (geranyl diphosphate mannose pyrophosphorylase), SHAGGY, RBX1 (RING-box protein 1), SRF receptor kinases and β-amylase, implicated in tuberous root formation in other plant species showed higher levels of expression in tuberous roots compared to shoots. A total of 15,487 transcription factors belonging to bHLH, MYB, bZIP families and 399 ABC transporters which regulate biosynthesis and accumulation of bioactive compounds were identified in root and shoot transcriptomes. The expression of 5 ABC transporters involved in tuberous root development was validated by quantitative PCR analysis. Network connectivity diagrams were drawn for starch-sucrose metabolism and isoquinoline alkaloid biosynthesis associated with tuberous root growth and secondary metabolism, respectively, in root transcriptome of A. heterophyllum. The current endeavor will be of practical importance in planning a suitable genetic intervention strategy for the improvement of A. heterophyllum.

Inflammatory Mediators Alter the Astrocyte Transcriptome and Calcium Signaling Elicited by Multiple G-Protein-Coupled Receptors

PubMed Central

Hamby, Mary E.; Coppola, Giovanni; Ao, Yan; Geschwind, Daniel H.; Khakh, Baljit S.; Sofroniew, Michael V.

2012-01-01

Inflammation features in CNS disorders such as stroke, trauma, neurodegeneration, infection, and autoimmunity in which astrocytes play critical roles. To elucidate how inflammatory mediators alter astrocyte functions, we examined effects of transforming growth factor-β1 (TGF-β1), lipopolysaccharide (LPS), and interferon-gamma (IFNγ), alone and in combination, on purified, mouse primary cortical astrocyte cultures. We used microarrays to conduct whole-genome expression profiling, and measured calcium signaling, which is implicated in mediating dynamic astrocyte functions. Combinatorial exposure to TGF-β1, LPS, and IFNγ significantly modulated astrocyte expression of >6800 gene probes, including >380 synergistic changes not predicted by summing individual treatment effects. Bioinformatic analyses revealed significantly and markedly upregulated molecular networks and pathways associated in particular with immune signaling and regulation of cell injury, death, growth, and proliferation. Highly regulated genes included chemokines, growth factors, enzymes, channels, transporters, and intercellular and intracellular signal transducers. Notably, numerous genes for G-protein-coupled receptors (GPCRs) and G-protein effectors involved in calcium signaling were significantly regulated, mostly down (for example, Cxcr4, Adra2a, Ednra, P2y1, Gnao1, Gng7), but some up (for example, P2y14, P2y6, Ccrl2, Gnb4). We tested selected cases and found that changes in GPCR gene expression were accompanied by significant, parallel changes in astrocyte calcium signaling evoked by corresponding GPCR-specific ligands. These findings identify pronounced changes in the astrocyte transcriptome induced by TGF-β1, LPS, and IFNγ, and show that these inflammatory stimuli upregulate astrocyte molecular networks associated with immune- and injury-related functions and significantly alter astrocyte calcium signaling stimulated by multiple GPCRs. PMID:23077035

Genetic variation and gene expression across multiple tissues and developmental stages in a non-human primate

PubMed Central

Jasinska, Anna J.; Zelaya, Ivette; Service, Susan K.; Peterson, Christine B.; Cantor, Rita M.; Choi, Oi-Wa; DeYoung, Joseph; Eskin, Eleazar; Fairbanks, Lynn A.; Fears, Scott; Furterer, Allison E.; Huang, Yu S.; Ramensky, Vasily; Schmitt, Christopher A.; Svardal, Hannes; Jorgensen, Matthew J.; Kaplan, Jay R.; Villar, Diego; Aken, Bronwen L.; Flicek, Paul; Nag, Rishi; Wong, Emily S.; Blangero, John; Dyer, Thomas D.; Bogomolov, Marina; Benjamini, Yoav; Weinstock, George M.; Dewar, Ken; Sabatti, Chiara; Wilson, Richard K.; Jentsch, J. David; Warren, Wesley; Coppola, Giovanni; Woods, Roger P.; Freimer, Nelson B.

2017-01-01

By analyzing multi-tissue gene expression and genome-wide genetic variation data in samples from a vervet monkey pedigree, we generated a transcriptome resource and produced the first catalogue of expression quantitative trait loci (eQTLs) in a non-human primate model. This catalogue contains more genome-wide significant eQTLs, per sample, than comparable human resources, and reveals sex and age-related expression patterns. Findings include a master regulatory locus that likely plays a role in immune function, and a locus regulating hippocampal long non-coding RNAs (lncRNAs), whose expression correlates with hippocampal volume. This resource will facilitate genetic investigation of quantitative traits, including brain and behavioral phenotypes relevant to neuropsychiatric disorders. PMID:29083405

Reduced representation approaches to interrogate genome diversity in large repetitive plant genomes.

PubMed

Hirsch, Cory D; Evans, Joseph; Buell, C Robin; Hirsch, Candice N

2014-07-01

Technology and software improvements in the last decade now provide methodologies to access the genome sequence of not only a single accession, but also multiple accessions of plant species. This provides a means to interrogate species diversity at the genome level. Ample diversity among accessions in a collection of species can be found, including single-nucleotide polymorphisms, insertions and deletions, copy number variation and presence/absence variation. For species with small, non-repetitive rich genomes, re-sequencing of query accessions is robust, highly informative, and economically feasible. However, for species with moderate to large sized repetitive-rich genomes, technical and economic barriers prevent en masse genome re-sequencing of accessions. Multiple approaches to access a focused subset of loci in species with larger genomes have been developed, including reduced representation sequencing, exome capture and transcriptome sequencing. Collectively, these approaches have enabled interrogation of diversity on a genome scale for large plant genomes, including crop species important to worldwide food security. © The Author 2014. Published by Oxford University Press. All rights reserved. For permissions, please email: journals.permissions@oup.com.

De novo assembly and analysis of the Artemisia argyi transcriptome and identification of genes involved in terpenoid biosynthesis.

PubMed

Liu, Miaomiao; Zhu, Jinhang; Wu, Shengbing; Wang, Chenkai; Guo, Xingyi; Wu, Jiawen; Zhou, Meiqi

2018-04-11

Artemisia argyi Lev. et Vant. (A. argyi) is widely utilized for moxibustion in Chinese medicine, and the mechanism underlying terpenoid biosynthesis in its leaves is suggested to play an important role in its medicinal use. However, the A. argyi transcriptome has not been sequenced. Herein, we performed RNA sequencing for A. argyi leaf, root and stem tissues to identify as many as possible of the transcribed genes. In total, 99,807 unigenes were assembled by analysing the expression profiles generated from the three tissue types, and 67,446 of those unigenes were annotated in public databases. We further performed differential gene expression analysis to compare leaf tissue with the other two tissue types and identified numerous genes that were specifically expressed or up-regulated in leaf tissue. Specifically, we identified multiple genes encoding significant enzymes or transcription factors related to terpenoid synthesis. This study serves as a valuable resource for transcriptome information, as many transcribed genes related to terpenoid biosynthesis were identified in the A. argyi transcriptome, providing a functional genomic basis for additional studies on molecular mechanisms underlying the medicinal use of A. argyi.

RNAseq versus genome-predicted transcriptomes: a large population of novel transcripts identified in an Illumina-454 Hydra transcriptome.

PubMed

Wenger, Yvan; Galliot, Brigitte

2013-03-25

Evolutionary studies benefit from deep sequencing technologies that generate genomic and transcriptomic sequences from a variety of organisms. Genome sequencing and RNAseq have complementary strengths. In this study, we present the assembly of the most complete Hydra transcriptome to date along with a comparative analysis of the specific features of RNAseq and genome-predicted transcriptomes currently available in the freshwater hydrozoan Hydra vulgaris. To produce an accurate and extensive Hydra transcriptome, we combined Illumina and 454 Titanium reads, giving the primacy to Illumina over 454 reads to correct homopolymer errors. This strategy yielded an RNAseq transcriptome that contains 48'909 unique sequences including splice variants, representing approximately 24'450 distinct genes. Comparative analysis to the available genome-predicted transcriptomes identified 10'597 novel Hydra transcripts that encode 529 evolutionarily-conserved proteins. The annotation of 170 human orthologs points to critical functions in protein biosynthesis, FGF and TOR signaling, vesicle transport, immunity, cell cycle regulation, cell death, mitochondrial metabolism, transcription and chromatin regulation. However, a majority of these novel transcripts encodes short ORFs, at least 767 of them corresponding to pseudogenes. This RNAseq transcriptome also lacks 11'270 predicted transcripts that correspond either to silent genes or to genes expressed below the detection level of this study. We established a simple and powerful strategy to combine Illumina and 454 reads and we produced, with genome assistance, an extensive and accurate Hydra transcriptome. The comparative analysis of the RNAseq transcriptome with genome-predicted transcriptomes lead to the identification of large populations of novel as well as missing transcripts that might reflect Hydra-specific evolutionary events.

RNAseq versus genome-predicted transcriptomes: a large population of novel transcripts identified in an Illumina-454 Hydra transcriptome

PubMed Central

2013-01-01

Background Evolutionary studies benefit from deep sequencing technologies that generate genomic and transcriptomic sequences from a variety of organisms. Genome sequencing and RNAseq have complementary strengths. In this study, we present the assembly of the most complete Hydra transcriptome to date along with a comparative analysis of the specific features of RNAseq and genome-predicted transcriptomes currently available in the freshwater hydrozoan Hydra vulgaris. Results To produce an accurate and extensive Hydra transcriptome, we combined Illumina and 454 Titanium reads, giving the primacy to Illumina over 454 reads to correct homopolymer errors. This strategy yielded an RNAseq transcriptome that contains 48’909 unique sequences including splice variants, representing approximately 24’450 distinct genes. Comparative analysis to the available genome-predicted transcriptomes identified 10’597 novel Hydra transcripts that encode 529 evolutionarily-conserved proteins. The annotation of 170 human orthologs points to critical functions in protein biosynthesis, FGF and TOR signaling, vesicle transport, immunity, cell cycle regulation, cell death, mitochondrial metabolism, transcription and chromatin regulation. However, a majority of these novel transcripts encodes short ORFs, at least 767 of them corresponding to pseudogenes. This RNAseq transcriptome also lacks 11’270 predicted transcripts that correspond either to silent genes or to genes expressed below the detection level of this study. Conclusions We established a simple and powerful strategy to combine Illumina and 454 reads and we produced, with genome assistance, an extensive and accurate Hydra transcriptome. The comparative analysis of the RNAseq transcriptome with genome-predicted transcriptomes lead to the identification of large populations of novel as well as missing transcripts that might reflect Hydra-specific evolutionary events. PMID:23530871

Circular RNA profiling reveals that circular RNAs from ANXA2 can be used as new biomarkers for multiple sclerosis.

PubMed

Iparraguirre, Leire; Muñoz-Culla, Maider; Prada-Luengo, Iñigo; Castillo-Triviño, Tamara; Olascoaga, Javier; Otaegui, David

2017-09-15

Multiple sclerosis is an autoimmune disease, with higher prevalence in women, in whom the immune system is dysregulated. This dysregulation has been shown to correlate with changes in transcriptome expression as well as in gene-expression regulators, such as non-coding RNAs (e.g. microRNAs). Indeed, some of these have been suggested as biomarkers for multiple sclerosis even though few biomarkers have reached the clinical practice. Recently, a novel family of non-coding RNAs, circular RNAs, has emerged as a new player in the complex network of gene-expression regulation. MicroRNA regulation function through a 'sponge system' and a RNA splicing regulation function have been proposed for the circular RNAs. This regulating role together with their high stability in biofluids makes them seemingly good candidates as biomarkers. Given the dysregulation of both protein-coding and non-coding transcriptome that have been reported in multiple sclerosis patients, we hypothesised that circular RNA expression may also be altered. Therefore, we carried out expression profiling of 13.617 circular RNAs in peripheral blood leucocytes from multiple sclerosis patients and healthy controls finding 406 differentially expressed (P-value < 0.05, Fold change > 1.5) and demonstrate after validation that, circ_0005402 and circ_0035560 are underexpressed in multiple sclerosis patients and could be used as biomarkers of the disease. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Tipping the balance of RNA stability by 3' editing of the transcriptome.

PubMed

Chung, Christina Z; Seidl, Lauren E; Mann, Mitchell R; Heinemann, Ilka U

2017-11-01

The regulation of active microRNAs (miRNAs) and maturation of messenger RNAs (mRNAs) that are competent for translation is a crucial point in the control of all cellular processes, with established roles in development and differentiation. Terminal nucleotidyltransferases (TNTases) are potent regulators of RNA metabolism. TNTases promote the addition of single or multiple nucleotides to an RNA transcript that can rapidly alter transcript stability. The well-known polyadenylation promotes transcript stability while the newly discovered but ubiquitious 3'-end polyuridylation marks RNA for degradation. Monoadenylation and uridylation are essential control mechanisms balancing mRNA and miRNA homeostasis. This review discusses the multiple functions of non-canonical TNTases, focusing on their substrate range, biological functions, and evolution. TNTases directly control mRNA and miRNA levels, with diverse roles in transcriptome stabilization, maturation, silencing, or degradation. We will summarize the current state of knowledge on non-canonical nucleotidyltransferases and their function in regulating miRNA and mRNA metabolism. We will review the discovery of uridylation as an RNA degradation pathway and discuss the evolution of nucleotidyltransferases along with their use in RNA labeling and future applications as therapeutic targets. The biochemically and evolutionarily highly related adenylyl- and uridylyltransferases play antagonizing roles in the cell. In general, RNA adenylation promotes stability, while uridylation marks RNA for degradation. Uridylyltransferases evolved from adenylyltransferases in multiple independent evolutionary events by the insertion of a histidine residue into the active site, altering nucleotide, but not RNA specificity. Understanding the mechanisms regulating RNA stability in the cell and controlling the transcriptome is essential for efforts aiming to influence cellular fate. Selectively enhancing or reducing RNA stability allows for alterations in the transcriptome, proteome, and downstream cellular processes. Genetic, biochemical, and clinical data suggest TNTases are potent targets for chemotherapeutics and have been exploited for RNA labeling applications. This article is part of a Special Issue entitled "Biochemistry of Synthetic Biology - Recent Developments" Guest Editor: Dr. Ilka Heinemann and Dr. Patrick O'Donoghue. Copyright © 2017 Elsevier B.V. All rights reserved.

Transcriptomic profile induced in bone marrow mesenchymal stromal cells after interaction with multiple myeloma cells: implications in myeloma progression and myeloma bone disease

PubMed Central

Garcia-Gomez, Antonio; Las Rivas, Javier De; Ocio, Enrique M.; Díaz-Rodríguez, Elena; Montero, Juan C.; Martín, Montserrat; Blanco, Juan F.; Sanchez-Guijo, Fermín M.; Pandiella, Atanasio; San Miguel, Jesús F.; Garayoa, Mercedes

2014-01-01

Despite evidence about the implication of the bone marrow (BM) stromal microenvironment in multiple myeloma (MM) cell growth and survival, little is known about the effects of myelomatous cells on BM stromal cells. Mesenchymal stromal cells (MSCs) from healthy donors (dMSCs) or myeloma patients (pMSCs) were co-cultured with the myeloma cell line MM.1S, and the transcriptomic profile of MSCs induced by this interaction was analyzed. Deregulated genes after co-culture common to both d/pMSCs revealed functional involvement in tumor microenvironment cross-talk, myeloma growth induction and drug resistance, angiogenesis and signals for osteoclast activation and osteoblast inhibition. Additional genes induced by co-culture were exclusively deregulated in pMSCs and predominantly associated to RNA processing, the ubiquitine-proteasome pathway, cell cycle regulation, cellular stress and non-canonical Wnt signaling. The upregulated expression of five genes after co-culture (CXCL1, CXCL5 and CXCL6 in d/pMSCs, and Neuregulin 3 and Norrie disease protein exclusively in pMSCs) was confirmed, and functional in vitro assays revealed putative roles in MM pathophysiology. The transcriptomic profile of pMSCs co-cultured with myeloma cells may better reflect that of MSCs in the BM of myeloma patients, and provides new molecular insights to the contribution of these cells to MM pathophysiology and to myeloma bone disease. PMID:25268740

Genome and transcriptome of the porcine whipworm Trichuris suis

PubMed Central

Jex, Aaron R.; Nejsum, Peter; Schwarz, Erich M.; Hu, Li; Young, Neil D.; Hall, Ross S.; Korhonen, Pasi K.; Liao, Shengguang; Thamsborg, Stig; Xia, Jinquan; Xu, Pengwei; Wang, Shaowei; Scheerlinck, Jean-Pierre Y.; Hofmann, Andreas; Sternberg, Paul W.; Wang, Jun; Gasser, Robin B.

2014-01-01

Trichuris (whipworm) infects 1 billion people worldwide, and causes a disease (trichuriasis) that results in major socioeconomic losses in both humans and pigs. Trichuriasis relates to an inflammation of the large intestine manifested in bloody diarrhoea, and chronic disease can cause malnourishment and stunting in children. Paradoxically, Trichuris of pigs has shown substantial promise as a treatment for human autoimmune disorders, including inflammatory bowel disease (IBD) and multiple sclerosis (MS). Here, we report ~80 megabase (Mb) draft assemblies of the genomes of adult male and female T. suis, and explore stage-, sex- and tissue-specific transcription of messenger and small non-coding RNAs. PMID:24929829

Genome and transcriptome of the porcine whipworm Trichuris suis.

PubMed

Jex, Aaron R; Nejsum, Peter; Schwarz, Erich M; Hu, Li; Young, Neil D; Hall, Ross S; Korhonen, Pasi K; Liao, Shengguang; Thamsborg, Stig; Xia, Jinquan; Xu, Pengwei; Wang, Shaowei; Scheerlinck, Jean-Pierre Y; Hofmann, Andreas; Sternberg, Paul W; Wang, Jun; Gasser, Robin B

2014-07-01

Trichuris (whipworm) infects 1 billion people worldwide and causes a disease (trichuriasis) that results in major socioeconomic losses in both humans and pigs. Trichuriasis relates to an inflammation of the large intestine manifested in bloody diarrhea, and chronic disease can cause malnourishment and stunting in children. Paradoxically, Trichuris of pigs has shown substantial promise as a treatment for human autoimmune disorders, including inflammatory bowel disease (IBD) and multiple sclerosis. Here we report whole-genome sequencing at ∼140-fold coverage of adult male and female T. suis and ∼80-Mb draft assemblies. We explore stage-, sex- and tissue-specific transcription of mRNAs and small noncoding RNAs.

Evidence for light perception in a bioluminescent organ

PubMed Central

Tong, Deyan; Rozas, Natalia S.; Oakley, Todd H.; Mitchell, Jane; Colley, Nansi J.; McFall-Ngai, Margaret J.

2009-01-01

Here we show that bioluminescent organs of the squid Euprymna scolopes possess the molecular, biochemical, and physiological capability for light detection. Transcriptome analyses revealed expression of genes encoding key visual transduction proteins in light-organ tissues, including the same isoform of opsin that occurs in the retina. Electroretinograms demonstrated that the organ responds physiologically to light, and immunocytochemistry experiments localized multiple proteins of visual transduction cascades to tissues housing light-producing bacterial symbionts. These data provide evidence that the light-organ tissues harboring the symbionts serve as extraocular photoreceptors, with the potential to perceive directly the bioluminescence produced by their bacterial partners. PMID:19509343

Metformin-Induced Changes of the Coding Transcriptome and Non-Coding RNAs in the Livers of Non-Alcoholic Fatty Liver Disease Mice.

PubMed

Guo, Jun; Zhou, Yuan; Cheng, Yafen; Fang, Weiwei; Hu, Gang; Wei, Jie; Lin, Yajun; Man, Yong; Guo, Lixin; Sun, Mingxiao; Cui, Qinghua; Li, Jian

2018-01-01

Recent studies have suggested that changes in non-coding mRNA play a key role in the progression of non-alcoholic fatty liver disease (NAFLD). Metformin is now recommended and effective for the treatment of NAFLD. We hope the current analyses of the non-coding mRNA transcriptome will provide a better presentation of the potential roles of mRNAs and long non-coding RNAs (lncRNAs) that underlie NAFLD and metformin intervention. The present study mainly analysed changes in the coding transcriptome and non-coding RNAs after the application of a five-week metformin intervention. Liver samples from three groups of mice were harvested for transcriptome profiling, which covered mRNA, lncRNA, microRNA (miRNA) and circular RNA (circRNA), using a microarray technique. A systematic alleviation of high-fat diet (HFD)-induced transcriptome alterations by metformin was observed. The metformin treatment largely reversed the correlations with diabetes-related pathways. Our analysis also suggested interaction networks between differentially expressed lncRNAs and known hepatic disease genes and interactions between circRNA and their disease-related miRNA partners. Eight HFD-responsive lncRNAs and three metformin-responsive lncRNAs were noted due to their widespread associations with disease genes. Moreover, seven miRNAs that interacted with multiple differentially expressed circRNAs were highlighted because they were likely to be associated with metabolic or liver diseases. The present study identified novel changes in the coding transcriptome and non-coding RNAs in the livers of NAFLD mice after metformin treatment that might shed light on the underlying mechanism by which metformin impedes the progression of NAFLD. © 2018 The Author(s). Published by S. Karger AG, Basel.

Optimized Probe Masking for Comparative Transcriptomics of Closely Related Species

PubMed Central

Poeschl, Yvonne; Delker, Carolin; Trenner, Jana; Ullrich, Kristian Karsten; Quint, Marcel; Grosse, Ivo

2013-01-01

Microarrays are commonly applied to study the transcriptome of specific species. However, many available microarrays are restricted to model organisms, and the design of custom microarrays for other species is often not feasible. Hence, transcriptomics approaches of non-model organisms as well as comparative transcriptomics studies among two or more species often make use of cost-intensive RNAseq studies or, alternatively, by hybridizing transcripts of a query species to a microarray of a closely related species. When analyzing these cross-species microarray expression data, differences in the transcriptome of the query species can cause problems, such as the following: (i) lower hybridization accuracy of probes due to mismatches or deletions, (ii) probes binding multiple transcripts of different genes, and (iii) probes binding transcripts of non-orthologous genes. So far, methods for (i) exist, but these neglect (ii) and (iii). Here, we propose an approach for comparative transcriptomics addressing problems (i) to (iii), which retains only transcript-specific probes binding transcripts of orthologous genes. We apply this approach to an Arabidopsis lyrata expression data set measured on a microarray designed for Arabidopsis thaliana, and compare it to two alternative approaches, a sequence-based approach and a genomic DNA hybridization-based approach. We investigate the number of retained probe sets, and we validate the resulting expression responses by qRT-PCR. We find that the proposed approach combines the benefit of sequence-based stringency and accuracy while allowing the expression analysis of much more genes than the alternative sequence-based approach. As an added benefit, the proposed approach requires probes to detect transcripts of orthologous genes only, which provides a superior base for biological interpretation of the measured expression responses. PMID:24260119

Comprehensive Assessments of RNA-seq by the SEQC Consortium: FDA-Led Efforts Advance Precision Medicine.

PubMed

Xu, Joshua; Gong, Binsheng; Wu, Leihong; Thakkar, Shraddha; Hong, Huixiao; Tong, Weida

2016-03-15

Studies on gene expression in response to therapy have led to the discovery of pharmacogenomics biomarkers and advances in precision medicine. Whole transcriptome sequencing (RNA-seq) is an emerging tool for profiling gene expression and has received wide adoption in the biomedical research community. However, its value in regulatory decision making requires rigorous assessment and consensus between various stakeholders, including the research community, regulatory agencies, and industry. The FDA-led SEquencing Quality Control (SEQC) consortium has made considerable progress in this direction, and is the subject of this review. Specifically, three RNA-seq platforms (Illumina HiSeq, Life Technologies SOLiD, and Roche 454) were extensively evaluated at multiple sites to assess cross-site and cross-platform reproducibility. The results demonstrated that relative gene expression measurements were consistently comparable across labs and platforms, but not so for the measurement of absolute expression levels. As part of the quality evaluation several studies were included to evaluate the utility of RNA-seq in clinical settings and safety assessment. The neuroblastoma study profiled tumor samples from 498 pediatric neuroblastoma patients by both microarray and RNA-seq. RNA-seq offers more utilities than microarray in determining the transcriptomic characteristics of cancer. However, RNA-seq and microarray-based models were comparable in clinical endpoint prediction, even when including additional features unique to RNA-seq beyond gene expression. The toxicogenomics study compared microarray and RNA-seq profiles of the liver samples from rats exposed to 27 different chemicals representing multiple toxicity modes of action. Cross-platform concordance was dependent on chemical treatment and transcript abundance. Though both RNA-seq and microarray are suitable for developing gene expression based predictive models with comparable prediction performance, RNA-seq offers advantages over microarray in profiling genes with low expression. The rat BodyMap study provided a comprehensive rat transcriptomic body map by performing RNA-Seq on 320 samples from 11 organs in either sex of juvenile, adolescent, adult and aged Fischer 344 rats. Lastly, the transferability study demonstrated that signature genes of predictive models are reciprocally transferable between microarray and RNA-seq data for model development using a comprehensive approach with two large clinical data sets. This result suggests continued usefulness of legacy microarray data in the coming RNA-seq era. In conclusion, the SEQC project enhances our understanding of RNA-seq and provides valuable guidelines for RNA-seq based clinical application and safety evaluation to advance precision medicine.

Developmental Transcriptome for a Facultatively Eusocial Bee, Megalopta genalis

PubMed Central

Jones, Beryl M.; Wcislo, William T.; Robinson, Gene E.

2015-01-01

Transcriptomes provide excellent foundational resources for mechanistic and evolutionary analyses of complex traits. We present a developmental transcriptome for the facultatively eusocial bee Megalopta genalis, which represents a potential transition point in the evolution of eusociality. A de novo transcriptome assembly of Megalopta genalis was generated using paired-end Illumina sequencing and the Trinity assembler. Males and females of all life stages were aligned to this transcriptome for analysis of gene expression profiles throughout development. Gene Ontology analysis indicates that stage-specific genes are involved in ion transport, cell–cell signaling, and metabolism. A number of distinct biological processes are upregulated in each life stage, and transitions between life stages involve shifts in dominant functional processes, including shifts from transcriptional regulation in embryos to metabolism in larvae, and increased lipid metabolism in adults. We expect that this transcriptome will provide a useful resource for future analyses to better understand the molecular basis of the evolution of eusociality and, more generally, phenotypic plasticity. PMID:26276382

Developmental Transcriptome for a Facultatively Eusocial Bee, Megalopta genalis.

PubMed

Jones, Beryl M; Wcislo, William T; Robinson, Gene E

2015-08-14

Transcriptomes provide excellent foundational resources for mechanistic and evolutionary analyses of complex traits. We present a developmental transcriptome for the facultatively eusocial bee Megalopta genalis, which represents a potential transition point in the evolution of eusociality. A de novo transcriptome assembly of Megalopta genalis was generated using paired-end Illumina sequencing and the Trinity assembler. Males and females of all life stages were aligned to this transcriptome for analysis of gene expression profiles throughout development. Gene Ontology analysis indicates that stage-specific genes are involved in ion transport, cell-cell signaling, and metabolism. A number of distinct biological processes are upregulated in each life stage, and transitions between life stages involve shifts in dominant functional processes, including shifts from transcriptional regulation in embryos to metabolism in larvae, and increased lipid metabolism in adults. We expect that this transcriptome will provide a useful resource for future analyses to better understand the molecular basis of the evolution of eusociality and, more generally, phenotypic plasticity. Copyright © 2015 Jones et al.

Transgenerational Epigenetic Programming of the Embryonic Testis Transcriptome

PubMed Central

Anway, Matthew D.; Rekow, Stephen S.; Skinner, Michael K.

2008-01-01

Embryonic exposure to the endocrine disruptor vinclozolin during gonadal sex determination appears to promote an epigenetic reprogramming of the male germ-line that is associated with transgenerational adult onset disease states. Transgenerational effects on the embryonic day 16 (E16) testis demonstrated reproducible changes in the testis transcriptome for multiple generations (F1-F3). The expression of 196 genes were found to be influenced, with the majority of gene expression being decreased or silenced. Dramatic changes in the gene expression of methyltransferases during gonadal sex determination were observed in the F1 and F2 vinclozolin generation (E16) embryonic testis, but the majority returned to control generation levels by the F3 generation. The most dramatic effects were on the germ-line associated Dnmt3A and Dnmt3L isoforms. Observations demonstrate that an embryonic exposure to vinclozolin appears to promote an epigenetic reprogramming of the male germ-line that correlates with transgenerational alterations in the testis transcriptome in subsequent generations. PMID:18042343

Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures

PubMed Central

Park, Paul J.; Fuchs, Robert; Wei, Lai; Jorgensen, Brian G.; Redelman, Doug; Ward, Sean M.; Sanders, Kenton M.

2017-01-01

Transcriptome-scale data can reveal essential clues into understanding the underlying molecular mechanisms behind specific cellular functions and biological processes. Transcriptomics is a continually growing field of research utilized in biomarker discovery. The transcriptomic profile of interstitial cells of Cajal (ICC), which serve as slow-wave electrical pacemakers for gastrointestinal (GI) smooth muscle, has yet to be uncovered. Using copGFP-labeled ICC mice and flow cytometry, we isolated ICC populations from the murine small intestine and colon and obtained their transcriptomes. In analyzing the transcriptome, we identified a unique set of ICC-restricted markers including transcription factors, epigenetic enzymes/regulators, growth factors, receptors, protein kinases/phosphatases, and ion channels/transporters. This analysis provides new and unique insights into the cellular and biological functions of ICC in GI physiology. Additionally, we constructed an interactive ICC genome browser (http://med.unr.edu/physio/transcriptome) based on the UCSC genome database. To our knowledge, this is the first online resource that provides a comprehensive library of all known genetic transcripts expressed in primary ICC. Our genome browser offers a new perspective into the alternative expression of genes in ICC and provides a valuable reference for future functional studies. PMID:28426719

Character trees from transcriptome data: Origin and individuation of morphological characters and the so-called "species signal".

PubMed

Musser, Jacob M; Wagner, Günter P

2015-11-01

We elaborate a framework for investigating the evolutionary history of morphological characters. We argue that morphological character trees generated by phylogenetic analysis of transcriptomes provide a useful tool for identifying causal gene expression differences underlying the development and evolution of morphological characters. They also enable rigorous testing of different models of morphological character evolution and origination, including the hypothesis that characters originate via divergence of repeated ancestral characters. Finally, morphological character trees provide evidence that character transcriptomes undergo concerted evolution. We argue that concerted evolution of transcriptomes can explain the so-called "species signal" found in several recent comparative transcriptome studies. The species signal is the phenomenon that transcriptomes cluster by species rather than character type, even though the characters are older than the respective species. We suggest the species signal is a natural consequence of concerted gene expression evolution resulting from mutations that alter gene regulatory network interactions shared by the characters under comparison. Thus, character trees generated from transcriptomes allow us to investigate the variational independence, or individuation, of morphological characters at the level of genetic programs. © 2015 Wiley Periodicals, Inc.

CO-Releasing Molecules Have Nonheme Targets in Bacteria: Transcriptomic, Mathematical Modeling and Biochemical Analyses of CORM-3 [Ru(CO)3Cl(glycinate)] Actions on a Heme-Deficient Mutant of Escherichia coli

PubMed Central

Wilson, Jayne Louise; Wareham, Lauren K.; McLean, Samantha; Begg, Ronald; Greaves, Sarah; Mann, Brian E.; Sanguinetti, Guido

2015-01-01

Abstract Aims: Carbon monoxide-releasing molecules (CORMs) are being developed with the ultimate goal of safely utilizing the therapeutic potential of CO clinically, including applications in antimicrobial therapy. Hemes are generally considered the prime targets of CO and CORMs, so we tested this hypothesis using heme-deficient bacteria, applying cellular, transcriptomic, and biochemical tools. Results: CORM-3 [Ru(CO)3Cl(glycinate)] readily penetrated Escherichia coli hemA bacteria and was inhibitory to these and Lactococcus lactis, even though they lack all detectable hemes. Transcriptomic analyses, coupled with mathematical modeling of transcription factor activities, revealed that the response to CORM-3 in hemA bacteria is multifaceted but characterized by markedly elevated expression of iron acquisition and utilization mechanisms, global stress responses, and zinc management processes. Cell membranes are disturbed by CORM-3. Innovation: This work has demonstrated for the first time that CORM-3 (and to a lesser extent its inactivated counterpart) has multiple cellular targets other than hemes. A full understanding of the actions of CORMs is vital to understand their toxic effects. Conclusion: This work has furthered our understanding of the key targets of CORM-3 in bacteria and raises the possibility that the widely reported antimicrobial effects cannot be attributed to classical biochemical targets of CO. This is a vital step in exploiting the potential, already demonstrated, for using optimized CORMs in antimicrobial therapy. Antioxid. Redox Signal. 23, 148–162. PMID:25811604

Transcriptomic changes throughout post-hatch development in Gallus gallus pituitary

PubMed Central

Lamont, Susan J; Schmidt, Carl J

2016-01-01

The pituitary gland is a neuroendocrine organ that works closely with the hypothalamus to affect multiple processes within the body including the stress response, metabolism, growth and immune function. Relative tissue expression (rEx) is a transcriptome analysis method that compares the genes expressed in a particular tissue to the genes expressed in all other tissues with available data. Using rEx, the aim of this study was to identify genes that are uniquely or more abundantly expressed in the pituitary when compared to all other collected chicken tissues. We applied rEx to define genes enriched in the chicken pituitaries at days 21, 22 and 42 post-hatch. rEx analysis identified 25 genes shared between all time points, 295 genes shared between days 21 and 22 and 407 genes unique to day 42. The 25 genes shared by all time points are involved in morphogenesis and general nervous tissue development. The 295 shared genes between days 21 and 22 are involved in neurogenesis and nervous system development and differentiation. The 407 unique day 42 genes are involved in pituitary development, endocrine system development and other hormonally related gene ontology terms. Overall, rEx analysis indicates a focus on nervous system/tissue development at days 21 and 22. By day 42, in addition to nervous tissue development, there is expression of genes involved in the endocrine system, possibly for maturation and preparation for reproduction. This study defines the transcriptome of the chicken pituitary gland and aids in understanding the expressed genes critical to its function and maturation. PMID:27856505

A high-resolution transcriptome map of cell cycle reveals novel connections between periodic genes and cancer

PubMed Central

Dominguez, Daniel; Tsai, Yi-Hsuan; Gomez, Nicholas; Jha, Deepak Kumar; Davis, Ian; Wang, Zefeng

2016-01-01

Progression through the cell cycle is largely dependent on waves of periodic gene expression, and the regulatory networks for these transcriptome dynamics have emerged as critical points of vulnerability in various aspects of tumor biology. Through RNA-sequencing of human cells during two continuous cell cycles (>2.3 billion paired reads), we identified over 1 000 mRNAs, non-coding RNAs and pseudogenes with periodic expression. Periodic transcripts are enriched in functions related to DNA metabolism, mitosis, and DNA damage response, indicating these genes likely represent putative cell cycle regulators. Using our set of periodic genes, we developed a new approach termed “mitotic trait” that can classify primary tumors and normal tissues by their transcriptome similarity to different cell cycle stages. By analyzing >4 000 tumor samples in The Cancer Genome Atlas (TCGA) and other expression data sets, we found that mitotic trait significantly correlates with genetic alterations, tumor subtype and, notably, patient survival. We further defined a core set of 67 genes with robust periodic expression in multiple cell types. Proteins encoded by these genes function as major hubs of protein-protein interaction and are mostly required for cell cycle progression. The core genes also have unique chromatin features including increased levels of CTCF/RAD21 binding and H3K36me3. Loss of these features in uterine and kidney cancers is associated with altered expression of the core 67 genes. Our study suggests new chromatin-associated mechanisms for periodic gene regulation and offers a predictor of cancer patient outcomes. PMID:27364684

Multi-tissue transcriptomics for construction of a comprehensive gene resource for the terrestrial snail Theba pisana.

PubMed

Zhao, M; Wang, T; Adamson, K J; Storey, K B; Cummins, S F

2016-02-08

The land snail Theba pisana is native to the Mediterranean region but has become one of the most abundant invasive species worldwide. Here, we present three transcriptomes of this agriculture pest derived from three tissues: the central nervous system, hepatopancreas (digestive gland), and foot muscle. Sequencing of the three tissues produced 339,479,092 high quality reads and a global de novo assembly generated a total of 250,848 unique transcripts (unigenes). BLAST analysis mapped 52,590 unigenes to NCBI non-redundant protein databases and further functional analysis annotated 21,849 unigenes with gene ontology. We report that T. pisana transcripts have representatives in all functional classes and a comparison of differentially expressed transcripts amongst all three tissues demonstrates enormous differences in their potential metabolic activities. The genes differentially expressed include those with sequence similarity to those genes associated with multiple bacterial diseases and neurological diseases. To provide a valuable resource that will assist functional genomics study, we have implemented a user-friendly web interface, ThebaDB (http://thebadb.bioinfo-minzhao.org/). This online database allows for complex text queries, sequence searches, and data browsing by enriched functional terms and KEGG mapping.

Genome-Wide Identification and Transcriptome-Based Expression Profiling of the Sox Gene Family in the Nile Tilapia (Oreochromis niloticus)

PubMed Central

Wei, Ling; Yang, Chao; Tao, Wenjing; Wang, Deshou

2016-01-01

The Sox transcription factor family is characterized with the presence of a Sry-related high-mobility group (HMG) box and plays important roles in various biological processes in animals, including sex determination and differentiation, and the development of multiple organs. In this study, 27 Sox genes were identified in the genome of the Nile tilapia (Oreochromis niloticus), and were classified into seven groups. The members of each group of the tilapia Sox genes exhibited a relatively conserved exon-intron structure. Comparative analysis showed that the Sox gene family has undergone an expansion in tilapia and other teleost fishes following their whole genome duplication, and group K only exists in teleosts. Transcriptome-based analysis demonstrated that most of the tilapia Sox genes presented stage-specific and/or sex-dimorphic expressions during gonadal development, and six of the group B Sox genes were specifically expressed in the adult brain. Our results provide a better understanding of gene structure and spatio-temporal expression of the Sox gene family in tilapia, and will be useful for further deciphering the roles of the Sox genes during sex determination and gonadal development in teleosts. PMID:26907269

Genome-Wide Identification and Transcriptome-Based Expression Profiling of the Sox Gene Family in the Nile Tilapia (Oreochromis niloticus).

PubMed

Wei, Ling; Yang, Chao; Tao, Wenjing; Wang, Deshou

2016-02-23

The Sox transcription factor family is characterized with the presence of a Sry-related high-mobility group (HMG) box and plays important roles in various biological processes in animals, including sex determination and differentiation, and the development of multiple organs. In this study, 27 Sox genes were identified in the genome of the Nile tilapia (Oreochromis niloticus), and were classified into seven groups. The members of each group of the tilapia Sox genes exhibited a relatively conserved exon-intron structure. Comparative analysis showed that the Sox gene family has undergone an expansion in tilapia and other teleost fishes following their whole genome duplication, and group K only exists in teleosts. Transcriptome-based analysis demonstrated that most of the tilapia Sox genes presented stage-specific and/or sex-dimorphic expressions during gonadal development, and six of the group B Sox genes were specifically expressed in the adult brain. Our results provide a better understanding of gene structure and spatio-temporal expression of the Sox gene family in tilapia, and will be useful for further deciphering the roles of the Sox genes during sex determination and gonadal development in teleosts.

Applications of RNA Indexes for Precision Oncology in Breast Cancer.

PubMed

Ma, Liming; Liang, Zirui; Zhou, Hui; Qu, Lianghu

2018-05-09

Precision oncology aims to offer the most appropriate treatments to cancer patients mainly based on their individual genetic information. Genomics has provided numerous valuable data on driver mutations and risk loci; however, it remains a formidable challenge to transform these data into therapeutic agents. Transcriptomics describes the multifarious expression patterns of both mRNAs and non-coding RNAs (ncRNAs), which facilitates the deciphering of genomic codes. In this review, we take breast cancer as an example to demonstrate the applications of these rich RNA resources in precision medicine exploration. These include the use of mRNA profiles in triple-negative breast cancer (TNBC) subtyping to inform corresponding candidate targeted therapies; current advancements and achievements of high-throughput RNA interference (RNAi) screening technologies in breast cancer; and microRNAs as functional signatures for defining cell identities and regulating the biological activities of breast cancer cells. We summarize the benefits of transcriptomic analyses in breast cancer management and propose that unscrambling the core signaling networks of cancer may be an important task of multiple-omic data integration for precision oncology. Copyright © 2018 The Authors. Production and hosting by Elsevier B.V. All rights reserved.

NvERTx: a gene expression database to compare embryogenesis and regeneration in the sea anemone Nematostella vectensis.

PubMed

Warner, Jacob F; Guerlais, Vincent; Amiel, Aldine R; Johnston, Hereroa; Nedoncelle, Karine; Röttinger, Eric

2018-05-17

For over a century, researchers have been comparing embryogenesis and regeneration hoping that lessons learned from embryonic development will unlock hidden regenerative potential. This problem has historically been a difficult one to investigate because the best regenerative model systems are poor embryonic models and vice versa. Recently, however, there has been renewed interest in this question, as emerging models have allowed researchers to investigate these processes in the same organism. This interest has been further fueled by the advent of high-throughput transcriptomic analyses that provide virtual mountains of data. Here, we present N ematostella vectensis Embryogenesis and Regeneration Transcriptomics (NvERTx), a platform for comparing gene expression during embryogenesis and regeneration. NvERTx consists of close to 50 transcriptomic data sets spanning embryogenesis and regeneration in Nematostella These data were used to perform a robust de novo transcriptome assembly, with which users can search, conduct BLAST analyses, and plot the expression of multiple genes during these two developmental processes. The site is also home to the results of gene clustering analyses, to further mine the data and identify groups of co-expressed genes. The site can be accessed at http://nvertx.kahikai.org. © 2018. Published by The Company of Biologists Ltd.

Single-Cell Sequencing for Drug Discovery and Drug Development.

PubMed

Wu, Hongjin; Wang, Charles; Wu, Shixiu

2017-01-01

Next-generation sequencing (NGS), particularly single-cell sequencing, has revolutionized the scale and scope of genomic and biomedical research. Recent technological advances in NGS and singlecell studies have made the deep whole-genome (DNA-seq), whole epigenome and whole-transcriptome sequencing (RNA-seq) at single-cell level feasible. NGS at the single-cell level expands our view of genome, epigenome and transcriptome and allows the genome, epigenome and transcriptome of any organism to be explored without a priori assumptions and with unprecedented throughput. And it does so with single-nucleotide resolution. NGS is also a very powerful tool for drug discovery and drug development. In this review, we describe the current state of single-cell sequencing techniques, which can provide a new, more powerful and precise approach for analyzing effects of drugs on treated cells and tissues. Our review discusses single-cell whole genome/exome sequencing (scWGS/scWES), single-cell transcriptome sequencing (scRNA-seq), single-cell bisulfite sequencing (scBS), and multiple omics of single-cell sequencing. We also highlight the advantages and challenges of each of these approaches. Finally, we describe, elaborate and speculate the potential applications of single-cell sequencing for drug discovery and drug development. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Analysis of drought-responsive signalling network in two contrasting rice cultivars using transcriptome-based approach

PubMed Central

Borah, Pratikshya; Sharma, Eshan; Kaur, Amarjot; Chandel, Girish; Mohapatra, Trilochan; Kapoor, Sanjay; Khurana, Jitendra P.

2017-01-01

Traditional cultivars of rice in India exhibit tolerance to drought stress due to their inherent genetic variations. Here we present comparative physiological and transcriptome analyses of two contrasting cultivars, drought tolerant Dhagaddeshi (DD) and susceptible IR20. Microarray analysis revealed several differentially expressed genes (DEGs) exclusively in DD as compared to IR20 seedlings exposed to 3 h drought stress. Physiologically, DD seedlings showed higher cell membrane stability and differential ABA accumulation in response to dehydration, coupled with rapid changes in gene expression. Detailed analyses of metabolic pathways enriched in expression data suggest interplay of ABA dependent along with secondary and redox metabolic networks that activate osmotic and detoxification signalling in DD. By co-localization of DEGs with QTLs from databases or published literature for physiological traits of DD and IR20, candidate genes were identified including those underlying major QTL qDTY1.1 in DD. Further, we identified previously uncharacterized genes from both DD and IR20 under drought conditions including OsWRKY51, OsVP1 and confirmed their expression by qPCR in multiple rice cultivars. OsFBK1 was also functionally validated in susceptible PB1 rice cultivar and Arabidopsis for providing drought tolerance. Some of the DEGs mapped to the known QTLs could thus, be of potential significance for marker-assisted breeding. PMID:28181537

Multiple plant hormones and cell wall metabolism regulate apple fruit maturation patterns and texture attributes

USDA-ARS?s Scientific Manuscript database

Molecular events regulating apple fruit ripening and sensory quality are largely unknown. Such knowledge is essential for genomic-assisted apple breeding and postharvest quality management. In this study, a parallel transcriptome profile analysis, scanning electron microscopic (SEM) examination and...

Cell type transcriptome atlas for the planarian Schmidtea mediterranea.

PubMed

Fincher, Christopher T; Wurtzel, Omri; de Hoog, Thom; Kravarik, Kellie M; Reddien, Peter W

2018-05-25

The transcriptome of a cell dictates its unique cell type biology. We used single-cell RNA sequencing to determine the transcriptomes for essentially every cell type of a complete animal: the regenerative planarian Schmidtea mediterranea. Planarians contain a diverse array of cell types, possess lineage progenitors for differentiated cells (including pluripotent stem cells), and constitutively express positional information, making them ideal for this undertaking. We generated data for 66,783 cells, defining transcriptomes for known and many previously unknown planarian cell types and for putative transition states between stem and differentiated cells. We also uncovered regionally expressed genes in muscle, which harbors positional information. Identifying the transcriptomes for potentially all cell types for many organisms should be readily attainable and represents a powerful approach to metazoan biology. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

High-confidence coding and noncoding transcriptome maps

PubMed Central

2017-01-01

The advent of high-throughput RNA sequencing (RNA-seq) has led to the discovery of unprecedentedly immense transcriptomes encoded by eukaryotic genomes. However, the transcriptome maps are still incomplete partly because they were mostly reconstructed based on RNA-seq reads that lack their orientations (known as unstranded reads) and certain boundary information. Methods to expand the usability of unstranded RNA-seq data by predetermining the orientation of the reads and precisely determining the boundaries of assembled transcripts could significantly benefit the quality of the resulting transcriptome maps. Here, we present a high-performing transcriptome assembly pipeline, called CAFE, that significantly improves the original assemblies, respectively assembled with stranded and/or unstranded RNA-seq data, by orienting unstranded reads using the maximum likelihood estimation and by integrating information about transcription start sites and cleavage and polyadenylation sites. Applying large-scale transcriptomic data comprising 230 billion RNA-seq reads from the ENCODE, Human BodyMap 2.0, The Cancer Genome Atlas, and GTEx projects, CAFE enabled us to predict the directions of about 220 billion unstranded reads, which led to the construction of more accurate transcriptome maps, comparable to the manually curated map, and a comprehensive lncRNA catalog that includes thousands of novel lncRNAs. Our pipeline should not only help to build comprehensive, precise transcriptome maps from complex genomes but also to expand the universe of noncoding genomes. PMID:28396519

Transcriptome profiling reveals regulatory mechanisms underlying Corolla Senescence in Petunia

USDA-ARS?s Scientific Manuscript database

Genetic regulatory mechanisms that govern petal natural senescence in petunia is complicated and unclear. To identify key genes and pathways that regulate the process, we initiated a transcriptome analysis in petunia petals at four developmental time points, including petal opening without anthesis ...

Ab initio reconstruction of transcriptomes of pluripotent and lineage committed cells reveals gene structures of thousands of lincRNAs

PubMed Central

Guttman, Mitchell; Garber, Manuel; Levin, Joshua Z.; Donaghey, Julie; Robinson, James; Adiconis, Xian; Fan, Lin; Koziol, Magdalena J.; Gnirke, Andreas; Nusbaum, Chad; Rinn, John L.; Lander, Eric S.; Regev, Aviv

2010-01-01

RNA-Seq provides an unbiased way to study a transcriptome, including both coding and non-coding genes. To date, most RNA-Seq studies have critically depended on existing annotations, and thus focused on expression levels and variation in known transcripts. Here, we present Scripture, a method to reconstruct the transcriptome of a mammalian cell using only RNA-Seq reads and the genome sequence. We apply it to mouse embryonic stem cells, neuronal precursor cells, and lung fibroblasts to accurately reconstruct the full-length gene structures for the vast majority of known expressed genes. We identify substantial variation in protein-coding genes, including thousands of novel 5′-start sites, 3′-ends, and internal coding exons. We then determine the gene structures of over a thousand lincRNA and antisense loci. Our results open the way to direct experimental manipulation of thousands of non-coding RNAs, and demonstrate the power of ab initio reconstruction to render a comprehensive picture of mammalian transcriptomes. PMID:20436462

Systems-wide RNAi analysis of CASP8AP2/FLASH shows transcriptional deregulation of the replication-dependent histone genes and extensive effects on the transcriptome of colorectal cancer cells

PubMed Central

2012-01-01

Background Colorectal carcinomas (CRC) carry massive genetic and transcriptional alterations that influence multiple cellular pathways. The study of proteins whose loss-of-function (LOF) alters the growth of CRC cells can be used to further understand the cellular processes cancer cells depend upon for survival. Results A small-scale RNAi screen of ~400 genes conducted in SW480 CRC cells identified several candidate genes as required for the viability of CRC cells, most prominently CASP8AP2/FLASH. To understand the function of this gene in maintaining the viability of CRC cells in an unbiased manner, we generated gene specific expression profiles following RNAi. Silencing of CASP8AP2/FLASH resulted in altered expression of over 2500 genes enriched for genes associated with cellular growth and proliferation. Loss of CASP8AP2/FLASH function was significantly associated with altered transcription of the genes encoding the replication-dependent histone proteins as a result of the expression of the non-canonical polyA variants of these transcripts. Silencing of CASP8AP2/FLASH also mediated enrichment of changes in the expression of targets of the NFκB and MYC transcription factors. These findings were confirmed by whole transcriptome analysis of CASP8AP2/FLASH silenced cells at multiple time points. Finally, we identified and validated that CASP8AP2/FLASH LOF increases the expression of neurofilament heavy polypeptide (NEFH), a protein recently linked to regulation of the AKT1/ß-catenin pathway. Conclusions We have used unbiased RNAi based approaches to identify and characterize the function of CASP8AP2/FLASH, a protein not previously reported as required for cell survival. This study further defines the role CASP8AP2/FLASH plays in the regulating expression of the replication-dependent histones and shows that its LOF results in broad and reproducible effects on the transcriptome of colorectal cancer cells including the induction of expression of the recently described tumor suppressor gene NEFH. PMID:22216762

Systems-wide RNAi analysis of CASP8AP2/FLASH shows transcriptional deregulation of the replication-dependent histone genes and extensive effects on the transcriptome of colorectal cancer cells.

PubMed

Hummon, Amanda B; Pitt, Jason J; Camps, Jordi; Emons, Georg; Skube, Susan B; Huppi, Konrad; Jones, Tamara L; Beissbarth, Tim; Kramer, Frank; Grade, Marian; Difilippantonio, Michael J; Ried, Thomas; Caplen, Natasha J

2012-01-04

Colorectal carcinomas (CRC) carry massive genetic and transcriptional alterations that influence multiple cellular pathways. The study of proteins whose loss-of-function (LOF) alters the growth of CRC cells can be used to further understand the cellular processes cancer cells depend upon for survival. A small-scale RNAi screen of ~400 genes conducted in SW480 CRC cells identified several candidate genes as required for the viability of CRC cells, most prominently CASP8AP2/FLASH. To understand the function of this gene in maintaining the viability of CRC cells in an unbiased manner, we generated gene specific expression profiles following RNAi. Silencing of CASP8AP2/FLASH resulted in altered expression of over 2500 genes enriched for genes associated with cellular growth and proliferation. Loss of CASP8AP2/FLASH function was significantly associated with altered transcription of the genes encoding the replication-dependent histone proteins as a result of the expression of the non-canonical polyA variants of these transcripts. Silencing of CASP8AP2/FLASH also mediated enrichment of changes in the expression of targets of the NFκB and MYC transcription factors. These findings were confirmed by whole transcriptome analysis of CASP8AP2/FLASH silenced cells at multiple time points. Finally, we identified and validated that CASP8AP2/FLASH LOF increases the expression of neurofilament heavy polypeptide (NEFH), a protein recently linked to regulation of the AKT1/ß-catenin pathway. We have used unbiased RNAi based approaches to identify and characterize the function of CASP8AP2/FLASH, a protein not previously reported as required for cell survival. This study further defines the role CASP8AP2/FLASH plays in the regulating expression of the replication-dependent histones and shows that its LOF results in broad and reproducible effects on the transcriptome of colorectal cancer cells including the induction of expression of the recently described tumor suppressor gene NEFH.

Molecular characteristics of the KCNJ5 mutated aldosterone-producing adenomas.

PubMed

Murakami, Masanori; Yoshimoto, Takanobu; Nakabayashi, Kazuhiko; Nakano, Yujiro; Fukaishi, Takahiro; Tsuchiya, Kyoichiro; Minami, Isao; Bouchi, Ryotaro; Okamura, Kohji; Fujii, Yasuhisa; Hashimoto, Koshi; Hata, Ken-Ichiro; Kihara, Kazunori; Ogawa, Yoshihiro

2017-10-01

The pathophysiology of aldosterone-producing adenomas (APAs) has been investigated via genetic approaches and the pathogenic significance of a series of somatic mutations, including KCNJ5 , has been uncovered. However, how the mutational status of an APA is associated with its molecular characteristics, including its transcriptome and methylome, has not been fully understood. This study was undertaken to explore the molecular characteristics of APAs, specifically focusing on APAs with KCNJ5 mutations as opposed to those without KCNJ5 mutations, by comparing their transcriptome and methylome status. Cortisol-producing adenomas (CPAs) were used as reference. We conducted transcriptome and methylome analyses of 29 APAs with KCNJ5 mutations, 8 APAs without KCNJ5 mutations and 5 CPAs. Genome-wide gene expression and CpG methylation profiles were obtained from RNA and DNA samples extracted from these 42 adrenal tumors. Cluster analysis of the transcriptome and methylome revealed molecular heterogeneity in APAs depending on their mutational status. DNA hypomethylation and gene expression changes in Wnt signaling and inflammatory response pathways were characteristic of APAs with KCNJ5 mutations. Comparisons between transcriptome data from our APAs and that from normal adrenal cortex obtained from the Gene Expression Omnibus suggested similarities between APAs with KCNJ5 mutations and zona glomerulosa. The present study, which is based on transcriptome and methylome analyses, indicates the molecular heterogeneity of APAs depends on their mutational status. Here, we report the unique characteristics of APAs with KCNJ5 mutations. © 2017 Society for Endocrinology.

A genome-wide transcriptome map of pistachio (Pistacia vera L.) provides novel insights into salinity-related genes and marker discovery.

PubMed

Moazzzam Jazi, Maryam; Seyedi, Seyed Mahdi; Ebrahimie, Esmaeil; Ebrahimi, Mansour; De Moro, Gianluca; Botanga, Christopher

2017-08-17

Pistachio (Pistacia vera L.) is one of the most important commercial nut crops worldwide. It is a salt-tolerant and long-lived tree, with the largest cultivation area in Iran. Climate change and subsequent increased soil salt content have adversely affected the pistachio yield in recent years. However, the lack of genomic/global transcriptomic sequences on P. vera impedes comprehensive researches at the molecular level. Hence, whole transcriptome sequencing is required to gain insight into functional genes and pathways in response to salt stress. RNA sequencing of a pooled sample representing 24 different tissues of two pistachio cultivars with contrasting salinity tolerance under control and salt treatment by Illumina Hiseq 2000 platform resulted in 368,953,262 clean 100 bp paired-ends reads (90 Gb). Following creating several assemblies and assessing their quality from multiple perspectives, we found that using the annotation-based metrics together with the length-based parameters allows an improved assessment of the transcriptome assembly quality, compared to the solely use of the length-based parameters. The generated assembly by Trinity was adopted for functional annotation and subsequent analyses. In total, 29,119 contigs annotated against all of five public databases, including NR, UniProt, TAIR10, KOG and InterProScan. Among 279 KEGG pathways supported by our assembly, we further examined the pathways involved in the plant hormone biosynthesis and signaling as well as those to be contributed to secondary metabolite biosynthesis due to their importance under salinity stress. In total, 11,337 SSRs were also identified, which the most abundant being dinucleotide repeats. Besides, 13,097 transcripts as candidate stress-responsive genes were identified. Expression of some of these genes experimentally validated through quantitative real-time PCR (qRT-PCR) that further confirmed the accuracy of the assembly. From this analysis, the contrasting expression pattern of NCED3 and SOS1 genes were observed between salt-sensitive and salt-tolerant cultivars. This study, as the first report on the whole transcriptome survey of P. vera, provides important resources and paves the way for functional and comparative genomic studies on this major tree to discover the salinity tolerance-related markers and stress response mechanisms for breeding of new pistachio cultivars with more salinity tolerance.

The genomic landscape of rapid, repeated evolutionary rescue from toxic pollution in wild fish

USDA-ARS?s Scientific Manuscript database

Here we describe evolutionary rescue from intense pollution via multiple modes of selection in killifish populations from 4 urban estuaries of the US eastern seaboard. Comparative transcriptomics and analysis of 384 whole genome sequences show that the functioning of a receptor-based signaling pathw...

Large-scale atlas of microarray data reveals biological landscape of gene expression in Arabidopsis

USDA-ARS?s Scientific Manuscript database

Transcriptome datasets from thousands of samples of the model plant Arabidopsis thaliana have been collectively generated by multiple individual labs. Although integration and meta-analysis of these samples has become routine in the plant research community, it is often hampered by the lack of metad...

Transcriptome analysis reveals the same 17 S-locus F-box genes in two haplotypes of the self-incompatibility locus of Petunia inflata.

PubMed

Williams, Justin S; Der, Joshua P; dePamphilis, Claude W; Kao, Teh-Hui

2014-07-01

Petunia possesses self-incompatibility, by which pistils reject self-pollen but accept non-self-pollen for fertilization. Self-/non-self-recognition between pollen and pistil is regulated by the pistil-specific S-RNase gene and by multiple pollen-specific S-locus F-box (SLF) genes. To date, 10 SLF genes have been identified by various methods, and seven have been shown to be involved in pollen specificity. For a given S-haplotype, each SLF interacts with a subset of its non-self S-RNases, and an as yet unknown number of SLFs are thought to collectively mediate ubiquitination and degradation of all non-self S-RNases to allow cross-compatible pollination. To identify a complete suite of SLF genes of P. inflata, we used a de novo RNA-seq approach to analyze the pollen transcriptomes of S2-haplotype and S3-haplotype, as well as the leaf transcriptome of the S3S3 genotype. We searched for genes that fit several criteria established from the properties of the known SLF genes and identified the same seven new SLF genes in S2-haplotype and S3-haplotype, suggesting that a total of 17 SLF genes constitute pollen specificity in each S-haplotype. This finding lays the foundation for understanding how multiple SLF genes evolved and the biochemical basis for differential interactions between SLF proteins and S-RNases. © 2014 American Society of Plant Biologists. All rights reserved.

Extreme diversity of scorpion venom peptides and proteins revealed by transcriptomic analysis: implication for proteome evolution of scorpion venom arsenal.

PubMed

Ma, Yibao; He, Yawen; Zhao, Ruiming; Wu, Yingliang; Li, Wenxin; Cao, Zhijian

2012-02-16

Venom is an important genetic development crucial to the survival of scorpions for over 400 million years. We studied the evolution of the scorpion venom arsenal by means of comparative transcriptome analysis of venom glands and phylogenetic analysis of shared types of venom peptides and proteins between buthids and euscorpiids. Fifteen types of venom peptides and proteins were sequenced during the venom gland transcriptome analyses of two Buthidae species (Lychas mucronatus and Isometrus maculatus) and one Euscorpiidae species (Scorpiops margerisonae). Great diversity has been observed in translated amino acid sequences of these transcripts for venom peptides and proteins. Seven types of venom peptides and proteins were shared between buthids and euscorpiids. Molecular phylogenetic analysis revealed that at least five of the seven common types of venom peptides and proteins were likely recruited into the scorpion venom proteome before the lineage split between Buthidae and Euscorpiidae with their corresponding genes undergoing individual or multiple gene duplication events. These are α-KTxs, βKSPNs (β-KTxs and scorpines), anionic peptides, La1-like peptides, and SPSVs (serine proteases from scorpion venom). Multiple types of venom peptides and proteins were demonstrated to be continuously recruited into the venom proteome during the evolution process of individual scorpion lineages. Our results provide an insight into the recruitment pattern of the scorpion venom arsenal for the first time. Copyright © 2011 Elsevier B.V. All rights reserved.

Insights from the pollination drop proteome and the ovule transcriptome of Cephalotaxus at the time of pollination drop production.

PubMed

Pirone-Davies, Cary; Prior, Natalie; von Aderkas, Patrick; Smith, Derek; Hardie, Darryl; Friedman, William E; Mathews, Sarah

2016-05-01

Many gymnosperms produce an ovular secretion, the pollination drop, during reproduction. The drops serve as a landing site for pollen, but also contain a suite of ions and organic compounds, including proteins, that suggests diverse roles for the drop during pollination. Proteins in the drops of species of Chamaecyparis, Juniperus, Taxus, Pseudotsuga, Ephedra and Welwitschia are thought to function in the conversion of sugars, defence against pathogens, and pollen growth and development. To better understand gymnosperm pollination biology, the pollination drop proteomes of pollination drops from two species of Cephalotaxus have been characterized and an ovular transcriptome for C. sinensis has been assembled. Mass spectrometry was used to identify proteins in the pollination drops of Cephalotaxus sinensis and C. koreana RNA-sequencing (RNA-Seq) was employed to assemble a transcriptome and identify transcripts present in the ovules of C. sinensis at the time of pollination drop production. About 30 proteins were detected in the pollination drops of both species. Many of these have been detected in the drops of other gymnosperms and probably function in defence, polysaccharide metabolism and pollen tube growth. Other proteins appear to be unique to Cephalotaxus, and their putative functions include starch and callose degradation, among others. Together, the proteins appear either to have been secreted into the drop or to occur there due to breakdown of ovular cells during drop production. Ovular transcripts represent a wide range of gene ontology categories, and some may be involved in drop formation, ovule development and pollen-ovule interactions. The proteome of Cephalotaxus pollination drops shares a number of components with those of other conifers and gnetophytes, including proteins for defence such as chitinases and for carbohydrate modification such as β-galactosidase. Proteins likely to be of intracellular origin, however, form a larger component of drops from Cephalotaxus than expected from studies of other conifers. This is consistent with the observation of nucellar breakdown during drop formation in Cephalotaxus The transcriptome data provide a framework for understanding multiple metabolic processes that occur within the ovule and the pollination drop just before fertilization. They reveal the deep conservation of WUSCHEL expression in ovules and raise questions about whether any of the S-locus transcripts in Cephalotaxus ovules might be involved in pollen-ovule recognition. © The Author 2016. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Current knowledge of detoxification mechanisms of xenobiotic in honey bees.

PubMed

Gong, Youhui; Diao, Qingyun

2017-01-01

The western honey bee Apis mellifera is the most important managed pollinator species in the world. Multiple factors have been implicated as potential causes or factors contributing to colony collapse disorder, including honey bee pathogens and nutritional deficiencies as well as exposure to pesticides. Honey bees' genome is characterized by a paucity of genes associated with detoxification, which makes them vulnerable to specific pesticides, especially to combinations of pesticides in real field environments. Many studies have investigated the mechanisms involved in detoxification of xenobiotics/pesticides in honey bees, from primal enzyme assays or toxicity bioassays to characterization of transcript gene expression and protein expression in response to xenobiotics/insecticides by using a global transcriptomic or proteomic approach, and even to functional characterizations. The global transcriptomic and proteomic approach allowed us to learn that detoxification mechanisms in honey bees involve multiple genes and pathways along with changes in energy metabolism and cellular stress response. P450 genes, is highly implicated in the direct detoxification of xenobiotics/insecticides in honey bees and their expression can be regulated by honey/pollen constitutes, resulting in the tolerance of honey bees to other xenobiotics or insecticides. P450s is also a key detoxification enzyme that mediate synergism interaction between acaricides/insecticides and fungicides through inhibition P450 activity by fungicides or competition for detoxification enzymes between acaricides. With the wide use of insecticides in agriculture, understanding the detoxification mechanism of insecticides in honey bees and how honeybees fight with the xenobiotis or insecticides to survive in the changing environment will finally benefit honeybees' management.

A high resolution atlas of gene expression in the domestic sheep (Ovis aries)

PubMed Central

Farquhar, Iseabail L.; Young, Rachel; Lefevre, Lucas; Pridans, Clare; Tsang, Hiu G.; Afrasiabi, Cyrus; Watson, Mick; Whitelaw, C. Bruce; Freeman, Tom C.; Archibald, Alan L.; Hume, David A.

2017-01-01

Sheep are a key source of meat, milk and fibre for the global livestock sector, and an important biomedical model. Global analysis of gene expression across multiple tissues has aided genome annotation and supported functional annotation of mammalian genes. We present a large-scale RNA-Seq dataset representing all the major organ systems from adult sheep and from several juvenile, neonatal and prenatal developmental time points. The Ovis aries reference genome (Oar v3.1) includes 27,504 genes (20,921 protein coding), of which 25,350 (19,921 protein coding) had detectable expression in at least one tissue in the sheep gene expression atlas dataset. Network-based cluster analysis of this dataset grouped genes according to their expression pattern. The principle of ‘guilt by association’ was used to infer the function of uncharacterised genes from their co-expression with genes of known function. We describe the overall transcriptional signatures present in the sheep gene expression atlas and assign those signatures, where possible, to specific cell populations or pathways. The findings are related to innate immunity by focusing on clusters with an immune signature, and to the advantages of cross-breeding by examining the patterns of genes exhibiting the greatest expression differences between purebred and crossbred animals. This high-resolution gene expression atlas for sheep is, to our knowledge, the largest transcriptomic dataset from any livestock species to date. It provides a resource to improve the annotation of the current reference genome for sheep, presenting a model transcriptome for ruminants and insight into gene, cell and tissue function at multiple developmental stages. PMID:28915238

A high resolution atlas of gene expression in the domestic sheep (Ovis aries).

PubMed

Clark, Emily L; Bush, Stephen J; McCulloch, Mary E B; Farquhar, Iseabail L; Young, Rachel; Lefevre, Lucas; Pridans, Clare; Tsang, Hiu G; Wu, Chunlei; Afrasiabi, Cyrus; Watson, Mick; Whitelaw, C Bruce; Freeman, Tom C; Summers, Kim M; Archibald, Alan L; Hume, David A

2017-09-01

Sheep are a key source of meat, milk and fibre for the global livestock sector, and an important biomedical model. Global analysis of gene expression across multiple tissues has aided genome annotation and supported functional annotation of mammalian genes. We present a large-scale RNA-Seq dataset representing all the major organ systems from adult sheep and from several juvenile, neonatal and prenatal developmental time points. The Ovis aries reference genome (Oar v3.1) includes 27,504 genes (20,921 protein coding), of which 25,350 (19,921 protein coding) had detectable expression in at least one tissue in the sheep gene expression atlas dataset. Network-based cluster analysis of this dataset grouped genes according to their expression pattern. The principle of 'guilt by association' was used to infer the function of uncharacterised genes from their co-expression with genes of known function. We describe the overall transcriptional signatures present in the sheep gene expression atlas and assign those signatures, where possible, to specific cell populations or pathways. The findings are related to innate immunity by focusing on clusters with an immune signature, and to the advantages of cross-breeding by examining the patterns of genes exhibiting the greatest expression differences between purebred and crossbred animals. This high-resolution gene expression atlas for sheep is, to our knowledge, the largest transcriptomic dataset from any livestock species to date. It provides a resource to improve the annotation of the current reference genome for sheep, presenting a model transcriptome for ruminants and insight into gene, cell and tissue function at multiple developmental stages.

DNA microarray‐based analysis of voluntary resistance wheel running reveals novel transcriptome leading robust hippocampal plasticity

PubMed Central

Lee, Min Chul; Rakwal, Randeep; Shibato, Junko; Inoue, Koshiro; Chang, Hyukki; Soya, Hideaki

2014-01-01

Abstract In two separate experiments, voluntary resistance wheel running with 30% of body weight (RWR), rather than wheel running (WR), led to greater enhancements, including adult hippocampal neurogenesis and cognitive functions, in conjunction with hippocampal brain‐derived neurotrophic factor (BDNF) signaling (Lee et al., J Appl Physiol, 2012; Neurosci Lett., 2013). Here we aimed to unravel novel molecular factors and gain insight into underlying molecular mechanisms for RWR‐enhanced hippocampal functions; a high‐throughput whole‐genome DNA microarray approach was applied to rats performing voluntary running for 4 weeks. RWR rats showed a significant decrease in average running distances although average work levels increased immensely, by about 11‐fold compared to WR, resulting in muscular adaptation for the fast‐twitch plantaris muscle. Global transcriptome profiling analysis identified 128 (sedentary × WR) and 169 (sedentary × RWR) up‐regulated (>1.5‐fold change), and 97 (sedentary × WR) and 468 (sedentary × RWR) down‐regulated (<0.75‐fold change) genes. Functional categorization using both pathway‐ or specific‐disease‐state‐focused gene classifications and Ingenuity Pathway Analysis (IPA) revealed expression pattern changes in the major categories of disease and disorders, molecular functions, and physiological system development and function. Genes specifically regulated with RWR include the newly identified factors of NFATc1, AVPR1A, and FGFR4, as well as previously known factors, BDNF and CREB mRNA. Interestingly, RWR down‐regulated multiple inflammatory cytokines (IL1B, IL2RA, and TNF) and chemokines (CXCL1, CXCL10, CCL2, and CCR4) with the SYCP3, PRL genes, which are potentially involved in regulating hippocampal neuroplastic changes. These results provide understanding of the voluntary‐RWR‐related hippocampal transcriptome, which will open a window to the underlying mechanisms of the positive effects of exercise, with therapeutic value for enhancing hippocampal functions. PMID:25413326

Transcriptome characterisation of Pinus tabuliformis and evolution of genes in the Pinus phylogeny

PubMed Central

2013-01-01

Background The Chinese pine (Pinus tabuliformis) is an indigenous conifer species in northern China but is relatively underdeveloped as a genomic resource; thus, limiting gene discovery and breeding. Large-scale transcriptome data were obtained using a next-generation sequencing platform to compensate for the lack of P. tabuliformis genomic information. Results The increasing amount of transcriptome data on Pinus provides an excellent resource for multi-gene phylogenetic analysis and studies on how conserved genes and functions are maintained in the face of species divergence. The first P. tabuliformis transcriptome from a normalised cDNA library of multiple tissues and individuals was sequenced in a full 454 GS-FLX run, producing 911,302 sequencing reads. The high quality overlapping expressed sequence tags (ESTs) were assembled into 46,584 putative transcripts, and more than 700 SSRs and 92,000 SNPs/InDels were characterised. Comparative analysis of the transcriptome of six conifer species yielded 191 orthologues, from which we inferred a phylogenetic tree, evolutionary patterns and calculated rates of gene diversion. We also identified 938 fast evolving sequences that may be useful for identifying genes that perhaps evolved in response to positive selection and might be responsible for speciation in the Pinus lineage. Conclusions A large collection of high-quality ESTs was obtained, de novo assembled and characterised, which represents a dramatic expansion of the current transcript catalogues of P. tabuliformis and which will gradually be applied in breeding programs of P. tabuliformis. Furthermore, these data will facilitate future studies of the comparative genomics of P. tabuliformis and other related species. PMID:23597112

CONVERGENT TRANSCRIPTOMICS AND PROTEOMICS OF ENVIRONMENTAL ENRICHMENT AND COCAINE IDENTIFIES NOVEL THERAPEUTIC STRATEGIES FOR ADDICTION

PubMed Central

ZHANG, YAFANG; CROFTON, ELIZABETH J.; FAN, XIUZHEN; LI, DINGGE; KONG, FANPING; SINHA, MALA; LUXON, BRUCE A.; SPRATT, HEIDI M.; LICHTI, CHERYL F.; GREEN, THOMAS A.

2016-01-01

Transcriptomic and proteomic approaches have separately proven effective at identifying novel mechanisms affecting addiction-related behavior; however, it is difficult to prioritize the many promising leads from each approach. A convergent secondary analysis of proteomic and transcriptomic results can glean additional information to help prioritize promising leads. The current study is a secondary analysis of the convergence of recently published separate transcriptomic and proteomic analyses of nucleus accumbens (NAc) tissue from rats subjected to environmental enrichment vs. isolation and cocaine self-administration vs. saline. Multiple bioinformatics approaches (e.g. Gene Ontology (GO) analysis, Ingenuity Pathway Analysis (IPA), and Gene Set Enrichment Analysis (GSEA)) were used to interrogate these rich data sets. Although there was little correspondence between mRNA vs. protein at the individual target level, good correspondence was found at the level of gene/protein sets, particularly for the environmental enrichment manipulation. These data identify gene sets where there is a positive relationship between changes in mRNA and protein (e.g. glycolysis, ATP synthesis, translation elongation factor activity, etc.) and gene sets where there is an inverse relationship (e.g. ribosomes, Rho GTPase signaling, protein ubiquitination, etc.). Overall environmental enrichment produced better correspondence than cocaine self-administration. The individual targets contributing to mRNA and protein effects were largely not overlapping. As a whole, these results confirm that robust transcriptomic and proteomic data sets can provide similar results at the gene/protein set level even when there is little correspondence at the individual target level and little overlap in the targets contributing to the effects. PMID:27717806

Deciphering the Developmental Dynamics of the Mouse Liver Transcriptome

PubMed Central

Gunewardena, Sumedha S.; Yoo, Byunggil; Peng, Lai; Lu, Hong; Zhong, Xiaobo; Klaassen, Curtis D.; Cui, Julia Yue

2015-01-01

During development, liver undergoes a rapid transition from a hematopoietic organ to a major organ for drug metabolism and nutrient homeostasis. However, little is known on a transcriptome level of the genes and RNA-splicing variants that are differentially regulated with age, and which up-stream regulators orchestrate age-specific biological functions in liver. We used RNA-Seq to interrogate the developmental dynamics of the liver transcriptome in mice at 12 ages from late embryonic stage (2-days before birth) to maturity (60-days after birth). Among 21,889 unique NCBI RefSeq-annotated genes, 9,641 were significantly expressed in at least one age, 7,289 were differently regulated with age, and 859 had multiple (> = 2) RNA splicing-variants. Factor analysis showed that the dynamics of hepatic genes fall into six distinct groups based on their temporal expression. The average expression of cytokines, ion channels, kinases, phosphatases, transcription regulators and translation regulators decreased with age, whereas the average expression of peptidases, enzymes and transmembrane receptors increased with age. The average expression of growth factors peak between Day-3 and Day-10, and decrease thereafter. We identified critical biological functions, upstream regulators, and putative transcription modules that seem to govern age-specific gene expression. We also observed differential ontogenic expression of known splicing variants of certain genes, and 1,455 novel splicing isoform candidates. In conclusion, the hepatic ontogeny of the transcriptome ontogeny has unveiled critical networks and up-stream regulators that orchestrate age-specific biological functions in liver, and suggest that age contributes to the complexity of the alternative splicing landscape of the hepatic transcriptome. PMID:26496202

Deciphering the Developmental Dynamics of the Mouse Liver Transcriptome.

PubMed

Gunewardena, Sumedha S; Yoo, Byunggil; Peng, Lai; Lu, Hong; Zhong, Xiaobo; Klaassen, Curtis D; Cui, Julia Yue

2015-01-01

During development, liver undergoes a rapid transition from a hematopoietic organ to a major organ for drug metabolism and nutrient homeostasis. However, little is known on a transcriptome level of the genes and RNA-splicing variants that are differentially regulated with age, and which up-stream regulators orchestrate age-specific biological functions in liver. We used RNA-Seq to interrogate the developmental dynamics of the liver transcriptome in mice at 12 ages from late embryonic stage (2-days before birth) to maturity (60-days after birth). Among 21,889 unique NCBI RefSeq-annotated genes, 9,641 were significantly expressed in at least one age, 7,289 were differently regulated with age, and 859 had multiple (> = 2) RNA splicing-variants. Factor analysis showed that the dynamics of hepatic genes fall into six distinct groups based on their temporal expression. The average expression of cytokines, ion channels, kinases, phosphatases, transcription regulators and translation regulators decreased with age, whereas the average expression of peptidases, enzymes and transmembrane receptors increased with age. The average expression of growth factors peak between Day-3 and Day-10, and decrease thereafter. We identified critical biological functions, upstream regulators, and putative transcription modules that seem to govern age-specific gene expression. We also observed differential ontogenic expression of known splicing variants of certain genes, and 1,455 novel splicing isoform candidates. In conclusion, the hepatic ontogeny of the transcriptome ontogeny has unveiled critical networks and up-stream regulators that orchestrate age-specific biological functions in liver, and suggest that age contributes to the complexity of the alternative splicing landscape of the hepatic transcriptome.

Novel biomarkers for cardiovascular risk assessment: current status and future directions.

PubMed

MacNamara, James; Eapen, Danny J; Quyyumi, Arshed; Sperling, Laurence

2015-09-01

Cardiovascular disease (CVD) is the leading cause of mortality in the modern world. Traditional risk algorithms may miss up to 20% of CVD events. Therefore, there is a need for new cardiac biomarkers. Many fields of research are dedicated to improving cardiac risk prediction, including genomics, transcriptomics and proteomics. To date, even the most promising biomarkers have only demonstrated modest associations and predictive ability. Few have undergone randomized control trials. A number of biomarkers are targets to new therapies aimed to reduce cardiovascular risk. Currently, some of the most promising risk prediction has been demonstrated with panels of multiple biomarkers. This article reviews the current state and future of proteomic biomarkers and aggregate biomarker panels.

Cell type-specific responses to salinity - the epidermal bladder cell transcriptome of Mesembryanthemum crystallinum.

PubMed

Oh, Dong-Ha; Barkla, Bronwyn J; Vera-Estrella, Rosario; Pantoja, Omar; Lee, Sang-Yeol; Bohnert, Hans J; Dassanayake, Maheshi

2015-08-01

Mesembryanthemum crystallinum (ice plant) exhibits extreme tolerance to salt. Epidermal bladder cells (EBCs), developing on the surface of aerial tissues and specialized in sodium sequestration and other protective functions, are critical for the plant's stress adaptation. We present the first transcriptome analysis of EBCs isolated from intact plants, to investigate cell type-specific responses during plant salt adaptation. We developed a de novo assembled, nonredundant EBC reference transcriptome. Using RNAseq, we compared the expression patterns of the EBC-specific transcriptome between control and salt-treated plants. The EBC reference transcriptome consists of 37 341 transcript-contigs, of which 7% showed significantly different expression between salt-treated and control samples. We identified significant changes in ion transport, metabolism related to energy generation and osmolyte accumulation, stress signalling, and organelle functions, as well as a number of lineage-specific genes of unknown function, in response to salt treatment. The salinity-induced EBC transcriptome includes active transcript clusters, refuting the view of EBCs as passive storage compartments in the whole-plant stress response. EBC transcriptomes, differing from those of whole plants or leaf tissue, exemplify the importance of cell type-specific resolution in understanding stress adaptive mechanisms. No claim to original US government works. New Phytologist © 2015 New Phytologist Trust.

A retrospective likelihood approach for efficient integration of multiple omics factors in case-control association studies.

PubMed

Balliu, Brunilda; Tsonaka, Roula; Boehringer, Stefan; Houwing-Duistermaat, Jeanine

2015-03-01

Integrative omics, the joint analysis of outcome and multiple types of omics data, such as genomics, epigenomics, and transcriptomics data, constitute a promising approach for powerful and biologically relevant association studies. These studies often employ a case-control design, and often include nonomics covariates, such as age and gender, that may modify the underlying omics risk factors. An open question is how to best integrate multiple omics and nonomics information to maximize statistical power in case-control studies that ascertain individuals based on the phenotype. Recent work on integrative omics have used prospective approaches, modeling case-control status conditional on omics, and nonomics risk factors. Compared to univariate approaches, jointly analyzing multiple risk factors with a prospective approach increases power in nonascertained cohorts. However, these prospective approaches often lose power in case-control studies. In this article, we propose a novel statistical method for integrating multiple omics and nonomics factors in case-control association studies. Our method is based on a retrospective likelihood function that models the joint distribution of omics and nonomics factors conditional on case-control status. The new method provides accurate control of Type I error rate and has increased efficiency over prospective approaches in both simulated and real data. © 2015 Wiley Periodicals, Inc.

Human-specific features of spatial gene expression and regulation in eight brain regions.

PubMed

Xu, Chuan; Li, Qian; Efimova, Olga; He, Liu; Tatsumoto, Shoji; Stepanova, Vita; Oishi, Takao; Udono, Toshifumi; Yamaguchi, Katsushi; Shigenobu, Shuji; Kakita, Akiyoshi; Nawa, Hiroyuki; Khaitovich, Philipp; Go, Yasuhiro

2018-06-13

Molecular maps of the human brain alone do not inform us of the features unique to humans. Yet, the identification of these features is important for understanding both the evolution and nature of human cognition. Here, we approached this question by analyzing gene expression and H3K27ac chromatin modification data collected in eight brain regions of humans, chimpanzees, gorillas, a gibbon and macaques. An analysis of spatial transcriptome trajectories across eight brain regions in four primate species revealed 1,851 genes showing human-specific transcriptome differences in one or multiple brain regions, in contrast to 240 chimpanzee-specific ones. More than half of these human-specific differences represented elevated expression of genes enriched in neuronal and astrocytic markers in the human hippocampus, while the rest were enriched in microglial markers and displayed human-specific expression in several frontal cortical regions and the cerebellum. An analysis of the predicted regulatory interactions driving these differences revealed the role of transcription factors in species-specific transcriptome changes, while epigenetic modifications were linked to spatial expression differences conserved across species. Published by Cold Spring Harbor Laboratory Press.

Digital RNA sequencing minimizes sequence-dependent bias and amplification noise with optimized single-molecule barcodes

PubMed Central

Shiroguchi, Katsuyuki; Jia, Tony Z.; Sims, Peter A.; Xie, X. Sunney

2012-01-01

RNA sequencing (RNA-Seq) is a powerful tool for transcriptome profiling, but is hampered by sequence-dependent bias and inaccuracy at low copy numbers intrinsic to exponential PCR amplification. We developed a simple strategy for mitigating these complications, allowing truly digital RNA-Seq. Following reverse transcription, a large set of barcode sequences is added in excess, and nearly every cDNA molecule is uniquely labeled by random attachment of barcode sequences to both ends. After PCR, we applied paired-end deep sequencing to read the two barcodes and cDNA sequences. Rather than counting the number of reads, RNA abundance is measured based on the number of unique barcode sequences observed for a given cDNA sequence. We optimized the barcodes to be unambiguously identifiable, even in the presence of multiple sequencing errors. This method allows counting with single-copy resolution despite sequence-dependent bias and PCR-amplification noise, and is analogous to digital PCR but amendable to quantifying a whole transcriptome. We demonstrated transcriptome profiling of Escherichia coli with more accurate and reproducible quantification than conventional RNA-Seq. PMID:22232676

A rat RNA-Seq transcriptomic BodyMap across 11 organs and 4 developmental stages

PubMed Central

Yu, Ying; Fuscoe, James C.; Zhao, Chen; Guo, Chao; Jia, Meiwen; Qing, Tao; Bannon, Desmond I.; Lancashire, Lee; Bao, Wenjun; Du, Tingting; Luo, Heng; Su, Zhenqiang; Jones, Wendell D.; Moland, Carrie L.; Branham, William S.; Qian, Feng; Ning, Baitang; Li, Yan; Hong, Huixiao; Guo, Lei; Mei, Nan; Shi, Tieliu; Wang, Kevin Y.; Wolfinger, Russell D.; Nikolsky, Yuri; Walker, Stephen J.; Duerksen-Hughes, Penelope; Mason, Christopher E.; Tong, Weida; Thierry-Mieg, Jean; Thierry-Mieg, Danielle; Shi, Leming; Wang, Charles

2014-01-01

The rat has been used extensively as a model for evaluating chemical toxicities and for understanding drug mechanisms. However, its transcriptome across multiple organs, or developmental stages, has not yet been reported. Here we show, as part of the SEQC consortium efforts, a comprehensive rat transcriptomic BodyMap created by performing RNA-Seq on 320 samples from 11 organs of both sexes of juvenile, adolescent, adult and aged Fischer 344 rats. We catalogue the expression profiles of 40,064 genes, 65,167 transcripts, 31,909 alternatively spliced transcript variants and 2,367 non-coding genes/non-coding RNAs (ncRNAs) annotated in AceView. We find that organ-enriched, differentially expressed genes reflect the known organ-specific biological activities. A large number of transcripts show organ-specific, age-dependent or sex-specific differential expression patterns. We create a web-based, open-access rat BodyMap database of expression profiles with crosslinks to other widely used databases, anticipating that it will serve as a primary resource for biomedical research using the rat model. PMID:24510058

A molecular atlas of the developing ectoderm defines neural, neural crest, placode, and nonneural progenitor identity in vertebrates.

PubMed

Plouhinec, Jean-Louis; Medina-Ruiz, Sofía; Borday, Caroline; Bernard, Elsa; Vert, Jean-Philippe; Eisen, Michael B; Harland, Richard M; Monsoro-Burq, Anne H

2017-10-01

During vertebrate neurulation, the embryonic ectoderm is patterned into lineage progenitors for neural plate, neural crest, placodes and epidermis. Here, we use Xenopus laevis embryos to analyze the spatial and temporal transcriptome of distinct ectodermal domains in the course of neurulation, during the establishment of cell lineages. In order to define the transcriptome of small groups of cells from a single germ layer and to retain spatial information, dorsal and ventral ectoderm was subdivided along the anterior-posterior and medial-lateral axes by microdissections. Principal component analysis on the transcriptomes of these ectoderm fragments primarily identifies embryonic axes and temporal dynamics. This provides a genetic code to define positional information of any ectoderm sample along the anterior-posterior and dorsal-ventral axes directly from its transcriptome. In parallel, we use nonnegative matrix factorization to predict enhanced gene expression maps onto early and mid-neurula embryos, and specific signatures for each ectoderm area. The clustering of spatial and temporal datasets allowed detection of multiple biologically relevant groups (e.g., Wnt signaling, neural crest development, sensory placode specification, ciliogenesis, germ layer specification). We provide an interactive network interface, EctoMap, for exploring synexpression relationships among genes expressed in the neurula, and suggest several strategies to use this comprehensive dataset to address questions in developmental biology as well as stem cell or cancer research.

Cross-disease transcriptomics: Unique IL-17A signaling in psoriasis lesions and an autoimmune PBMC signature

PubMed Central

Sarkar, Mrinal K.; Liang, Yun; Xing, Xianying; Gudjonsson, Johann E.

2016-01-01

Transcriptome studies of psoriasis have identified robust changes in mRNA expression through large-scale analysis of patient cohorts. These studies, however, have analyzed all mRNA changes in aggregate, without distinguishing between disease-specific and non-specific differentially expressed genes (DEGs). In this study, RNA-seq meta-analysis was used to identify (1) psoriasis-specific DEGs altered in few diseases besides psoriasis and (2) non-specific DEGs similarly altered in many other skin conditions. We show that few cutaneous DEGs are psoriasis-specific and that the two DEG classes differ in their cell type and cytokine associations. Psoriasis-specific DEGs are expressed by keratinocytes and induced by IL-17A, whereas non-specific DEGs are expressed by inflammatory cells and induced by IFN-gamma and TNF. PBMC-derived DEGs were more psoriasis-specific than cutaneous DEGs. Nonetheless, PBMC DEGs associated with MHC class I and NK cells were commonly downregulated in psoriasis and other autoimmune diseases (e.g., multiple sclerosis, sarcoidosis and juvenile rheumatoid arthritis). These findings demonstrate “cross-disease” transcriptomics as an approach to gain insights into the cutaneous and non-cutaneous psoriasis transcriptomes. This highlighted unique contributions of IL-17A to the cytokine network and uncovered a blood-based gene signature that links psoriasis to other diseases of autoimmunity. PMID:27206706

A molecular atlas of the developing ectoderm defines neural, neural crest, placode, and nonneural progenitor identity in vertebrates

PubMed Central

Borday, Caroline; Bernard, Elsa; Vert, Jean-Philippe; Eisen, Michael B.; Harland, Richard M.

2017-01-01

During vertebrate neurulation, the embryonic ectoderm is patterned into lineage progenitors for neural plate, neural crest, placodes and epidermis. Here, we use Xenopus laevis embryos to analyze the spatial and temporal transcriptome of distinct ectodermal domains in the course of neurulation, during the establishment of cell lineages. In order to define the transcriptome of small groups of cells from a single germ layer and to retain spatial information, dorsal and ventral ectoderm was subdivided along the anterior-posterior and medial-lateral axes by microdissections. Principal component analysis on the transcriptomes of these ectoderm fragments primarily identifies embryonic axes and temporal dynamics. This provides a genetic code to define positional information of any ectoderm sample along the anterior-posterior and dorsal-ventral axes directly from its transcriptome. In parallel, we use nonnegative matrix factorization to predict enhanced gene expression maps onto early and mid-neurula embryos, and specific signatures for each ectoderm area. The clustering of spatial and temporal datasets allowed detection of multiple biologically relevant groups (e.g., Wnt signaling, neural crest development, sensory placode specification, ciliogenesis, germ layer specification). We provide an interactive network interface, EctoMap, for exploring synexpression relationships among genes expressed in the neurula, and suggest several strategies to use this comprehensive dataset to address questions in developmental biology as well as stem cell or cancer research. PMID:29049289

Prediction of G protein-coupled receptor encoding sequences from the synganglion transcriptome of the cattle tick, Rhipicephalus microplus

USDA-ARS?s Scientific Manuscript database

The cattle tick, Rhipicephalus (Boophilus) microplus, is a pest which causes multiple health complications in cattle. The G-protein coupled receptor (GPCR) super-family presents an interesting target for developing novel tick control methods. However, GPCRs share limited sequence similarity among or...

De novo transcript sequence reconstruction from RNA-Seq: reference generation and analysis with Trinity

PubMed Central

Yassour, Moran; Grabherr, Manfred; Blood, Philip D.; Bowden, Joshua; Couger, Matthew Brian; Eccles, David; Li, Bo; Lieber, Matthias; MacManes, Matthew D.; Ott, Michael; Orvis, Joshua; Pochet, Nathalie; Strozzi, Francesco; Weeks, Nathan; Westerman, Rick; William, Thomas; Dewey, Colin N.; Henschel, Robert; LeDuc, Richard D.; Friedman, Nir; Regev, Aviv

2013-01-01

De novo assembly of RNA-Seq data allows us to study transcriptomes without the need for a genome sequence, such as in non-model organisms of ecological and evolutionary importance, cancer samples, or the microbiome. In this protocol, we describe the use of the Trinity platform for de novo transcriptome assembly from RNA-Seq data in non-model organisms. We also present Trinity’s supported companion utilities for downstream applications, including RSEM for transcript abundance estimation, R/Bioconductor packages for identifying differentially expressed transcripts across samples, and approaches to identify protein coding genes. In an included tutorial we provide a workflow for genome-independent transcriptome analysis leveraging the Trinity platform. The software, documentation and demonstrations are freely available from http://trinityrnaseq.sf.net. PMID:23845962

The evolution of neuropeptide signalling: insights from echinoderms.

PubMed

Semmens, Dean C; Elphick, Maurice R

2017-09-01

Neuropeptides are evolutionarily ancient mediators of neuronal signalling that regulate a wide range of physiological processes and behaviours in animals. Neuropeptide signalling has been investigated extensively in vertebrates and protostomian invertebrates, which include the ecdysozoans Drosophila melanogaster (Phylum Arthropoda) and Caenorhabditis elegans (Phylum Nematoda). However, until recently, an understanding of evolutionary relationships between neuropeptide signalling systems in vertebrates and protostomes has been impaired by a lack of genome/transcriptome sequence data from non-ecdysozoan invertebrates. The echinoderms-a deuterostomian phylum that includes sea urchins, sea cucumbers and starfish-have been particularly important in providing new insights into neuropeptide evolution. Sequencing of the genome of the sea urchin Strongylocentrotus purpuratus (Class Echinoidea) enabled discovery of (i) the first invertebrate thyrotropin-releasing hormone-type precursor, (ii) the first deuterostomian pedal peptide/orcokinin-type precursors and (iii) NG peptides-the 'missing link' between neuropeptide S in tetrapod vertebrates and crustacean cardioactive peptide in protostomes. More recently, sequencing of the neural transcriptome of the starfish Asterias rubens (Class Asteroidea) enabled identification of 40 neuropeptide precursors, including the first kisspeptin and melanin-concentrating hormone-type precursors to be identified outside of the chordates. Furthermore, the characterization of a corazonin-type neuropeptide signalling system in A. rubens has provided important new insights into the evolution of gonadotropin-releasing hormone-related neuropeptides. Looking forward, the discovery of multiple neuropeptide signalling systems in echinoderms provides opportunities to investigate how these systems are used to regulate physiological and behavioural processes in the unique context of a decentralized, pentaradial bauplan. © The Author 2017. Published by Oxford University Press.

Assessing the Gene Content of the Megagenome: Sugar Pine (Pinus lambertiana)

PubMed Central

Gonzalez-Ibeas, Daniel; Martinez-Garcia, Pedro J.; Famula, Randi A.; Delfino-Mix, Annette; Stevens, Kristian A.; Loopstra, Carol A.; Langley, Charles H.; Neale, David B.; Wegrzyn, Jill L.

2016-01-01

Sugar pine (Pinus lambertiana Douglas) is within the subgenus Strobus with an estimated genome size of 31 Gbp. Transcriptomic resources are of particular interest in conifers due to the challenges presented in their megagenomes for gene identification. In this study, we present the first comprehensive survey of the P. lambertiana transcriptome through deep sequencing of a variety of tissue types to generate more than 2.5 billion short reads. Third generation, long reads generated through PacBio Iso-Seq have been included for the first time in conifers to combat the challenges associated with de novo transcriptome assembly. A technology comparison is provided here to contribute to the otherwise scarce comparisons of second and third generation transcriptome sequencing approaches in plant species. In addition, the transcriptome reference was essential for gene model identification and quality assessment in the parallel project responsible for sequencing and assembly of the entire genome. In this study, the transcriptomic data were also used to address questions surrounding lineage-specific Dicer-like proteins in conifers. These proteins play a role in the control of transposable element proliferation and the related genome expansion in conifers. PMID:27799338

False negative rates in Drosophila cell-based RNAi screens: a case study

PubMed Central

2011-01-01

Background High-throughput screening using RNAi is a powerful gene discovery method but is often complicated by false positive and false negative results. Whereas false positive results associated with RNAi reagents has been a matter of extensive study, the issue of false negatives has received less attention. Results We performed a meta-analysis of several genome-wide, cell-based Drosophila RNAi screens, together with a more focused RNAi screen, and conclude that the rate of false negative results is at least 8%. Further, we demonstrate how knowledge of the cell transcriptome can be used to resolve ambiguous results and how the number of false negative results can be reduced by using multiple, independently-tested RNAi reagents per gene. Conclusions RNAi reagents that target the same gene do not always yield consistent results due to false positives and weak or ineffective reagents. False positive results can be partially minimized by filtering with transcriptome data. RNAi libraries with multiple reagents per gene also reduce false positive and false negative outcomes when inconsistent results are disambiguated carefully. PMID:21251254

Byssus Structure and Protein Composition in the Highly Invasive Fouling Mussel Limnoperna fortunei

PubMed Central

Li, Shiguo; Xia, Zhiqiang; Chen, Yiyong; Gao, Yangchun; Zhan, Aibin

2018-01-01

Biofouling mediated by byssus adhesion in invasive bivalves has become a global environmental problem in aquatic ecosystems, resulting in negative ecological and economic consequences. Previous studies suggested that mechanisms responsible for byssus adhesion largely vary among bivalves, but it is poorly understood in freshwater species. Understanding of byssus structure and protein composition is the prerequisite for revealing these mechanisms. Here, we used multiple methods, including scanning electron microscope, liquid chromatography–tandem mass spectrometry, transcriptome sequencing, real-time quantitative PCR, inductively coupled plasma mass spectrometry, to investigate structure, and protein composition of byssus in the highly invasive freshwater mussel Limnoperna fortunei. The results indicated that the structure characteristics of adhesive plaque, proximal and distal threads were conducive to byssus adhesion, contributing to the high biofouling capacity of this species. The 3,4-dihydroxyphenyl-α-alanine (Dopa) is a major post-transnationally modification in L. fortunei byssus. We identified 16 representative foot proteins with typical repetitive motifs and conserved domains by integrating transcriptomic and proteomic approaches. In these proteins, Lfbp-1, Lffp-2, and Lfbp-3 were specially located in foot tissue and highly expressed in the rapid byssus formation period, suggesting the involvement of these foot proteins in byssus production and adhesion. Multiple metal irons, including Ca2+, Mg2+, Zn2+, Al3+, and Fe3+, were abundant in both foot tissue and byssal thread. The heavy metals in these irons may be directly accumulated by L. fortunei from surrounding environments. Nevertheless, some metal ions (e.g., Ca2+) corresponded well with amino acid preferences of L. fortunei foot proteins, suggesting functional roles of these metal ions by interacting with foot proteins in byssus adhesion. Overall, this study provides structural and molecular bases of adhesive mechanisms of byssus in L. fortunei, and findings here are expected to develop strategies against biofouling by freshwater organisms. PMID:29713291

Mouse models rarely mimic the transcriptome of human neurodegenerative diseases: A systematic bioinformatics-based critique of preclinical models.

PubMed

Burns, Terry C; Li, Matthew D; Mehta, Swapnil; Awad, Ahmed J; Morgan, Alexander A

2015-07-15

Translational research for neurodegenerative disease depends intimately upon animal models. Unfortunately, promising therapies developed using mouse models mostly fail in clinical trials, highlighting uncertainty about how well mouse models mimic human neurodegenerative disease at the molecular level. We compared the transcriptional signature of neurodegeneration in mouse models of Alzheimer׳s disease (AD), Parkinson׳s disease (PD), Huntington׳s disease (HD) and amyotrophic lateral sclerosis (ALS) to human disease. In contrast to aging, which demonstrated a conserved transcriptome between humans and mice, only 3 of 19 animal models showed significant enrichment for gene sets comprising the most dysregulated up- and down-regulated human genes. Spearman׳s correlation analysis revealed even healthy human aging to be more closely related to human neurodegeneration than any mouse model of AD, PD, ALS or HD. Remarkably, mouse models frequently upregulated stress response genes that were consistently downregulated in human diseases. Among potential alternate models of neurodegeneration, mouse prion disease outperformed all other disease-specific models. Even among the best available animal models, conserved differences between mouse and human transcriptomes were found across multiple animal model versus human disease comparisons, surprisingly, even including aging. Relative to mouse models, mouse disease signatures demonstrated consistent trends toward preserved mitochondrial function protein catabolism, DNA repair responses, and chromatin maintenance. These findings suggest a more complex and multifactorial pathophysiology in human neurodegeneration than is captured through standard animal models, and suggest that even among conserved physiological processes such as aging, mice are less prone to exhibit neurodegeneration-like changes. This work may help explain the poor track record of mouse-based translational therapies for neurodegeneration and provides a path forward to critically evaluate and improve animal models of human disease. Copyright © 2015 Elsevier B.V. All rights reserved.

Transcriptomic Analysis Reveals Mechanisms of Sterile and Fertile Flower Differentiation and Development in Viburnum macrocephalum f. keteleeri

PubMed Central

Lu, Zhaogeng; Xu, Jing; Li, Weixing; Zhang, Li; Cui, Jiawen; He, Qingsong; Wang, Li; Jin, Biao

2017-01-01

Sterile and fertile flowers are an important evolutionary developmental (evo-devo) phenotype in angiosperm flowers, playing important roles in pollinator attraction and sexual reproductive success. However, the gene regulatory mechanisms underlying fertile and sterile flower differentiation and development remain largely unknown. Viburnum macrocephalum f. keteleeri, which possesses fertile and sterile flowers in a single inflorescence, is a useful candidate species for investigating the regulatory networks in differentiation and development. We developed a de novo-assembled flower reference transcriptome. Using RNA sequencing (RNA-seq), we compared the expression patterns of fertile and sterile flowers isolated from the same inflorescence over its rapid developmental stages. The flower reference transcriptome consisted of 105,683 non-redundant transcripts, of which 5,675 transcripts showed significant differential expression between fertile and sterile flowers. Combined with morphological and cytological changes between fertile and sterile flowers, we identified expression changes of many genes potentially involved in reproductive processes, phytohormone signaling, and cell proliferation and expansion using RNA-seq and qRT-PCR. In particular, many transcription factors (TFs), including MADS-box family members and ABCDE-class genes, were identified, and expression changes in TFs involved in multiple functions were analyzed and highlighted to determine their roles in regulating fertile and sterile flower differentiation and development. Our large-scale transcriptional analysis of fertile and sterile flowers revealed the dynamics of transcriptional networks and potentially key components in regulating differentiation and development of fertile and sterile flowers in Viburnum macrocephalum f. keteleeri. Our data provide a useful resource for Viburnum transcriptional research and offer insights into gene regulation of differentiation of diverse evo-devo processes in flowers. PMID:28298915

The Carcinogenic Liver Fluke, Clonorchis sinensis: New Assembly, Reannotation and Analysis of the Genome and Characterization of Tissue Transcriptomes

PubMed Central

Wang, Xiaoyun; Liu, Hailiang; Chen, Yangyi; Guo, Lei; Luo, Fang; Sun, Jiufeng; Mao, Qiang; Liang, Pei; Xie, Zhizhi; Zhou, Chenhui; Tian, Yanli; Lv, Xiaoli; Huang, Lisi; Zhou, Juanjuan; Hu, Yue; Li, Ran; Zhang, Fan; Lei, Huali; Li, Wenfang; Hu, Xuchu; Liang, Chi; Xu, Jin; Li, Xuerong; Yu, Xinbing

2013-01-01

Clonorchis sinensis (C. sinensis), an important food-borne parasite that inhabits the intrahepatic bile duct and causes clonorchiasis, is of interest to both the public health field and the scientific research community. To learn more about the migration, parasitism and pathogenesis of C. sinensis at the molecular level, the present study developed an upgraded genomic assembly and annotation by sequencing paired-end and mate-paired libraries. We also performed transcriptome sequence analyses on multiple C. sinensis tissues (sucker, muscle, ovary and testis). Genes encoding molecules involved in responses to stimuli and muscle-related development were abundantly expressed in the oral sucker. Compared with other species, genes encoding molecules that facilitate the recognition and transport of cholesterol were observed in high copy numbers in the genome and were highly expressed in the oral sucker. Genes encoding transporters for fatty acids, glucose, amino acids and oxygen were also highly expressed, along with other molecules involved in metabolizing these substrates. All genes involved in energy metabolism pathways, including the β-oxidation of fatty acids, the citrate cycle, oxidative phosphorylation, and fumarate reduction, were expressed in the adults. Finally, we also provide valuable insights into the mechanism underlying the process of pathogenesis by characterizing the secretome of C. sinensis. The characterization and elaborate analysis of the upgraded genome and the tissue transcriptomes not only form a detailed and fundamental C. sinensis resource but also provide novel insights into the physiology and pathogenesis of C. sinensis. We anticipate that this work will aid the development of innovative strategies for the prevention and control of clonorchiasis. PMID:23382950

Multi-level evaluation of Escherichia coli polyphosphate related mutants using global transcriptomic, proteomic and phenomic analyses.

PubMed

Varas, Macarena; Valdivieso, Camilo; Mauriaca, Cecilia; Ortíz-Severín, Javiera; Paradela, Alberto; Poblete-Castro, Ignacio; Cabrera, Ricardo; Chávez, Francisco P

2017-04-01

Polyphosphate (polyP) is a linear biopolymer found in all living cells. In bacteria, mutants lacking polyphosphate kinase 1 (PPK1), the enzyme responsible for synthesis of most polyP, have many structural and functional defects. However, little is known about the causes of these pleiotropic alterations. The link between ppk1 deletion and those numerous phenotypes observed can be the result of complex molecular interactions that can be elucidated via a systems biology approach. By integrating different omics levels (transcriptome, proteome and phenome), we described the functioning of various metabolic pathways among Escherichia coli polyphosphate mutant strains (Δppk1, Δppx, and ΔpolyP). Bioinformatic analyses reveal the complex metabolic and regulatory bases of the phenotypes unique to polyP mutants. Our results suggest that during polyP deficiency (Δppk1 mutant), metabolic pathways needed for energy supply are up-regulated, including fermentation, aerobic and anaerobic respiration. Transcriptomic and q-proteomic contrasting changes between Δppk1 and Δppx mutant strains were observed in those central metabolic pathways and confirmed by using Phenotypic microarrays. In addition, our results suggest a regulatory connection between polyP, second messenger metabolism, alternative Sigma/Anti-Sigma factors and type-II toxin-antitoxin (TA) systems. We suggest a broader role for polyP via regulation of ATP-dependent proteolysis of type II toxin-antitoxin system and alternative Sigma/Anti-Sigma factors, that could explain the multiple structural and functional deficiencies described due to alteration of polyP metabolism. Understanding the interplay of polyP in bacterial metabolism using a systems biology approach can help to improve design of novel antimicrobials toward pathogens. Copyright © 2017 Elsevier B.V. All rights reserved.

Chlorophyll a fluorescence and transcriptome reveal the toxicological effects of bisphenol A on an invasive cyanobacterium, Cylindrospermopsis raciborskii.

PubMed

Xiang, Rong; Shi, Junqiong; Zhang, Hongbo; Dong, Congcong; Liu, Li; Fu, JunKe; He, Xinyu; Yan, Yanjun; Wu, Zhongxing

2018-05-09

Bisphenol A has attracted worldwide attention due to its harmful effects on humans, animals and plants. In this study, the toxicological effects of BPA on Cylindrospermopsis raciborskii were assessed based on chlorophyll a fluorescence and transcriptome analyses. The results showed that the growth of C. raciborskii was significantly inhibited when BPA exceeded 0.1 mg L -1 . A marked rise of phase J was observed at a concentration greater than 0.1 mg L -1 , while a K phase appeared at 20 mg L -1 . The chlorophyll a fluorescence parameters of RC/CS 0 , F 0 , φ P0 , φ E0 , and ψ 0 , underwent a significant decline under all treatments of BPA, whereas a significant increase in both V J and M 0 occurred under all concentrations of BPA. Additionally, ABS/RC and DIo/RC markedly increased at 10 mg L -1 and 20 mg L -1 . The transcriptome analysis revealed that the genes of photosynthesis, including psbA, psbB, psbC, psbD, apcA, apcB, cpcA, and cpcB, as well as those of chlorophyll and carotenoid biosynthesis, namely hemN, acsF, chlL, chlN, chlP, crtB, pds, were all down-regulated. Moreover, BPA also inhibited the oxidative phosphorylation, glycolysis/gluconeogenesis, citrate cycle (TCA cycle), and fatty acid metabolism in C. raciborskii. Taken together, these results suggest BPA can negatively affect the expression of multiple genes and the vital energy metabolism process to arrest the growth and photosynthesis of C. raciborskii. Copyright © 2018 Elsevier B.V. All rights reserved.

In-depth characterization of the microRNA transcriptome in a leukemia progression model

PubMed Central

Kuchenbauer, Florian; Morin, Ryan D.; Argiropoulos, Bob; Petriv, Oleh I.; Griffith, Malachi; Heuser, Michael; Yung, Eric; Piper, Jessica; Delaney, Allen; Prabhu, Anna-Liisa; Zhao, Yongjun; McDonald, Helen; Zeng, Thomas; Hirst, Martin; Hansen, Carl L.; Marra, Marco A.; Humphries, R. Keith

2008-01-01

MicroRNAs (miRNAs) have been shown to play important roles in physiological as well as multiple malignant processes, including acute myeloid leukemia (AML). In an effort to gain further insight into the role of miRNAs in AML, we have applied the Illumina massively parallel sequencing platform to carry out an in-depth analysis of the miRNA transcriptome in a murine leukemia progression model. This model simulates the stepwise conversion of a myeloid progenitor cell by an engineered overexpression of the nucleoporin 98 (NUP98)–homeobox HOXD13 fusion gene (ND13), to aggressive AML inducing cells upon transduction with the oncogenic collaborator Meis1. From this data set, we identified 307 miRNA/miRNA* species in the ND13 cells and 306 miRNA/miRNA* species in ND13+Meis1 cells, corresponding to 223 and 219 miRNA genes. Sequence counts varied between two and 136,558, indicating a remarkable expression range between the detected miRNA species. The large number of miRNAs expressed and the nature of differential expression suggest that leukemic progression as modeled here is dictated by the repertoire of shared, but differentially expressed miRNAs. Our finding of extensive sequence variations (isomiRs) for almost all miRNA and miRNA* species adds additional complexity to the miRNA transcriptome. A stringent target prediction analysis coupled with in vitro target validation revealed the potential for miRNA-mediated release of oncogenes that facilitates leukemic progression from the preleukemic to leukemia inducing state. Finally, 55 novel miRNAs species were identified in our data set, adding further complexity to the emerging world of small RNAs. PMID:18849523

Characterization of the Pratylenchus penetrans transcriptome including data mining of putative nematode genes involved in plant parasitism

USDA-ARS?s Scientific Manuscript database

The root lesion nematode Pratylenchus penetrans is considered one of the most economically important species within the genus. Host range studies have shown that nearly 400 plant species can be parasitized by this species. To obtain insight into the transcriptome of this migratory plant-parasitic ne...

Transcriptome of Aspergillus flavus aswA (AFLA_085170) deletion strain related to sclerotial development and production of secondary metabolites

USDA-ARS?s Scientific Manuscript database

Aspergillus flavus produces many secondary metabolites including aflatoxins. Besides conidia, the fungus uses sclerotia as another type of propagule. We obtained transcriptomes from four growth conditions of the aswA mutant, a strain impaired in sclerotial development and production of sclerotium-sp...

Transcriptome sequencing of newly molted adult female cattle ticks, Rhipicephalus microplus: Raw Illumina reads.

USDA-ARS?s Scientific Manuscript database

Illumina paired end oligo-dT sequencing technology was used to sequence the transcriptome from newly molted adult females from the cattle tick, Rhipicephalus microplus. These samples include newly molted unfed whole adult females, newly molted whole adult females feeding for 2 hours on a bovine host...

Comparative Transcriptomes and EVO-DEVO Studies Depending on Next Generation Sequencing.

PubMed

Liu, Tiancheng; Yu, Lin; Liu, Lei; Li, Hong; Li, Yixue

2015-01-01

High throughput technology has prompted the progressive omics studies, including genomics and transcriptomics. We have reviewed the improvement of comparative omic studies, which are attributed to the high throughput measurement of next generation sequencing technology. Comparative genomics have been successfully applied to evolution analysis while comparative transcriptomics are adopted in comparison of expression profile from two subjects by differential expression or differential coexpression, which enables their application in evolutionary developmental biology (EVO-DEVO) studies. EVO-DEVO studies focus on the evolutionary pressure affecting the morphogenesis of development and previous works have been conducted to illustrate the most conserved stages during embryonic development. Old measurements of these studies are based on the morphological similarity from macro view and new technology enables the micro detection of similarity in molecular mechanism. Evolutionary model of embryo development, which includes the "funnel-like" model and the "hourglass" model, has been evaluated by combination of these new comparative transcriptomic methods with prior comparative genomic information. Although the technology has promoted the EVO-DEVO studies into a new era, technological and material limitation still exist and further investigations require more subtle study design and procedure.

A General Framework for Interrogation of mRNA Stability Programs Identifies RNA-Binding Proteins that Govern Cancer Transcriptomes.

PubMed

Perron, Gabrielle; Jandaghi, Pouria; Solanki, Shraddha; Safisamghabadi, Maryam; Storoz, Cristina; Karimzadeh, Mehran; Papadakis, Andreas I; Arseneault, Madeleine; Scelo, Ghislaine; Banks, Rosamonde E; Tost, Jorg; Lathrop, Mark; Tanguay, Simon; Brazma, Alvis; Huang, Sidong; Brimo, Fadi; Najafabadi, Hamed S; Riazalhosseini, Yasser

2018-05-08

Widespread remodeling of the transcriptome is a signature of cancer; however, little is known about the post-transcriptional regulatory factors, including RNA-binding proteins (RBPs) that regulate mRNA stability, and the extent to which RBPs contribute to cancer-associated pathways. Here, by modeling the global change in gene expression based on the effect of sequence-specific RBPs on mRNA stability, we show that RBP-mediated stability programs are recurrently deregulated in cancerous tissues. Particularly, we uncovered several RBPs that contribute to the abnormal transcriptome of renal cell carcinoma (RCC), including PCBP2, ESRP2, and MBNL2. Modulation of these proteins in cancer cell lines alters the expression of pathways that are central to the disease and highlights RBPs as driving master regulators of RCC transcriptome. This study presents a framework for the screening of RBP activities based on computational modeling of mRNA stability programs in cancer and highlights the role of post-transcriptional gene dysregulation in RCC. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Analysis of insecticide resistance-related genes of the Carmine spider mite Tetranychus cinnabarinus based on a de novo assembled transcriptome.

PubMed

Xu, Zhifeng; Zhu, Wenyi; Liu, Yanchao; Liu, Xing; Chen, Qiushuang; Peng, Miao; Wang, Xiangzun; Shen, Guangmao; He, Lin

2014-01-01

The carmine spider mite (CSM), Tetranychus cinnabarinus, is an important pest mite in agriculture, because it can develop insecticide resistance easily. To gain valuable gene information and molecular basis for the future insecticide resistance study of CSM, the first transcriptome analysis of CSM was conducted. A total of 45,016 contigs and 25,519 unigenes were generated from the de novo transcriptome assembly, and 15,167 unigenes were annotated via BLAST querying against current databases, including nr, SwissProt, the Clusters of Orthologous Groups (COGs), Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO). Aligning the transcript to Tetranychus urticae genome, the 19255 (75.45%) of the transcripts had significant (e-value <10-5) matches to T. urticae DNA genome, 19111 sequences matched to T. urticae proteome with an average protein length coverage of 42.55%. Core Eukaryotic Genes Mapping Approach (CEGMA) analysis identified 435 core eukaryotic genes (CEGs) in the CSM dataset corresponding to 95% coverage. Ten gene categories that relate to insecticide resistance in arthropod were generated from CSM transcriptome, including 53 P450-, 22 GSTs-, 23 CarEs-, 1 AChE-, 7 GluCls-, 9 nAChRs-, 8 GABA receptor-, 1 sodium channel-, 6 ATPase- and 12 Cyt b genes. We developed significant molecular resources for T. cinnabarinus putatively involved in insecticide resistance. The transcriptome assembly analysis will significantly facilitate our study on the mechanism of adapting environmental stress (including insecticide) in CSM at the molecular level, and will be very important for developing new control strategies against this pest mite.

Analyses of advanced rice anther transcriptomes reveal global tapetum secretory functions and potential proteins for lipid exine formation.

PubMed

Huang, Ming-Der; Wei, Fu-Jin; Wu, Cheng-Cheih; Hsing, Yue-Ie Caroline; Huang, Anthony H C

2009-02-01

The anthers in flowers perform important functions in sexual reproduction. Several recent studies used microarrays to study anther transcriptomes to explore genes controlling anther development. To analyze the secretion and other functions of the tapetum, we produced transcriptomes of anthers of rice (Oryza sativa subsp. japonica) at six progressive developmental stages and pollen with sequencing-by-synthesis technology. The transcriptomes included at least 18,000 unique transcripts, about 25% of which had antisense transcripts. In silico anther-minus-pollen subtraction produced transcripts largely unique to the tapetum; these transcripts include all the reported tapetum-specific transcripts of orthologs in other species. The differential developmental profiles of the transcripts and their antisense transcripts signify extensive regulation of gene expression in the anther, especially the tapetum, during development. The transcriptomes were used to dissect two major cell/biochemical functions of the tapetum. First, we categorized and charted the developmental profiles of all transcripts encoding secretory proteins present in the cellular exterior; these transcripts represent about 12% and 30% of the those transcripts having more than 100 and 1,000 transcripts per million, respectively. Second, we successfully selected from hundreds of transcripts several transcripts encoding potential proteins for lipid exine synthesis during early anther development. These proteins include cytochrome P450, acyltransferases, and lipid transfer proteins in our hypothesized mechanism of exine synthesis in and export from the tapetum. Putative functioning of these proteins in exine formation is consistent with proteins and metabolites detected in the anther locule fluid obtained by micropipetting.

A house finch (Haemorhous mexicanus) spleen transcriptome reveals intra- and interspecific patterns of gene expression, alternative splicing and genetic diversity in passerines.

PubMed

Zhang, Qu; Hill, Geoffrey E; Edwards, Scott V; Backström, Niclas

2014-04-24

With its plumage color dimorphism and unique history in North America, including a recent population expansion and an epizootic of Mycoplasma gallisepticum (MG), the house finch (Haemorhous mexicanus) is a model species for studying sexual selection, plumage coloration and host-parasite interactions. As part of our ongoing efforts to make available genomic resources for this species, here we report a transcriptome assembly derived from genes expressed in spleen. We characterize transcriptomes from two populations with different histories of demography and disease exposure: a recently founded population in the eastern US that has been exposed to MG for over a decade and a native population from the western range that has never been exposed to MG. We utilize this resource to quantify conservation in gene expression in passerine birds over approximately 50 MY by comparing splenic expression profiles for 9,646 house finch transcripts and those from zebra finch and find that less than half of all genes expressed in spleen in either species are expressed in both species. Comparative gene annotations from several vertebrate species suggest that the house finch transcriptomes contain ~15 genes not yet found in previously sequenced vertebrate genomes. The house finch transcriptomes harbour ~85,000 SNPs, ~20,000 of which are non-synonymous. Although not yet validated by biological or technical replication, we identify a set of genes exhibiting differences between populations in gene expression (n = 182; 2% of all transcripts), allele frequencies (76 FST ouliers) and alternative splicing as well as genes with several fixed non-synonymous substitutions; this set includes genes with functions related to double-strand break repair and immune response. The two house finch spleen transcriptome profiles will add to the increasing data on genome and transcriptome sequence information from natural populations. Differences in splenic expression between house finch and zebra finch imply either significant evolutionary turnover of splenic expression patterns or different physiological states of the individuals examined. The transcriptome resource will enhance the potential to annotate an eventual house finch genome, and the set of gene-based high-quality SNPs will help clarify the genetic underpinnings of host-pathogen interactions and sexual selection.

CRISPR/Cas9-mediated heterozygous knockout of the autism gene CHD8 and characterization of its transcriptional networks in neurodevelopment.

PubMed

Wang, Ping; Lin, Mingyan; Pedrosa, Erika; Hrabovsky, Anastasia; Zhang, Zheng; Guo, Wenjun; Lachman, Herbert M; Zheng, Deyou

2015-01-01

Disruptive mutation in the CHD8 gene is one of the top genetic risk factors in autism spectrum disorders (ASDs). Previous analyses of genome-wide CHD8 occupancy and reduced expression of CHD8 by shRNA knockdown in committed neural cells showed that CHD8 regulates multiple cell processes critical for neural functions, and its targets are enriched with ASD-associated genes. To further understand the molecular links between CHD8 functions and ASD, we have applied the CRISPR/Cas9 technology to knockout one copy of CHD8 in induced pluripotent stem cells (iPSCs) to better mimic the loss-of-function status that would exist in the developing human embryo prior to neuronal differentiation. We then carried out transcriptomic and bioinformatic analyses of neural progenitors and neurons derived from the CHD8 mutant iPSCs. Transcriptome profiling revealed that CHD8 hemizygosity (CHD8 (+/-)) affected the expression of several thousands of genes in neural progenitors and early differentiating neurons. The differentially expressed genes were enriched for functions of neural development, β-catenin/Wnt signaling, extracellular matrix, and skeletal system development. They also exhibited significant overlap with genes previously associated with autism and schizophrenia, as well as the downstream transcriptional targets of multiple genes implicated in autism. Providing important insight into how CHD8 mutations might give rise to macrocephaly, we found that seven of the twelve genes associated with human brain volume or head size by genome-wide association studies (e.g., HGMA2) were dysregulated in CHD8 (+/-) neural progenitors or neurons. We have established a renewable source of CHD8 (+/-) iPSC lines that would be valuable for investigating the molecular and cellular functions of CHD8. Transcriptomic profiling showed that CHD8 regulates multiple genes implicated in ASD pathogenesis and genes associated with brain volume.

Transcriptional Analysis of Fracture Healing and the Induction of Embryonic Stem Cell–Related Genes

PubMed Central

Bais, Manish; McLean, Jody; Sebastiani, Paola; Young, Megan; Wigner, Nathan; Smith, Temple; Kotton, Darrell N.; Einhorn, Thomas A.; Gerstenfeld, Louis C.

2009-01-01

Fractures are among the most common human traumas. Fracture healing represents a unique temporarily definable post-natal process in which to study the complex interactions of multiple molecular events that regulate endochondral skeletal tissue formation. Because of the regenerative nature of fracture healing, it is hypothesized that large numbers of post-natal stem cells are recruited and contribute to formation of the multiple cell lineages that contribute to this process. Bayesian modeling was used to generate the temporal profiles of the transcriptome during fracture healing. The temporal relationships between ontologies that are associated with various biologic, metabolic, and regulatory pathways were identified and related to developmental processes associated with skeletogenesis, vasculogenesis, and neurogenesis. The complement of all the expressed BMPs, Wnts, FGFs, and their receptors were related to the subsets of transcription factors that were concurrently expressed during fracture healing. We further defined during fracture healing the temporal patterns of expression for 174 of the 193 genes known to be associated with human genetic skeletal disorders. In order to identify the common regulatory features that might be present in stem cells that are recruited during fracture healing to other types of stem cells, we queried the transcriptome of fracture healing against that seen in embryonic stem cells (ESCs) and mesenchymal stem cells (MSCs). Approximately 300 known genes that are preferentially expressed in ESCs and ∼350 of the known genes that are preferentially expressed in MSCs showed induction during fracture healing. Nanog, one of the central epigenetic regulators associated with ESC stem cell maintenance, was shown to be associated in multiple forms or bone repair as well as MSC differentiation. In summary, these data present the first temporal analysis of the transcriptome of an endochondral bone formation process that takes place during fracture healing. They show that neurogenesis as well as vasculogenesis are predominant components of skeletal tissue formation and suggest common pathways are shared between post-natal stem cells and those seen in ESCs. PMID:19415118

Histological eosinophilic gastritis is a systemic disorder associated with blood and extra-gastric eosinophilia, Th2 immunity, and a unique gastric transcriptome

PubMed Central

Caldwell, Julie M.; Collins, Margaret H.; Stucke, Emily M.; Putnam, Philip E.; Franciosi, James P.; Kushner, Jonathan P.; Abonia, J. Pablo; Rothenberg, Marc E.

2014-01-01

Background The definition of eosinophilic gastritis (EG) is currently limited to histological EG based on the tissue eosinophil count. Objective We aimed to provide additional fundamental information about the molecular, histopathological, and clinical characteristics of EG. Methods Genome-wide transcript profiles and histological features of gastric biopsies as well as blood eosinophil numbers were analyzed in EG and control patients (n = 15 each). Results The peak gastric antrum eosinophil count was 282.7 ± 163.9 eosinophils/400X high-power field (HPF) in EG and 11.0 ± 8.5 eosinophils/HPF in control patients (P = 6.1 × 10−7). EG patients (87%) had co-existing eosinophilic inflammation in multiple gastrointestinal segments; the esophagus represented the most common secondary site. Elevated peripheral blood eosinophil numbers (EG 1.09 ± 0.88 × 103 [K]/μl vs. control 0.09 ± 0.08 K/μl, P = .0027) positively correlated with peak gastric eosinophil counts (Pearson r2 = .8102, P < .0001). MIB-1+ (proliferating), CD117+ (mast cells), and FOXP3+ cells (regulatory and/or activated T cells) were increased in EG. Transcript profiling revealed changes in 8% of the genome in EG gastric tissue. Only 7% of this EG transcriptome overlapped with the eosinophilic esophagitis (EoE) transcriptome. Significantly increased IL4, IL5, IL13, IL17, CCL26 and mast cell-specific transcripts and decreased IL33 were observed. Conclusion EG is a systemic disorder involving profound blood and gastrointestinal tract eosinophilia, Th2 immunity, and a conserved gastric transcriptome markedly distinct from the EoE transcriptome. The data herein define germane cellular and molecular pathways of EG and provide a basis for improving diagnosis and treatment. PMID:25234644

Leveraging CyVerse Resources for De Novo Comparative Transcriptomics of Underserved (Non-model) Organisms

PubMed Central

Joyce, Blake L.; Haug-Baltzell, Asher K.; Hulvey, Jonathan P.; McCarthy, Fiona; Devisetty, Upendra Kumar; Lyons, Eric

2017-01-01

This workflow allows novice researchers to leverage advanced computational resources such as cloud computing to carry out pairwise comparative transcriptomics. It also serves as a primer for biologists to develop data scientist computational skills, e.g. executing bash commands, visualization and management of large data sets. All command line code and further explanations of each command or step can be found on the wiki (https://wiki.cyverse.org/wiki/x/dgGtAQ). The Discovery Environment and Atmosphere platforms are connected together through the CyVerse Data Store. As such, once the initial raw sequencing data has been uploaded there is no more need to transfer large data files over an Internet connection, minimizing the amount of time needed to conduct analyses. This protocol is designed to analyze only two experimental treatments or conditions. Differential gene expression analysis is conducted through pairwise comparisons, and will not be suitable to test multiple factors. This workflow is also designed to be manual rather than automated. Each step must be executed and investigated by the user, yielding a better understanding of data and analytical outputs, and therefore better results for the user. Once complete, this protocol will yield de novo assembled transcriptome(s) for underserved (non-model) organisms without the need to map to previously assembled reference genomes (which are usually not available in underserved organism). These de novo transcriptomes are further used in pairwise differential gene expression analysis to investigate genes differing between two experimental conditions. Differentially expressed genes are then functionally annotated to understand the genetic response organisms have to experimental conditions. In total, the data derived from this protocol is used to test hypotheses about biological responses of underserved organisms. PMID:28518075

Modeling hormonal and inflammatory contributions to preterm and term labor using uterine temporal transcriptomics.

PubMed

Migale, Roberta; MacIntyre, David A; Cacciatore, Stefano; Lee, Yun S; Hagberg, Henrik; Herbert, Bronwen R; Johnson, Mark R; Peebles, Donald; Waddington, Simon N; Bennett, Phillip R

2016-06-13

Preterm birth is now recognized as the primary cause of infant mortality worldwide. Interplay between hormonal and inflammatory signaling in the uterus modulates the onset of contractions; however, the relative contribution of each remains unclear. In this study we aimed to characterize temporal transcriptome changes in the uterus preceding term labor and preterm labor (PTL) induced by progesterone withdrawal or inflammation in the mouse and compare these findings with human data. Myometrium was collected at multiple time points during gestation and labor from three murine models of parturition: (1) term gestation; (2) PTL induced by RU486; and (3) PTL induced by lipopolysaccharide (LPS). RNA was extracted and cDNA libraries were prepared and sequenced using the Illumina HiSeq 2000 system. Resulting RNA-Seq data were analyzed using multivariate modeling approaches as well as pathway and causal network analyses and compared against human myometrial transcriptome data. We identified a core set of temporal myometrial gene changes associated with term labor and PTL in the mouse induced by either inflammation or progesterone withdrawal. Progesterone withdrawal initiated labor without inflammatory gene activation, yet LPS activation of uterine inflammation was sufficient to override the repressive effects of progesterone and induce a laboring phenotype. Comparison of human and mouse uterine transcriptomic datasets revealed that human labor more closely resembles inflammation-induced PTL in the mouse. Labor in the mouse can be achieved through inflammatory gene activation yet these changes are not a requisite for labor itself. Human labor more closely resembles LPS-induced PTL in the mouse, supporting an essential role for inflammatory mediators in human "functional progesterone withdrawal." This improved understanding of inflammatory and progesterone influence on the uterine transcriptome has important implications for the development of PTL prevention strategies.

Deep Sequencing Reveals Uncharted Isoform Heterogeneity of the Protein-Coding Transcriptome in Cerebral Ischemia.

PubMed

Bhattarai, Sunil; Aly, Ahmed; Garcia, Kristy; Ruiz, Diandra; Pontarelli, Fabrizio; Dharap, Ashutosh

2018-06-03

Gene expression in cerebral ischemia has been a subject of intense investigations for several years. Studies utilizing probe-based high-throughput methodologies such as microarrays have contributed significantly to our existing knowledge but lacked the capacity to dissect the transcriptome in detail. Genome-wide RNA-sequencing (RNA-seq) enables comprehensive examinations of transcriptomes for attributes such as strandedness, alternative splicing, alternative transcription start/stop sites, and sequence composition, thus providing a very detailed account of gene expression. Leveraging this capability, we conducted an in-depth, genome-wide evaluation of the protein-coding transcriptome of the adult mouse cortex after transient focal ischemia at 6, 12, or 24 h of reperfusion using RNA-seq. We identified a total of 1007 transcripts at 6 h, 1878 transcripts at 12 h, and 1618 transcripts at 24 h of reperfusion that were significantly altered as compared to sham controls. With isoform-level resolution, we identified 23 splice variants arising from 23 genes that were novel mRNA isoforms. For a subset of genes, we detected reperfusion time-point-dependent splice isoform switching, indicating an expression and/or functional switch for these genes. Finally, for 286 genes across all three reperfusion time-points, we discovered multiple, distinct, simultaneously expressed and differentially altered isoforms per gene that were generated via alternative transcription start/stop sites. Of these, 165 isoforms derived from 109 genes were novel mRNAs. Together, our data unravel the protein-coding transcriptome of the cerebral cortex at an unprecedented depth to provide several new insights into the flexibility and complexity of stroke-related gene transcription and transcript organization.

The First Chameleon Transcriptome: Comparative Genomic Analysis of the OXPHOS System Reveals Loss of COX8 in Iguanian Lizards

PubMed Central

Bar-Yaacov, Dan; Bouskila, Amos; Mishmar, Dan

2013-01-01

Recently, we found dramatic mitochondrial DNA divergence of Israeli Chamaeleo chamaeleon populations into two geographically distinct groups. We aimed to examine whether the same pattern of divergence could be found in nuclear genes. However, no genomic resource is available for any chameleon species. Here we present the first chameleon transcriptome, obtained using deep sequencing (SOLiD). Our analysis identified 164,000 sequence contigs of which 19,000 yielded unique BlastX hits. To test the efficacy of our sequencing effort, we examined whether the chameleon and other available reptilian transcriptomes harbored complete sets of genes comprising known biochemical pathways, focusing on the nDNA-encoded oxidative phosphorylation (OXPHOS) genes as a model. As a reference for the screen, we used the human 86 (including isoforms) known structural nDNA-encoded OXPHOS subunits. Analysis of 34 publicly available vertebrate transcriptomes revealed orthologs for most human OXPHOS genes. However, OXPHOS subunit COX8 (Cytochrome C oxidase subunit 8), including all its known isoforms, was consistently absent in transcriptomes of iguanian lizards, implying loss of this subunit during the radiation of this suborder. The lack of COX8 in the suborder Iguania is intriguing, since it is important for cellular respiration and ATP production. Our sequencing effort added a new resource for comparative genomic studies, and shed new light on the evolutionary dynamics of the OXPHOS system. PMID:24009133

The first Chameleon transcriptome: comparative genomic analysis of the OXPHOS system reveals loss of COX8 in Iguanian lizards.

PubMed

Bar-Yaacov, Dan; Bouskila, Amos; Mishmar, Dan

2013-01-01

Recently, we found dramatic mitochondrial DNA divergence of Israeli Chamaeleo chamaeleon populations into two geographically distinct groups. We aimed to examine whether the same pattern of divergence could be found in nuclear genes. However, no genomic resource is available for any chameleon species. Here we present the first chameleon transcriptome, obtained using deep sequencing (SOLiD). Our analysis identified 164,000 sequence contigs of which 19,000 yielded unique BlastX hits. To test the efficacy of our sequencing effort, we examined whether the chameleon and other available reptilian transcriptomes harbored complete sets of genes comprising known biochemical pathways, focusing on the nDNA-encoded oxidative phosphorylation (OXPHOS) genes as a model. As a reference for the screen, we used the human 86 (including isoforms) known structural nDNA-encoded OXPHOS subunits. Analysis of 34 publicly available vertebrate transcriptomes revealed orthologs for most human OXPHOS genes. However, OXPHOS subunit COX8 (Cytochrome C oxidase subunit 8), including all its known isoforms, was consistently absent in transcriptomes of iguanian lizards, implying loss of this subunit during the radiation of this suborder. The lack of COX8 in the suborder Iguania is intriguing, since it is important for cellular respiration and ATP production. Our sequencing effort added a new resource for comparative genomic studies, and shed new light on the evolutionary dynamics of the OXPHOS system.

Increasing the source/sink ratio in Vitis vinifera (cv Sangiovese) induces extensive transcriptome reprogramming and modifies berry ripening

PubMed Central

2011-01-01

Background Cluster thinning is an agronomic practice in which a proportion of berry clusters are removed from the vine to increase the source/sink ratio and improve the quality of the remaining berries. Until now no transcriptomic data have been reported describing the mechanisms that underlie the agronomic and biochemical effects of thinning. Results We profiled the transcriptome of Vitis vinifera cv. Sangiovese berries before and after thinning at veraison using a genome-wide microarray representing all grapevine genes listed in the latest V1 gene prediction. Thinning increased the source/sink ratio from 0.6 to 1.2 m2 leaf area per kg of berries and boosted the sugar and anthocyanin content at harvest. Extensive transcriptome remodeling was observed in thinned vines 2 weeks after thinning and at ripening. This included the enhanced modulation of genes that are normally regulated during berry development and the induction of a large set of genes that are not usually expressed. Conclusion Cluster thinning has a profound effect on several important cellular processes and metabolic pathways including carbohydrate metabolism and the synthesis and transport of secondary products. The integrated agronomic, biochemical and transcriptomic data revealed that the positive impact of cluster thinning on final berry composition reflects a much more complex outcome than simply enhancing the normal ripening process. PMID:22192855

Transcriptomic Studies of Malaria: a Paradigm for Investigation of Systemic Host-Pathogen Interactions

PubMed Central

2018-01-01

SUMMARY Transcriptomics, the analysis of genome-wide RNA expression, is a common approach to investigate host and pathogen processes in infectious diseases. Technical and bioinformatic advances have permitted increasingly thorough analyses of the association of RNA expression with fundamental biology, immunity, pathogenesis, diagnosis, and prognosis. Transcriptomic approaches can now be used to realize a previously unattainable goal, the simultaneous study of RNA expression in host and pathogen, in order to better understand their interactions. This exciting prospect is not without challenges, especially as focus moves from interactions in vitro under tightly controlled conditions to tissue- and systems-level interactions in animal models and natural and experimental infections in humans. Here we review the contribution of transcriptomic studies to the understanding of malaria, a parasitic disease which has exerted a major influence on human evolution and continues to cause a huge global burden of disease. We consider malaria a paradigm for the transcriptomic assessment of systemic host-pathogen interactions in humans, because much of the direct host-pathogen interaction occurs within the blood, a readily sampled compartment of the body. We illustrate lessons learned from transcriptomic studies of malaria and how these lessons may guide studies of host-pathogen interactions in other infectious diseases. We propose that the potential of transcriptomic studies to improve the understanding of malaria as a disease remains partly untapped because of limitations in study design rather than as a consequence of technological constraints. Further advances will require the integration of transcriptomic data with analytical approaches from other scientific disciplines, including epidemiology and mathematical modeling. PMID:29695497

Transcriptomic Studies of Malaria: a Paradigm for Investigation of Systemic Host-Pathogen Interactions.

PubMed

Lee, Hyun Jae; Georgiadou, Athina; Otto, Thomas D; Levin, Michael; Coin, Lachlan J; Conway, David J; Cunnington, Aubrey J

2018-06-01

Transcriptomics, the analysis of genome-wide RNA expression, is a common approach to investigate host and pathogen processes in infectious diseases. Technical and bioinformatic advances have permitted increasingly thorough analyses of the association of RNA expression with fundamental biology, immunity, pathogenesis, diagnosis, and prognosis. Transcriptomic approaches can now be used to realize a previously unattainable goal, the simultaneous study of RNA expression in host and pathogen, in order to better understand their interactions. This exciting prospect is not without challenges, especially as focus moves from interactions in vitro under tightly controlled conditions to tissue- and systems-level interactions in animal models and natural and experimental infections in humans. Here we review the contribution of transcriptomic studies to the understanding of malaria, a parasitic disease which has exerted a major influence on human evolution and continues to cause a huge global burden of disease. We consider malaria a paradigm for the transcriptomic assessment of systemic host-pathogen interactions in humans, because much of the direct host-pathogen interaction occurs within the blood, a readily sampled compartment of the body. We illustrate lessons learned from transcriptomic studies of malaria and how these lessons may guide studies of host-pathogen interactions in other infectious diseases. We propose that the potential of transcriptomic studies to improve the understanding of malaria as a disease remains partly untapped because of limitations in study design rather than as a consequence of technological constraints. Further advances will require the integration of transcriptomic data with analytical approaches from other scientific disciplines, including epidemiology and mathematical modeling. Copyright © 2018 Lee et al.

Comparative transcriptome analysis of pepper (Capsicum annuum) revealed common regulons in multiple stress conditions and hormone treatments.

PubMed

Lee, Sanghyeob; Choi, Doil

2013-09-01

Global transcriptome analysis revealed common regulons for biotic/abiotic stresses, and some of these regulons encoding signaling components in both stresses were newly identified in this study. In this study, we aimed to identify plant responses to multiple stress conditions and discover the common regulons activated under a variety of stress conditions. Global transcriptome analysis revealed that salicylic acid (SA) may affect the activation of abiotic stress-responsive genes in pepper. Our data indicate that methyl jasmonate (MeJA) and ethylene (ET)-responsive genes were primarily activated by biotic stress, while abscisic acid (ABA)-responsive genes were activated under both types of stresses. We also identified differentially expressed gene (DEG) responses to specific stress conditions. Biotic stress induces more DEGs than those induced by abiotic and hormone applications. The clustering analysis using DEGs indicates that there are common regulons for biotic or abiotic stress conditions. Although SA and MeJA have an antagonistic effect on gene expression levels, SA and MeJA show a largely common regulation as compared to the regulation at the DEG expression level induced by other hormones. We also monitored the expression profiles of DEG encoding signaling components. Twenty-two percent of these were commonly expressed in both stress conditions. The importance of this study is that several genes commonly regulated by both stress conditions may have future applications for creating broadly stress-tolerant pepper plants. This study revealed that there are complex regulons in pepper plant to both biotic and abiotic stress conditions.

Microglia Transcriptome Changes in a Model of Depressive Behavior after Immune Challenge

PubMed Central

Gonzalez-Pena, Dianelys; Nixon, Scott E.; O’Connor, Jason C.; Southey, Bruce R.; Lawson, Marcus A.; McCusker, Robert H.; Borras, Tania; Machuca, Debbie; Hernandez, Alvaro G.; Dantzer, Robert; Kelley, Keith W.; Rodriguez-Zas, Sandra L.

2016-01-01

Depression symptoms following immune response to a challenge have been reported after the recovery from sickness. A RNA-Seq study of the dysregulation of the microglia transcriptome in a model of inflammation-associated depressive behavior was undertaken. The transcriptome of microglia from mice at day 7 after Bacille Calmette Guérin (BCG) challenge was compared to that from unchallenged Control mice and to the transcriptome from peripheral macrophages from the same mice. Among the 562 and 3,851 genes differentially expressed between BCG-challenged and Control mice in microglia and macrophages respectively, 353 genes overlapped between these cells types. Among the most differentially expressed genes in the microglia, serum amyloid A3 (Saa3) and cell adhesion molecule 3 (Cadm3) were over-expressed and coiled-coil domain containing 162 (Ccdc162) and titin-cap (Tcap) were under-expressed in BCG-challenged relative to Control. Many of the differentially expressed genes between BCG-challenged and Control mice were associated with neurological disorders encompassing depression symptoms. Across cell types, S100 calcium binding protein A9 (S100A9), interleukin 1 beta (Il1b) and kynurenine 3-monooxygenase (Kmo) were differentially expressed between challenged and control mice. Immune response, chemotaxis, and chemokine activity were among the functional categories enriched by the differentially expressed genes. Functional categories enriched among the 9,117 genes differentially expressed between cell types included leukocyte regulation and activation, chemokine and cytokine activities, MAP kinase activity, and apoptosis. More than 200 genes exhibited alternative splicing events between cell types including WNK lysine deficient protein kinase 1 (Wnk1) and microtubule-actin crosslinking factor 1(Macf1). Network visualization revealed the capability of microglia to exhibit transcriptome dysregulation in response to immune challenge still after resolution of sickness symptoms, albeit lower than that observed in macrophages. The persistent transcriptome dysregulation in the microglia shared patterns with neurological disorders indicating that the associated persistent depressive symptoms share a common transcriptome basis. PMID:26959683

Microglia Transcriptome Changes in a Model of Depressive Behavior after Immune Challenge.

PubMed

Gonzalez-Pena, Dianelys; Nixon, Scott E; O'Connor, Jason C; Southey, Bruce R; Lawson, Marcus A; McCusker, Robert H; Borras, Tania; Machuca, Debbie; Hernandez, Alvaro G; Dantzer, Robert; Kelley, Keith W; Rodriguez-Zas, Sandra L

2016-01-01

Depression symptoms following immune response to a challenge have been reported after the recovery from sickness. A RNA-Seq study of the dysregulation of the microglia transcriptome in a model of inflammation-associated depressive behavior was undertaken. The transcriptome of microglia from mice at day 7 after Bacille Calmette Guérin (BCG) challenge was compared to that from unchallenged Control mice and to the transcriptome from peripheral macrophages from the same mice. Among the 562 and 3,851 genes differentially expressed between BCG-challenged and Control mice in microglia and macrophages respectively, 353 genes overlapped between these cells types. Among the most differentially expressed genes in the microglia, serum amyloid A3 (Saa3) and cell adhesion molecule 3 (Cadm3) were over-expressed and coiled-coil domain containing 162 (Ccdc162) and titin-cap (Tcap) were under-expressed in BCG-challenged relative to Control. Many of the differentially expressed genes between BCG-challenged and Control mice were associated with neurological disorders encompassing depression symptoms. Across cell types, S100 calcium binding protein A9 (S100A9), interleukin 1 beta (Il1b) and kynurenine 3-monooxygenase (Kmo) were differentially expressed between challenged and control mice. Immune response, chemotaxis, and chemokine activity were among the functional categories enriched by the differentially expressed genes. Functional categories enriched among the 9,117 genes differentially expressed between cell types included leukocyte regulation and activation, chemokine and cytokine activities, MAP kinase activity, and apoptosis. More than 200 genes exhibited alternative splicing events between cell types including WNK lysine deficient protein kinase 1 (Wnk1) and microtubule-actin crosslinking factor 1(Macf1). Network visualization revealed the capability of microglia to exhibit transcriptome dysregulation in response to immune challenge still after resolution of sickness symptoms, albeit lower than that observed in macrophages. The persistent transcriptome dysregulation in the microglia shared patterns with neurological disorders indicating that the associated persistent depressive symptoms share a common transcriptome basis.

Transcriptome Assembly, Gene Annotation and Tissue Gene Expression Atlas of the Rainbow Trout

PubMed Central

Salem, Mohamed; Paneru, Bam; Al-Tobasei, Rafet; Abdouni, Fatima; Thorgaard, Gary H.; Rexroad, Caird E.; Yao, Jianbo

2015-01-01

Efforts to obtain a comprehensive genome sequence for rainbow trout are ongoing and will be complemented by transcriptome information that will enhance genome assembly and annotation. Previously, transcriptome reference sequences were reported using data from different sources. Although the previous work added a great wealth of sequences, a complete and well-annotated transcriptome is still needed. In addition, gene expression in different tissues was not completely addressed in the previous studies. In this study, non-normalized cDNA libraries were sequenced from 13 different tissues of a single doubled haploid rainbow trout from the same source used for the rainbow trout genome sequence. A total of ~1.167 billion paired-end reads were de novo assembled using the Trinity RNA-Seq assembler yielding 474,524 contigs > 500 base-pairs. Of them, 287,593 had homologies to the NCBI non-redundant protein database. The longest contig of each cluster was selected as a reference, yielding 44,990 representative contigs. A total of 4,146 contigs (9.2%), including 710 full-length sequences, did not match any mRNA sequences in the current rainbow trout genome reference. Mapping reads to the reference genome identified an additional 11,843 transcripts not annotated in the genome. A digital gene expression atlas revealed 7,678 housekeeping and 4,021 tissue-specific genes. Expression of about 16,000–32,000 genes (35–71% of the identified genes) accounted for basic and specialized functions of each tissue. White muscle and stomach had the least complex transcriptomes, with high percentages of their total mRNA contributed by a small number of genes. Brain, testis and intestine, in contrast, had complex transcriptomes, with a large numbers of genes involved in their expression patterns. This study provides comprehensive de novo transcriptome information that is suitable for functional and comparative genomics studies in rainbow trout, including annotation of the genome. PMID:25793877

Genetic signatures of adaptation revealed from transcriptome sequencing of Arctic and red foxes.

PubMed

Kumar, Vikas; Kutschera, Verena E; Nilsson, Maria A; Janke, Axel

2015-08-07

The genus Vulpes (true foxes) comprises numerous species that inhabit a wide range of habitats and climatic conditions, including one species, the Arctic fox (Vulpes lagopus) which is adapted to the arctic region. A close relative to the Arctic fox, the red fox (Vulpes vulpes), occurs in subarctic to subtropical habitats. To study the genetic basis of their adaptations to different environments, transcriptome sequences from two Arctic foxes and one red fox individual were generated and analyzed for signatures of positive selection. In addition, the data allowed for a phylogenetic analysis and divergence time estimate between the two fox species. The de novo assembly of reads resulted in more than 160,000 contigs/transcripts per individual. Approximately 17,000 homologous genes were identified using human and the non-redundant databases. Positive selection analyses revealed several genes involved in various metabolic and molecular processes such as energy metabolism, cardiac gene regulation, apoptosis and blood coagulation to be under positive selection in foxes. Branch site tests identified four genes to be under positive selection in the Arctic fox transcriptome, two of which are fat metabolism genes. In the red fox transcriptome eight genes are under positive selection, including molecular process genes, notably genes involved in ATP metabolism. Analysis of the three transcriptomes and five Sanger re-sequenced genes in additional individuals identified a lower genetic variability within Arctic foxes compared to red foxes, which is consistent with distribution range differences and demographic responses to past climatic fluctuations. A phylogenomic analysis estimated that the Arctic and red fox lineages diverged about three million years ago. Transcriptome data are an economic way to generate genomic resources for evolutionary studies. Despite not representing an entire genome, this transcriptome analysis identified numerous genes that are relevant to arctic adaptation in foxes. Similar to polar bears, fat metabolism seems to play a central role in adaptation of Arctic foxes to the cold climate, as has been identified in the polar bear, another arctic specialist.

Transcriptomic Changes Drive Physiological Responses to Progressive Drought Stress and Rehydration in Tomato

PubMed Central

Iovieno, Paolo; Punzo, Paola; Guida, Gianpiero; Mistretta, Carmela; Van Oosten, Michael J.; Nurcato, Roberta; Bostan, Hamed; Colantuono, Chiara; Costa, Antonello; Bagnaresi, Paolo; Chiusano, Maria L.; Albrizio, Rossella; Giorio, Pasquale; Batelli, Giorgia; Grillo, Stefania

2016-01-01

Tomato is a major crop in the Mediterranean basin, where the cultivation in the open field is often vulnerable to drought. In order to adapt and survive to naturally occurring cycles of drought stress and recovery, plants employ a coordinated array of physiological, biochemical, and molecular responses. Transcriptomic studies on tomato responses to drought and subsequent recovery are few in number. As the search for novel traits to improve the genetic tolerance to drought increases, a better understanding of these responses is required. To address this need we designed a study in which we induced two cycles of prolonged drought stress and a single recovery by rewatering in tomato. In order to dissect the complexity of plant responses to drought, we analyzed the physiological responses (stomatal conductance, CO2 assimilation, and chlorophyll fluorescence), abscisic acid (ABA), and proline contents. In addition to the physiological and metabolite assays, we generated transcriptomes for multiple points during the stress and recovery cycles. Cluster analysis of differentially expressed genes (DEGs) between the conditions has revealed potential novel components in stress response. The observed reduction in leaf gas exchanges and efficiency of the photosystem PSII was concomitant with a general down-regulation of genes belonging to the photosynthesis, light harvesting, and photosystem I and II category induced by drought stress. Gene ontology (GO) categories such as cell proliferation and cell cycle were also significantly enriched in the down-regulated fraction of genes upon drought stress, which may contribute to explain the observed growth reduction. Several histone variants were also repressed during drought stress, indicating that chromatin associated processes are also affected by drought. As expected, ABA accumulated after prolonged water deficit, driving the observed enrichment of stress related GOs in the up-regulated gene fractions, which included transcripts putatively involved in stomatal movements. This transcriptomic study has yielded promising candidate genes that merit further functional studies to confirm their involvement in drought tolerance and recovery. Together, our results contribute to a better understanding of the coordinated responses taking place under drought stress and recovery in adult plants of tomato. PMID:27066027

Acute Hepatopancreatic Necrosis Disease (AHPND) related microRNAs in Litopenaeus vannamei infected with AHPND-causing strain of Vibrio parahemolyticus.

PubMed

Zheng, Zhihong; Aweya, Jude Juventus; Wang, Fan; Yao, Defu; Lun, Jingsheng; Li, Shengkang; Ma, Hongyu; Zhang, Yueling

2018-05-08

Acute hepatopancreatic necrosis disease (AHPND) has emerged as a major debilitating disease that causes massive shrimp death resulting in substantial economic losses in shrimp aquaculture. Given that several diseases and infections have been associated with microRNAs (miRNAs), we conducted a comparative transcriptomic analysis using the AHPND (VA) and non-AHPND (VN) strains of Vibrio parahemolyticus to identify miRNAs potentially involved in AHPND pathogenesis in Litopenaeus vannamei. A total of 83 miRNAs (47 upregulated and 36 downregulated) were significantly differentially expressed between the VA and VN challenged groups, while 222 target genes of these miRNAs were predicted. Functional enrichment analysis revealed that the miRNAs target genes were involved in multiple biological processes including metabolic pathways, amoebiasis, Vibrio cholerae infection etc. Finally, interaction network and qPCR (Real-time Quantitative PCR) analysis of 12 potential key AHPND-related miRNAs and their predicted target genes, revealed their possible involvement in modulating several immune-related processes in the pathogenesis of AHPND. We have shown using comparative transcriptomic analysis, miRNAs and their target genes that are responsive to AHPND V. parahemolyticus infection in shrimp, therefore suggesting their possible role in defense response to AHPND V. parahemolyticus infection.

Molecular characterization of firefly nuptial gifts: a multi-omics approach sheds light on postcopulatory sexual selection.

PubMed

Al-Wathiqui, Nooria; Fallon, Timothy R; South, Adam; Weng, Jing-Ke; Lewis, Sara M

2016-12-22

Postcopulatory sexual selection is recognized as a key driver of reproductive trait evolution, including the machinery required to produce endogenous nuptial gifts. Despite the importance of such gifts, the molecular composition of the non-gametic components of male ejaculates and their interactions with female reproductive tracts remain poorly understood. During mating, male Photinus fireflies transfer to females a spermatophore gift manufactured by multiple reproductive glands. Here we combined transcriptomics of both male and female reproductive glands with proteomics and metabolomics to better understand the synthesis, composition and fate of the spermatophore in the common Eastern firefly, Photinus pyralis. Our transcriptome of male glands revealed up-regulation of proteases that may enhance male fertilization success and activate female immune response. Using bottom-up proteomics we identified 208 functionally annotated proteins that males transfer to the female in their spermatophore. Targeted metabolomic analysis also provided the first evidence that Photinus nuptial gifts contain lucibufagin, a firefly defensive toxin. The reproductive tracts of female fireflies showed increased gene expression for several proteases that may be involved in egg production. This study offers new insights into the molecular composition of male spermatophores, and extends our understanding of how nuptial gifts may mediate postcopulatory interactions between the sexes.

Applications of Single-Cell Sequencing for Multiomics.

PubMed

Xu, Yungang; Zhou, Xiaobo

2018-01-01

Single-cell sequencing interrogates the sequence or chromatin information from individual cells with advanced next-generation sequencing technologies. It provides a higher resolution of cellular differences and a better understanding of the underlying genetic and epigenetic mechanisms of an individual cell in the context of its survival and adaptation to microenvironment. However, it is more challenging to perform single-cell sequencing and downstream data analysis, owing to the minimal amount of starting materials, sample loss, and contamination. In addition, due to the picogram level of the amount of nucleic acids used, heavy amplification is often needed during sample preparation of single-cell sequencing, resulting in the uneven coverage, noise, and inaccurate quantification of sequencing data. All these unique properties raise challenges in and thus high demands for computational methods that specifically fit single-cell sequencing data. We here comprehensively survey the current strategies and challenges for multiple single-cell sequencing, including single-cell transcriptome, genome, and epigenome, beginning with a brief introduction to multiple sequencing techniques for single cells.

Global Transcriptome Analysis of Staphylococcus aureus Response to Hydrogen Peroxide†

PubMed Central

Chang, Wook; Small, David A.; Toghrol, Freshteh; Bentley, William E.

2006-01-01

Staphylococcus aureus responds with protective strategies against phagocyte-derived reactive oxidants to infect humans. Herein, we report the transcriptome analysis of the cellular response of S. aureus to hydrogen peroxide-induced oxidative stress. The data indicate that the oxidative response includes the induction of genes involved in virulence, DNA repair, and notably, anaerobic metabolism. PMID:16452450

The green ash transcriptome and identification of genes responding to abiotic and biotic stresses

Treesearch

Thomas Lane; Teodora Best; Nicole Zembower; Jack Davitt; Nathan Henry; Yi Xu; Jennifer Koch; Haiying Liang; John McGraw; Stephan Schuster; Donghwan Shim; Mark V. Coggeshall; John E. Carlson; Margaret E. Staton

2016-01-01

Background: To develop a set of transcriptome sequences to support research on environmental stress responses in green ash (Fraxinus pennsylvanica), we undertook deep RNA sequencing of green ash tissues under various stress treatments. The treatments, including emerald ash borer (EAB) feeding, heat, drought, cold and ozone, were selected to mimic...

Haematobia irritans dataset of raw sequence reads from Illumina-based transcriptome sequencing of specific tissues and life stages

USDA-ARS?s Scientific Manuscript database

Illumina HiSeq technology was used to sequence the transcriptome from various dissected tissues and life stages from the horn fly, Haematobia irritans. These samples include eggs (0, 2, 4, and 9 hours post-oviposition), adult fly gut, adult fly legs, adult fly malpighian tubule, adult fly ovary, adu...

Gleason Score 7 Prostate Cancers Emerge through Branched Evolution of Clonal Gleason Pattern 3 and 4.

PubMed

Sowalsky, Adam G; Kissick, Haydn T; Gerrin, Sean J; Schaefer, Rachel J; Xia, Zheng; Russo, Joshua W; Arredouani, M Simo; Bubley, Glenn J; Sanda, Martin G; Li, Wei; Ye, Huihui; Balk, Steven P

2017-07-15

Purpose: The molecular features that account for the distinct histology and aggressive biological behavior of Gleason pattern 4 (Gp4) versus Gp3 prostate cancer, and whether Gp3 tumors progress directly to Gp4, remain to be established. Experimental Design: Whole-exome sequencing and transcriptome profiling of laser capture-microdissected adjacent Gp3 and cribiform Gp4 were used to determine the relationship between these entities. Results: Sequencing confirmed that adjacent Gp3 and Gp4 were clonal based on multiple shared genomic alterations. However, large numbers of unique mutations in the Gp3 and Gp4 tumors showed that the Gp4 were not derived directly from the Gp3. Remarkably, the Gp3 tumors retain their indolent-appearing morphology despite acquisition of multiple genomic alterations, including tumor suppressor losses. Although there were no consistent genomic alterations that distinguished Gp3 from Gp4, pairwise transcriptome analyses identified increased c-Myc and decreased p53 activity in Gp4 versus adjacent clonal Gp3 foci. Conclusions: These findings establish that at least a subset of Gp3 and aggressive Gp4 tumors have a common origin, and support a branched evolution model wherein the Gp3 and Gp4 tumors emerge early from a common precursor and subsequently undergo substantial divergence. Genomic alterations detectable in the Gp3 may distinguish these tumors from truly indolent Gp3. Screening for a panel of these genomic alterations in men who have prostate biopsies showing only Gp3 (Gleason score 6, Gs6) may allow for more precise selection of men who can be safely managed by active surveillance versus those who may benefit from further intervention. Clin Cancer Res; 23(14); 3823-33. ©2017 AACR . ©2017 American Association for Cancer Research.

A systems biology approach to the analysis of subset-specific responses to lipopolysaccharide in dendritic cells.

PubMed

Hancock, David G; Shklovskaya, Elena; Guy, Thomas V; Falsafi, Reza; Fjell, Chris D; Ritchie, William; Hancock, Robert E W; Fazekas de St Groth, Barbara

2014-01-01

Dendritic cells (DCs) are critical for regulating CD4 and CD8 T cell immunity, controlling Th1, Th2, and Th17 commitment, generating inducible Tregs, and mediating tolerance. It is believed that distinct DC subsets have evolved to control these different immune outcomes. However, how DC subsets mount different responses to inflammatory and/or tolerogenic signals in order to accomplish their divergent functions remains unclear. Lipopolysaccharide (LPS) provides an excellent model for investigating responses in closely related splenic DC subsets, as all subsets express the LPS receptor TLR4 and respond to LPS in vitro. However, previous studies of the LPS-induced DC transcriptome have been performed only on mixed DC populations. Moreover, comparisons of the in vivo response of two closely related DC subsets to LPS stimulation have not been reported in the literature to date. We compared the transcriptomes of murine splenic CD8 and CD11b DC subsets after in vivo LPS stimulation, using RNA-Seq and systems biology approaches. We identified subset-specific gene signatures, which included multiple functional immune mediators unique to each subset. To explain the observed subset-specific differences, we used a network analysis approach. While both DC subsets used a conserved set of transcription factors and major signalling pathways, the subsets showed differential regulation of sets of genes that 'fine-tune' the network Hubs expressed in common. We propose a model in which signalling through common pathway components is 'fine-tuned' by transcriptional control of subset-specific modulators, thus allowing for distinct functional outcomes in closely related DC subsets. We extend this analysis to comparable datasets from the literature and confirm that our model can account for cell subset-specific responses to LPS stimulation in multiple subpopulations in mouse and man.

Blood transcriptomic diagnosis of pulmonary and extrapulmonary tuberculosis

PubMed Central

Roe, Jennifer K; Thomas, Niclas; Gil, Eliza; Best, Katharine; Tsaliki, Evdokia; Morris‑Jones, Stephen; Stafford, Sian; Simpson, Nandi; Witt, Karolina D; Chain, Benjamin; Miller, Robert F; Martineau, Adrian

2016-01-01

BACKGROUND. Novel rapid diagnostics for active tuberculosis (TB) are required to overcome the time delays and inadequate sensitivity of current microbiological tests that are critically dependent on sampling the site of disease. Multiparametric blood transcriptomic signatures of TB have been described as potential diagnostic tests. We sought to identify the best transcript candidates as host biomarkers for active TB, extend the evaluation of their specificity by comparison with other infectious diseases, and to test their performance in both pulmonary and extrapulmonary TB. METHODS. Support vector machine learning, combined with feature selection, was applied to new and previously published blood transcriptional profiles in order to identify the minimal TB‑specific transcriptional signature shared by multiple patient cohorts including pulmonary and extrapulmonary TB, and individuals with and without HIV-1 coinfection. RESULTS. We identified and validated elevated blood basic leucine zipper transcription factor 2 (BATF2) transcript levels as a single sensitive biomarker that discriminated active pulmonary and extrapulmonary TB from healthy individuals, with receiver operating characteristic (ROC) area under the curve (AUC) scores of 0.93 to 0.99 in multiple cohorts of HIV-1–negative individuals, and 0.85 in HIV-1–infected individuals. In addition, we identified and validated a potentially novel 4-gene signature comprising CD177, haptoglobin, immunoglobin J chain, and galectin 10 that discriminated active pulmonary and extrapulmonary TB from other febrile infections, giving ROC AUCs of 0.94 to 1. CONCLUSIONS. Elevated blood BATF2 transcript levels provide a sensitive biomarker that discriminates active TB from healthy individuals, and a potentially novel 4-gene transcriptional signature differentiates between active TB and other infectious diseases in individuals presenting with fever. FUNDING. MRC, Wellcome Trust, Rosetrees Trust, British Lung Foundation, NIHR. PMID:27734027

Blood transcriptomic diagnosis of pulmonary and extrapulmonary tuberculosis.

PubMed

Roe, Jennifer K; Thomas, Niclas; Gil, Eliza; Best, Katharine; Tsaliki, Evdokia; Morris-Jones, Stephen; Stafford, Sian; Simpson, Nandi; Witt, Karolina D; Chain, Benjamin; Miller, Robert F; Martineau, Adrian; Noursadeghi, Mahdad

2016-10-06

BACKGROUND. Novel rapid diagnostics for active tuberculosis (TB) are required to overcome the time delays and inadequate sensitivity of current microbiological tests that are critically dependent on sampling the site of disease. Multiparametric blood transcriptomic signatures of TB have been described as potential diagnostic tests. We sought to identify the best transcript candidates as host biomarkers for active TB, extend the evaluation of their specificity by comparison with other infectious diseases, and to test their performance in both pulmonary and extrapulmonary TB. METHODS. Support vector machine learning, combined with feature selection, was applied to new and previously published blood transcriptional profiles in order to identify the minimal TB‑specific transcriptional signature shared by multiple patient cohorts including pulmonary and extrapulmonary TB, and individuals with and without HIV-1 coinfection. RESULTS. We identified and validated elevated blood basic leucine zipper transcription factor 2 ( BATF2 ) transcript levels as a single sensitive biomarker that discriminated active pulmonary and extrapulmonary TB from healthy individuals, with receiver operating characteristic (ROC) area under the curve (AUC) scores of 0.93 to 0.99 in multiple cohorts of HIV-1-negative individuals, and 0.85 in HIV-1-infected individuals. In addition, we identified and validated a potentially novel 4-gene signature comprising CD177, haptoglobin, immunoglobin J chain, and galectin 10 that discriminated active pulmonary and extrapulmonary TB from other febrile infections, giving ROC AUCs of 0.94 to 1. CONCLUSIONS. Elevated blood BATF2 transcript levels provide a sensitive biomarker that discriminates active TB from healthy individuals, and a potentially novel 4-gene transcriptional signature differentiates between active TB and other infectious diseases in individuals presenting with fever. FUNDING. MRC, Wellcome Trust, Rosetrees Trust, British Lung Foundation, NIHR.

De novo Transcriptome Assembly of Common Wild Rice (Oryza rufipogon Griff.) and Discovery of Drought-Response Genes in Root Tissue Based on Transcriptomic Data.

PubMed

Tian, Xin-Jie; Long, Yan; Wang, Jiao; Zhang, Jing-Wen; Wang, Yan-Yan; Li, Wei-Min; Peng, Yu-Fa; Yuan, Qian-Hua; Pei, Xin-Wu

2015-01-01

The perennial O. rufipogon (common wild rice), which is considered to be the ancestor of Asian cultivated rice species, contains many useful genetic resources, including drought resistance genes. However, few studies have identified the drought resistance and tissue-specific genes in common wild rice. In this study, transcriptome sequencing libraries were constructed, including drought-treated roots (DR) and control leaves (CL) and roots (CR). Using Illumina sequencing technology, we generated 16.75 million bases of high-quality sequence data for common wild rice and conducted de novo assembly and annotation of genes without prior genome information. These reads were assembled into 119,332 unigenes with an average length of 715 bp. A total of 88,813 distinct sequences (74.42% of unigenes) significantly matched known genes in the NCBI NT database. Differentially expressed gene (DEG) analysis showed that 3617 genes were up-regulated and 4171 genes were down-regulated in the CR library compared with the CL library. Among the DEGs, 535 genes were expressed in roots but not in shoots. A similar comparison between the DR and CR libraries showed that 1393 genes were up-regulated and 315 genes were down-regulated in the DR library compared with the CR library. Finally, 37 genes that were specifically expressed in roots were screened after comparing the DEGs identified in the above-described analyses. This study provides a transcriptome sequence resource for common wild rice plants and establishes a digital gene expression profile of wild rice plants under drought conditions using the assembled transcriptome data as a reference. Several tissue-specific and drought-stress-related candidate genes were identified, representing a fully characterized transcriptome and providing a valuable resource for genetic and genomic studies in plants.

ReprOlive: a database with linked data for the olive tree (Olea europaea L.) reproductive transcriptome

PubMed Central

Carmona, Rosario; Zafra, Adoración; Seoane, Pedro; Castro, Antonio J.; Guerrero-Fernández, Darío; Castillo-Castillo, Trinidad; Medina-García, Ana; Cánovas, Francisco M.; Aldana-Montes, José F.; Navas-Delgado, Ismael; Alché, Juan de Dios; Claros, M. Gonzalo

2015-01-01

Plant reproductive transcriptomes have been analyzed in different species due to the agronomical and biotechnological importance of plant reproduction. Here we presented an olive tree reproductive transcriptome database with samples from pollen and pistil at different developmental stages, and leaf and root as control vegetative tissues http://reprolive.eez.csic.es). It was developed from 2,077,309 raw reads to 1,549 Sanger sequences. Using a pre-defined workflow based on open-source tools, sequences were pre-processed, assembled, mapped, and annotated with expression data, descriptions, GO terms, InterPro signatures, EC numbers, KEGG pathways, ORFs, and SSRs. Tentative transcripts (TTs) were also annotated with the corresponding orthologs in Arabidopsis thaliana from TAIR and RefSeq databases to enable Linked Data integration. It results in a reproductive transcriptome comprising 72,846 contigs with average length of 686 bp, of which 63,965 (87.8%) included at least one functional annotation, and 55,356 (75.9%) had an ortholog. A minimum of 23,568 different TTs was identified and 5,835 of them contain a complete ORF. The representative reproductive transcriptome can be reduced to 28,972 TTs for further gene expression studies. Partial transcriptomes from pollen, pistil, and vegetative tissues as control were also constructed. ReprOlive provides free access and download capability to these results. Retrieval mechanisms for sequences and transcript annotations are provided. Graphical localization of annotated enzymes into KEGG pathways is also possible. Finally, ReprOlive has included a semantic conceptualisation by means of a Resource Description Framework (RDF) allowing a Linked Data search for extracting the most updated information related to enzymes, interactions, allergens, structures, and reactive oxygen species. PMID:26322066

Transcriptomics reveals multiple resistance mechanisms against cotton leaf curl disease in a naturally immune cotton species, Gossypium arboreum

USDA-ARS?s Scientific Manuscript database

Cotton is an economically important crop affected by a number of abiotic and biotic stresses. Cotton leaf curl disease (CLCuD) is caused by virus in the genus Begomovirus (family Geminiviridae), collectively called cotton leaf curl viruses (CLCuVs). It is one of the most devastating virual diseases ...

Conifer DBMagic: A database housing multiple de novo transcriptome assemblies for twelve diverse conifer species

Treesearch

W. Walter Lorenz; Savavanaraj Ayyampalayam; John M. Bordeaux; Glenn T. Howe; Kathleen D. Jermstad; David B. Neale; Deborah L. Rogers; Jeffrey F.D. Dean

2012-01-01

Conifers comprise an ancient and widespread plant lineage of enormous commercial and ecological value. However, compared to model woody angiosperms, such as Populus and Eucalyptus, our understanding of conifers remains quite limited at a genomic level. Large genome sizes (10,000-40,000 Mbp) and large amounts of repetitive DNA...

Integrated Molecular Characterization of Uterine Carcinosarcoma.

PubMed

Cherniack, Andrew D; Shen, Hui; Walter, Vonn; Stewart, Chip; Murray, Bradley A; Bowlby, Reanne; Hu, Xin; Ling, Shiyun; Soslow, Robert A; Broaddus, Russell R; Zuna, Rosemary E; Robertson, Gordon; Laird, Peter W; Kucherlapati, Raju; Mills, Gordon B; Weinstein, John N; Zhang, Jiashan; Akbani, Rehan; Levine, Douglas A

2017-03-13

We performed genomic, epigenomic, transcriptomic, and proteomic characterizations of uterine carcinosarcomas (UCSs). Cohort samples had extensive copy-number alterations and highly recurrent somatic mutations. Frequent mutations were found in TP53, PTEN, PIK3CA, PPP2R1A, FBXW7, and KRAS, similar to endometrioid and serous uterine carcinomas. Transcriptome sequencing identified a strong epithelial-to-mesenchymal transition (EMT) gene signature in a subset of cases that was attributable to epigenetic alterations at microRNA promoters. The range of EMT scores in UCS was the largest among all tumor types studied via The Cancer Genome Atlas. UCSs shared proteomic features with gynecologic carcinomas and sarcomas with intermediate EMT features. Multiple somatic mutations and copy-number alterations in genes that are therapeutic targets were identified. Copyright © 2017 Elsevier Inc. All rights reserved.

Single-cell analysis of the transcriptome and its application in the characterization of stem cells and early embryos.

PubMed

Liu, Na; Liu, Lin; Pan, Xinghua

2014-07-01

Cellular heterogeneity within a cell population is a common phenomenon in multicellular organisms, tissues, cultured cells, and even FACS-sorted subpopulations. Important information may be masked if the cells are studied as a mass. Transcriptome profiling is a parameter that has been intensively studied, and relatively easier to address than protein composition. To understand the basis and importance of heterogeneity and stochastic aspects of the cell function and its mechanisms, it is essential to examine transcriptomes of a panel of single cells. High-throughput technologies, starting from microarrays and now RNA-seq, provide a full view of the expression of transcriptomes but are limited by the amount of RNA for analysis. Recently, several new approaches for amplification and sequencing the transcriptome of single cells or a limited low number of cells have been developed and applied. In this review, we summarize these major strategies, such as PCR-based methods, IVT-based methods, phi29-DNA polymerase-based methods, and several other methods, including their principles, characteristics, advantages, and limitations, with representative applications in cancer stem cells, early development, and embryonic stem cells. The prospects for development of future technology and application of transcriptome analysis in a single cell are also discussed.

Insecticide resistance is mediated by multiple mechanisms in recently introduced Aedes aegypti from Madeira Island (Portugal).

PubMed

Seixas, Gonçalo; Grigoraki, Linda; Weetman, David; Vicente, José Luís; Silva, Ana Clara; Pinto, João; Vontas, John; Sousa, Carla Alexandra

2017-07-01

Aedes aegypti is a major mosquito vector of arboviruses, including dengue, chikungunya and Zika. In 2005, Ae. aegypti was identified for the first time in Madeira Island. Despite an initial insecticide-based vector control program, the species expanded throughout the Southern coast of the island, suggesting the presence of insecticide resistance. Here, we characterized the insecticide resistance status and the underlying mechanisms of two populations of Ae. aegypti from Madeira Island, Funchal and Paúl do Mar. WHO susceptibility bioassays indicated resistance to cyfluthrin, permethrin, fenitrothion and bendiocarb. Use of synergists significantly increased mortality rates, and biochemical assays indicated elevated activities of detoxification enzymes, suggesting the importance of metabolic resistance. Microarray-based transcriptome analysis detected significant upregulation in both populations of nine cytochrome P450 oxidase genes (including four known pyrethroid metabolizing enzymes), the organophosphate metabolizer CCEae3a, Glutathione-S-transferases, and multiple putative cuticle proteins. Genotyping of knockdown resistance loci linked to pyrethroid resistance revealed fixation of the 1534C mutation, and presence with moderate frequencies of the V1016I mutation in each population. Significant resistance to three major insecticide classes (pyrethroid, carbamate and organophosphate) is present in Ae. aegypti from Madeira Island, and appears to be mediated by multiple mechanisms. Implementation of appropriate resistance management strategies including rotation of insecticides with alternative modes of action, and methods other than chemical-based vector control are strongly advised to delay or reverse the spread of resistance and achieve efficient control.

ATGC transcriptomics: a web-based application to integrate, explore and analyze de novo transcriptomic data.

PubMed

Gonzalez, Sergio; Clavijo, Bernardo; Rivarola, Máximo; Moreno, Patricio; Fernandez, Paula; Dopazo, Joaquín; Paniego, Norma

2017-02-22

In the last years, applications based on massively parallelized RNA sequencing (RNA-seq) have become valuable approaches for studying non-model species, e.g., without a fully sequenced genome. RNA-seq is a useful tool for detecting novel transcripts and genetic variations and for evaluating differential gene expression by digital measurements. The large and complex datasets resulting from functional genomic experiments represent a challenge in data processing, management, and analysis. This problem is especially significant for small research groups working with non-model species. We developed a web-based application, called ATGC transcriptomics, with a flexible and adaptable interface that allows users to work with new generation sequencing (NGS) transcriptomic analysis results using an ontology-driven database. This new application simplifies data exploration, visualization, and integration for a better comprehension of the results. ATGC transcriptomics provides access to non-expert computer users and small research groups to a scalable storage option and simple data integration, including database administration and management. The software is freely available under the terms of GNU public license at http://atgcinta.sourceforge.net .

Transcriptomic responses to wounding: meta-analysis of gene expression microarray data.

PubMed

Sass, Piotr Andrzej; Dąbrowski, Michał; Charzyńska, Agata; Sachadyn, Paweł

2017-11-07

A vast amount of microarray data on transcriptomic response to injury has been collected so far. We designed the analysis in order to identify the genes displaying significant changes in expression after wounding in different organisms and tissues. This meta-analysis is the first study to compare gene expression profiles in response to wounding in as different tissues as heart, liver, skin, bones, and spinal cord, and species, including rat, mouse and human. We collected available microarray transcriptomic profiles obtained from different tissue injury experiments and selected the genes showing a minimum twofold change in expression in response to wounding in prevailing number of experiments for each of five wound healing stages we distinguished: haemostasis & early inflammation, inflammation, early repair, late repair and remodelling. During the initial phases after wounding, haemostasis & early inflammation and inflammation, the transcriptomic responses showed little consistency between different tissues and experiments. For the later phases, wound repair and remodelling, we identified a number of genes displaying similar transcriptional responses in all examined tissues. As revealed by ontological analyses, activation of certain pathways was rather specific for selected phases of wound healing, such as e.g. responses to vitamin D pronounced during inflammation. Conversely, we observed induction of genes encoding inflammatory agents and extracellular matrix proteins in all wound healing phases. Further, we selected several genes differentially upregulated throughout different stages of wound response, including established factors of wound healing in addition to those previously unreported  in this context such as PTPRC and AQP4. We found that transcriptomic responses to wounding showed similar traits in a diverse selection of tissues including skin, muscles, internal organs and nervous system. Notably, we distinguished transcriptional induction of inflammatory genes not only in the initial response to wounding, but also later, during wound repair and tissue remodelling.

Analysis of the Salivary Gland Transcriptome of Frankliniella occidentalis

PubMed Central

Stafford-Banks, Candice A.; Rotenberg, Dorith; Johnson, Brian R.; Whitfield, Anna E.; Ullman, Diane E.

2014-01-01

Saliva is known to play a crucial role in insect feeding behavior and virus transmission. Currently, little is known about the salivary glands and saliva of thrips, despite the fact that Frankliniella occidentalis (Pergande) (the western flower thrips) is a serious pest due to its destructive feeding, wide host range, and transmission of tospoviruses. As a first step towards characterizing thrips salivary gland functions, we sequenced the transcriptome of the primary salivary glands of F. occidentalis using short read sequencing (Illumina) technology. A de novo-assembled transcriptome revealed 31,392 high quality contigs with an average size of 605 bp. A total of 12,166 contigs had significant BLASTx or tBLASTx hits (E≤1.0E−6) to known proteins, whereas a high percentage (61.24%) of contigs had no apparent protein or nucleotide hits. Comparison of the F. occidentalis salivary gland transcriptome (sialotranscriptome) against a published F. occidentalis full body transcriptome assembled from Roche-454 reads revealed several contigs with putative annotations associated with salivary gland functions. KEGG pathway analysis of the sialotranscriptome revealed that the majority (18 out of the top 20 predicted KEGG pathways) of the salivary gland contig sequences match proteins involved in metabolism. We identified several genes likely to be involved in detoxification and inhibition of plant defense responses including aldehyde dehydrogenase, metalloprotease, glucose oxidase, glucose dehydrogenase, and regucalcin. We also identified several genes that may play a role in the extra-oral digestion of plant structural tissues including β-glucosidase and pectin lyase; and the extra-oral digestion of sugars, including α-amylase, maltase, sucrase, and α-glucosidase. This is the first analysis of a sialotranscriptome for any Thysanopteran species and it provides a foundational tool to further our understanding of how thrips interact with their plant hosts and the viruses they transmit. PMID:24736614

Analysis of the salivary gland transcriptome of Frankliniella occidentalis.

PubMed

Stafford-Banks, Candice A; Rotenberg, Dorith; Johnson, Brian R; Whitfield, Anna E; Ullman, Diane E

2014-01-01

Saliva is known to play a crucial role in insect feeding behavior and virus transmission. Currently, little is known about the salivary glands and saliva of thrips, despite the fact that Frankliniella occidentalis (Pergande) (the western flower thrips) is a serious pest due to its destructive feeding, wide host range, and transmission of tospoviruses. As a first step towards characterizing thrips salivary gland functions, we sequenced the transcriptome of the primary salivary glands of F. occidentalis using short read sequencing (Illumina) technology. A de novo-assembled transcriptome revealed 31,392 high quality contigs with an average size of 605 bp. A total of 12,166 contigs had significant BLASTx or tBLASTx hits (E≤1.0E-6) to known proteins, whereas a high percentage (61.24%) of contigs had no apparent protein or nucleotide hits. Comparison of the F. occidentalis salivary gland transcriptome (sialotranscriptome) against a published F. occidentalis full body transcriptome assembled from Roche-454 reads revealed several contigs with putative annotations associated with salivary gland functions. KEGG pathway analysis of the sialotranscriptome revealed that the majority (18 out of the top 20 predicted KEGG pathways) of the salivary gland contig sequences match proteins involved in metabolism. We identified several genes likely to be involved in detoxification and inhibition of plant defense responses including aldehyde dehydrogenase, metalloprotease, glucose oxidase, glucose dehydrogenase, and regucalcin. We also identified several genes that may play a role in the extra-oral digestion of plant structural tissues including β-glucosidase and pectin lyase; and the extra-oral digestion of sugars, including α-amylase, maltase, sucrase, and α-glucosidase. This is the first analysis of a sialotranscriptome for any Thysanopteran species and it provides a foundational tool to further our understanding of how thrips interact with their plant hosts and the viruses they transmit.

Transcriptome analysis of functional differentiation between haploid and diploid cells of Emiliania huxleyi, a globally significant photosynthetic calcifying cell.

PubMed

von Dassow, Peter; Ogata, Hiroyuki; Probert, Ian; Wincker, Patrick; Da Silva, Corinne; Audic, Stéphane; Claverie, Jean-Michel; de Vargas, Colomban

2009-01-01

Eukaryotes are classified as either haplontic, diplontic, or haplo-diplontic, depending on which ploidy levels undergo mitotic cell division in the life cycle. Emiliania huxleyi is one of the most abundant phytoplankton species in the ocean, playing an important role in global carbon fluxes, and represents haptophytes, an enigmatic group of unicellular organisms that diverged early in eukaryotic evolution. This species is haplo-diplontic. Little is known about the haploid cells, but they have been hypothesized to allow persistence of the species between the yearly blooms of diploid cells. We sequenced over 38,000 expressed sequence tags from haploid and diploid E. huxleyi normalized cDNA libraries to identify genes involved in important processes specific to each life phase (2N calcification or 1N motility), and to better understand the haploid phase of this prominent haplo-diplontic organism. The haploid and diploid transcriptomes showed a dramatic differentiation, with approximately 20% greater transcriptome richness in diploid cells than in haploid cells and only

Transcriptome analysis of functional differentiation between haploid and diploid cells of Emiliania huxleyi, a globally significant photosynthetic calcifying cell

PubMed Central

2009-01-01

Background Eukaryotes are classified as either haplontic, diplontic, or haplo-diplontic, depending on which ploidy levels undergo mitotic cell division in the life cycle. Emiliania huxleyi is one of the most abundant phytoplankton species in the ocean, playing an important role in global carbon fluxes, and represents haptophytes, an enigmatic group of unicellular organisms that diverged early in eukaryotic evolution. This species is haplo-diplontic. Little is known about the haploid cells, but they have been hypothesized to allow persistence of the species between the yearly blooms of diploid cells. We sequenced over 38,000 expressed sequence tags from haploid and diploid E. huxleyi normalized cDNA libraries to identify genes involved in important processes specific to each life phase (2N calcification or 1N motility), and to better understand the haploid phase of this prominent haplo-diplontic organism. Results The haploid and diploid transcriptomes showed a dramatic differentiation, with approximately 20% greater transcriptome richness in diploid cells than in haploid cells and only ≤ 50% of transcripts estimated to be common between the two phases. The major functional category of transcripts differentiating haploids included signal transduction and motility genes. Diploid-specific transcripts included Ca2+, H+, and HCO3- pumps. Potential factors differentiating the transcriptomes included haploid-specific Myb transcription factor homologs and an unusual diploid-specific histone H4 homolog. Conclusions This study permitted the identification of genes likely involved in diploid-specific biomineralization, haploid-specific motility, and transcriptional control. Greater transcriptome richness in diploid cells suggests they may be more versatile for exploiting a diversity of rich environments whereas haploid cells are intrinsically more streamlined. PMID:19832986

The maize (Zea mays ssp. mays var. B73) genome encodes 33 members of the purple acid phosphatase family

PubMed Central

González-Muñoz, Eliécer; Avendaño-Vázquez, Aida-Odette; Montes, Ricardo A. Chávez; de Folter, Stefan; Andrés-Hernández, Liliana; Abreu-Goodger, Cei; Sawers, Ruairidh J. H.

2015-01-01

Purple acid phosphatases (PAPs) play an important role in plant phosphorus nutrition, both by liberating phosphorus from organic sources in the soil and by modulating distribution within the plant throughout growth and development. Furthermore, members of the PAP protein family have been implicated in a broader role in plant mineral homeostasis, stress responses and development. We have identified 33 candidate PAP encoding gene models in the maize (Zea mays ssp. mays var. B73) reference genome. The maize Pap family includes a clear single-copy ortholog of the Arabidopsis gene AtPAP26, shown previously to encode both major intracellular and secreted acid phosphatase activities. Certain groups of PAPs present in Arabidopsis, however, are absent in maize, while the maize family contains a number of expansions, including a distinct radiation not present in Arabidopsis. Analysis of RNA-sequencing based transcriptome data revealed accumulation of maize Pap transcripts in multiple plant tissues at multiple stages of development, and increased accumulation of specific transcripts under low phosphorus availability. These data suggest the maize PAP family as a whole to have broad significance throughout the plant life cycle, while highlighting potential functional specialization of individual family members. PMID:26042133

Whole Transcriptome Sequencing Enables Discovery and Analysis of Viruses in Archived Primary Central Nervous System Lymphomas

PubMed Central

DeBoever, Christopher; Reid, Erin G.; Smith, Erin N.; Wang, Xiaoyun; Dumaop, Wilmar; Harismendy, Olivier; Carson, Dennis; Richman, Douglas; Masliah, Eliezer; Frazer, Kelly A.

2013-01-01

Primary central nervous system lymphomas (PCNSL) have a dramatically increased prevalence among persons living with AIDS and are known to be associated with human Epstein Barr virus (EBV) infection. Previous work suggests that in some cases, co-infection with other viruses may be important for PCNSL pathogenesis. Viral transcription in tumor samples can be measured using next generation transcriptome sequencing. We demonstrate the ability of transcriptome sequencing to identify viruses, characterize viral expression, and identify viral variants by sequencing four archived AIDS-related PCNSL tissue samples and analyzing raw sequencing reads. EBV was detected in all four PCNSL samples and cytomegalovirus (CMV), JC polyomavirus (JCV), and HIV were also discovered, consistent with clinical diagnoses. CMV was found to express three long non-coding RNAs recently reported as expressed during active infection. Single nucleotide variants were observed in each of the viruses observed and three indels were found in CMV. No viruses were found in several control tumor types including 32 diffuse large B-cell lymphoma samples. This study demonstrates the ability of next generation transcriptome sequencing to accurately identify viruses, including DNA viruses, in solid human cancer tissue samples. PMID:24023918

Large-scale atlas of microarray data reveals the distinct expression landscape of different tissues in Arabidopsis

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

He, Fei; Maslov, Sergei; Yoo, Shinjae

Here, transcriptome datasets from thousands of samples of the model plant Arabidopsis thaliana have been collectively generated by multiple individual labs. Although integration and meta-analysis of these samples has become routine in the plant research community, it is often hampered by the lack of metadata or differences in annotation styles by different labs. In this study, we carefully selected and integrated 6,057 Arabidopsis microarray expression samples from 304 experiments deposited to NCBI GEO. Metadata such as tissue type, growth condition, and developmental stage were manually curated for each sample. We then studied global expression landscape of the integrated dataset andmore » found that samples of the same tissue tend to be more similar to each other than to samples of other tissues, even in different growth conditions or developmental stages. Root has the most distinct transcriptome compared to aerial tissues, but the transcriptome of cultured root is more similar to those of aerial tissues as the former samples lost their cellular identity. Using a simple computational classification method, we showed that the tissue type of a sample can be successfully predicted based on its expression profile, opening the door for automatic metadata extraction and facilitating re-use of plant transcriptome data. As a proof of principle we applied our automated annotation pipeline to 708 RNA-seq samples from public repositories and verified accuracy of our predictions with samples’ metadata provided by authors.« less

Large-scale atlas of microarray data reveals the distinct expression landscape of different tissues in Arabidopsis

DOE PAGES

He, Fei; Maslov, Sergei; Yoo, Shinjae; ...

2016-05-25

Here, transcriptome datasets from thousands of samples of the model plant Arabidopsis thaliana have been collectively generated by multiple individual labs. Although integration and meta-analysis of these samples has become routine in the plant research community, it is often hampered by the lack of metadata or differences in annotation styles by different labs. In this study, we carefully selected and integrated 6,057 Arabidopsis microarray expression samples from 304 experiments deposited to NCBI GEO. Metadata such as tissue type, growth condition, and developmental stage were manually curated for each sample. We then studied global expression landscape of the integrated dataset andmore » found that samples of the same tissue tend to be more similar to each other than to samples of other tissues, even in different growth conditions or developmental stages. Root has the most distinct transcriptome compared to aerial tissues, but the transcriptome of cultured root is more similar to those of aerial tissues as the former samples lost their cellular identity. Using a simple computational classification method, we showed that the tissue type of a sample can be successfully predicted based on its expression profile, opening the door for automatic metadata extraction and facilitating re-use of plant transcriptome data. As a proof of principle we applied our automated annotation pipeline to 708 RNA-seq samples from public repositories and verified accuracy of our predictions with samples’ metadata provided by authors.« less

Gene expression analysis of induced pluripotent stem cells from aneuploid chromosomal syndromes

PubMed Central

2013-01-01

Background Human aneuploidy is the leading cause of early pregnancy loss, mental retardation, and multiple congenital anomalies. Due to the high mortality associated with aneuploidy, the pathophysiological mechanisms of aneuploidy syndrome remain largely unknown. Previous studies focused mostly on whether dosage compensation occurs, and the next generation transcriptomics sequencing technology RNA-seq is expected to eventually uncover the mechanisms of gene expression regulation and the related pathological phenotypes in human aneuploidy. Results Using next generation transcriptomics sequencing technology RNA-seq, we profiled the transcriptomes of four human aneuploid induced pluripotent stem cell (iPSC) lines generated from monosomy × (Turner syndrome), trisomy 8 (Warkany syndrome 2), trisomy 13 (Patau syndrome), and partial trisomy 11:22 (Emanuel syndrome) as well as two umbilical cord matrix iPSC lines as euploid controls to examine how phenotypic abnormalities develop with aberrant karyotype. A total of 466 M (50-bp) reads were obtained from the six iPSC lines, and over 13,000 mRNAs were identified by gene annotation. Global analysis of gene expression profiles and functional analysis of differentially expressed (DE) genes were implemented. Over 5000 DE genes are determined between aneuploidy and euploid iPSCs respectively while 9 KEGG pathways are overlapped enriched in four aneuploidy samples. Conclusions Our results demonstrate that the extra or missing chromosome has extensive effects on the whole transcriptome. Functional analysis of differentially expressed genes reveals that the genes most affected in aneuploid individuals are related to central nervous system development and tumorigenesis. PMID:24564826

De novo Assembly and Analysis of the Chilean Pencil Catfish Trichomycterus areolatus Transcriptome

PubMed Central

Schulze, Thomas T.; Ali, Jonathan M.; Bartlett, Maggie L.; McFarland, Madalyn M.; Clement, Emalie J.; Won, Harim I.; Sanford, Austin G.; Monzingo, Elyssa B.; Martens, Matthew C.; Hemsley, Ryan M.; Kumar, Sidharta; Gouin, Nicolas; Kolok, Alan S.; Davis, Paul H.

2016-01-01

Trichomycterus areolatus is an endemic species of pencil catfish that inhabits the riffles and rapids of many freshwater ecosystems of Chile. Despite its unique adaptation to Chile's high gradient watersheds and therefore potential application in the investigation of ecosystem integrity and environmental contamination, relatively little is known regarding the molecular biology of this environmental sentinel. Here, we detail the assembly of the Trichomycterus areolatus transcriptome, a molecular resource for the study of this organism and its molecular response to the environment. RNA-Seq reads were obtained by next-generation sequencing with an Illumina® platform and processed using PRINSEQ. The transcriptome assembly was performed using TRINITY assembler. Transcriptome validation was performed by functional characterization with KOG, KEGG, and GO analyses. Additionally, differential expression analysis highlights sex-specific expression patterns, and a list of endocrine and oxidative stress related transcripts are included. PMID:27672404

A large-scale full-length cDNA analysis to explore the budding yeast transcriptome

PubMed Central

Miura, Fumihito; Kawaguchi, Noriko; Sese, Jun; Toyoda, Atsushi; Hattori, Masahira; Morishita, Shinichi; Ito, Takashi

2006-01-01

We performed a large-scale cDNA analysis to explore the transcriptome of the budding yeast Saccharomyces cerevisiae. We sequenced two cDNA libraries, one from the cells exponentially growing in a minimal medium and the other from meiotic cells. Both libraries were generated by using a vector-capping method that allows the accurate mapping of transcription start sites (TSSs). Consequently, we identified 11,575 TSSs associated with 3,638 annotated genomic features, including 3,599 ORFs, to suggest that most yeast genes have two or more TSSs. In addition, we identified 45 previously undescribed introns, including those affecting current ORF annotations and those spliced alternatively. Furthermore, the analysis revealed 667 transcription units in the intergenic regions and transcripts derived from antisense strands of 367 known features. We also found that 348 ORFs carry TSSs in their 3′-halves to generate sense transcripts starting from inside the ORFs. These results indicate that the budding yeast transcriptome is considerably more complex than previously thought, and it shares many recently revealed characteristics with the transcriptomes of mammals and other higher eukaryotes. Thus, the genome-wide active transcription that generates novel classes of transcripts appears to be an intrinsic feature of the eukaryotic cells. The budding yeast will serve as a versatile model for the studies on these aspects of transcriptome, and the full-length cDNA clones can function as an invaluable resource in such studies. PMID:17101987

Isoform Sequencing Provides a More Comprehensive View of the Panax ginseng Transcriptome.

PubMed

Jo, Ick-Hyun; Lee, Jinsu; Hong, Chi Eun; Lee, Dong Jin; Bae, Wonsil; Park, Sin-Gi; Ahn, Yong Ju; Kim, Young Chang; Kim, Jang Uk; Lee, Jung Woo; Hyun, Dong Yun; Rhee, Sung-Keun; Hong, Chang Pyo; Bang, Kyong Hwan; Ryu, Hojin

2017-09-15

Korean ginseng ( Panax ginseng C.A. Meyer) has been widely used for medicinal purposes and contains potent plant secondary metabolites, including ginsenosides. To obtain transcriptomic data that offers a more comprehensive view of functional genomics in P. ginseng , we generated genome-wide transcriptome data from four different P. ginseng tissues using PacBio isoform sequencing (Iso-Seq) technology. A total of 135,317 assembled transcripts were generated with an average length of 3.2 kb and high assembly completeness. Of those unigenes, 67.5% were predicted to be complete full-length (FL) open reading frames (ORFs) and exhibited a high gene annotation rate. Furthermore, we successfully identified unique full-length genes involved in triterpenoid saponin synthesis and plant hormonal signaling pathways, including auxin and cytokinin. Studies on the functional genomics of P. ginseng seedlings have confirmed the rapid upregulation of negative feed-back loops by auxin and cytokinin signaling cues. The conserved evolutionary mechanisms in the auxin and cytokinin canonical signaling pathways of P. ginseng are more complex than those in Arabidopsis thaliana . Our analysis also revealed a more detailed view of transcriptome-wide alternative isoforms for 88 genes. Finally, transposable elements (TEs) were also identified, suggesting transcriptional activity of TEs in P. ginseng . In conclusion, our results suggest that long-read, full-length or partial-unigene data with high-quality assemblies are invaluable resources as transcriptomic references in P. ginseng and can be used for comparative analyses in closely related medicinal plants.

Identification of differentially expressed placental transcripts during multiple gestations in the Eurasian beaver (Castor fiber L.).

PubMed

Lipka, A; Paukszto, L; Majewska, M; Jastrzebski, J P; Myszczynski, K; Panasiewicz, G; Szafranska, B

2017-09-01

The Eurasian beaver is one of the largest rodents that, despite its high impact on the environment, is a non-model species that lacks a reference genome. Characterising genes critical for pregnancy outcome can serve as a basis for identifying mechanisms underlying effective reproduction, which is required for the success of endangered species conservation programs. In the present study, high-throughput RNA sequencing (RNA-seq) was used to analyse global changes in the Castor fiber subplacenta transcriptome during multiple pregnancy. De novo reconstruction of the C. fiber subplacenta transcriptome was used to identify genes that were differentially expressed in placentas (n=5) from two females (in advanced twin and triple pregnancy). Analyses of the expression values revealed 124 contigs with significantly different expression; of these, 55 genes were identified using MegaBLAST. Within this group of differentially expressed genes (DEGs), 18 were upregulated and 37 were downregulated in twins. Most DEGs were associated with the following gene ontology terms: cellular process, single organism process, response to stimulus, metabolic process and biological regulation. Some genes were also assigned to the developmental process, the reproductive process or reproduction. Among this group, four genes (namely keratin 19 (Krt19) and wingless-type MMTV integration site family - member 2 (Wnt2), which were downregulated in twins, and Nik-related kinase (Nrk) and gap junction protein β2 (Gjb2), which were upregulated in twins) were assigned to placental development and nine (Krt19, Wnt2 and integrin α 7 (Itga7), downregulated in twins, and Nrk, gap junction protein β6 (Gjb6), GATA binding protein 6 (Gata6), apolipoprotein A-I (ApoA1), apolipoprotein B (ApoB) and haemoglobin subunit α 1 (HbA1), upregulated in twins) were assigned to embryo development. The results of the present study indicate that the number of fetuses affects the expression profile in the C. fiber subplacental transcriptome. Enhancement of transcriptomic resources for C. fiber will improve understanding of the pathways relevant to proper placental development and successful reproduction.

An Integrated Proteomics/Transcriptomics Approach Points to Oxygen as the Main Electron Sink for Methanol Metabolism in Methylotenera mobilis▿†

PubMed Central

Beck, David A. C.; Hendrickson, Erik L.; Vorobev, Alexey; Wang, Tiansong; Lim, Sujung; Kalyuzhnaya, Marina G.; Lidstrom, Mary E.; Hackett, Murray; Chistoserdova, Ludmila

2011-01-01

Methylotenera species, unlike their close relatives in the genera Methylophilus, Methylobacillus, and Methylovorus, neither exhibit the activity of methanol dehydrogenase nor possess mxaFI genes encoding this enzyme, yet they are able to grow on methanol. In this work, we integrated a genome-wide proteomics approach, shotgun proteomics, and a genome-wide transcriptomics approach, shotgun transcriptome sequencing (RNA-seq), of Methylotenera mobilis JLW8 to identify genes and enzymes potentially involved in methanol oxidation, with special attention to alternative nitrogen sources, to address the question of whether nitrate could play a role as an electron acceptor in place of oxygen. Both proteomics and transcriptomics identified a limited number of genes and enzymes specifically responding to methanol. This set includes genes involved in oxidative stress response systems, a number of oxidoreductases, including XoxF-type alcohol dehydrogenases, a type II secretion system, and proteins without a predicted function. Nitrate stimulated expression of some genes in assimilatory nitrate reduction and denitrification pathways, while ammonium downregulated some of the nitrogen metabolism genes. However, none of these genes appeared to respond to methanol, which suggests that oxygen may be the main electron sink during growth on methanol. This study identifies initial targets for future focused physiological studies, including mutant analysis, which will provide further details into this novel process. PMID:21764938

Molecular diversity of toxic components from the scorpion Heterometrus petersii venom revealed by proteomic and transcriptome analysis.

PubMed

Ma, Yibao; Zhao, Yong; Zhao, Ruiming; Zhang, Weiping; He, Yawen; Wu, Yingliang; Cao, Zhijian; Guo, Lin; Li, Wenxin

2010-07-01

Scorpion venoms contain a vast untapped reservoir of natural products, which have the potential for medicinal value in drug discovery. In this study, toxin components from the scorpion Heterometrus petersii venom were evaluated by transcriptome and proteome analysis.Ten known families of venom peptides and proteins were identified, which include: two families of potassium channel toxins, four families of antimicrobial and cytolytic peptides,and one family from each of the calcium channel toxins, La1-like peptides, phospholipase A2,and the serine proteases. In addition, we also identified 12 atypical families, which include the acid phosphatases, diuretic peptides, and ten orphan families. From the data presented here, the extreme diversity and convergence of toxic components in scorpion venom was uncovered. Our work demonstrates the power of combining transcriptomic and proteomic approaches in the study of animal venoms.

Transcriptome analysis and related databases of Lactococcus lactis.

PubMed

Kuipers, Oscar P; de Jong, Anne; Baerends, Richard J S; van Hijum, Sacha A F T; Zomer, Aldert L; Karsens, Harma A; den Hengst, Chris D; Kramer, Naomi E; Buist, Girbe; Kok, Jan

2002-08-01

Several complete genome sequences of Lactococcus lactis and their annotations will become available in the near future, next to the already published genome sequence of L. lactis ssp. lactis IL 1403. This will allow intraspecies comparative genomics studies as well as functional genomics studies aimed at a better understanding of physiological processes and regulatory networks operating in lactococci. This paper describes the initial set-up of a DNA-microarray facility in our group, to enable transcriptome analysis of various Gram-positive bacteria, including a ssp. lactis and a ssp. cremoris strain of Lactococcus lactis. Moreover a global description will be given of the hardware and software requirements for such a set-up, highlighting the crucial integration of relevant bioinformatics tools and methods. This includes the development of MolGenIS, an information system for transcriptome data storage and retrieval, and LactococCye, a metabolic pathway/genome database of Lactococcus lactis.

Transcriptome Analysis Revealed Changes of Multiple Genes Involved in Haliotis discus hannai Innate Immunity during Vibrio parahemolyticus Infection.

PubMed

Nam, Bo-Hye; Jung, Myunghee; Subramaniyam, Sathiyamoorthy; Yoo, Seung-il; Markkandan, Kesavan; Moon, Ji-Young; Kim, Young-Ok; Kim, Dong-Gyun; An, Cheul Min; Shin, Younhee; Jung, Ho-jin; Park, Jun-hyung

2016-01-01

Abalone (Haliotis discus hannai) is one of the most valuable marine aquatic species in Korea, Japan and China. Tremendous exposure to bacterial infection is common in aquaculture environment, especially by Vibrio sp. infections. It's therefore necessary and urgent to understand the mechanism of H. discus hannai host defense against Vibrio parahemolyticus infection. However studies on its immune system are hindered by the lack of genomic resources. In the present study, we sequenced the transcriptome of control and bacterial challenged H. discus hannai tissues. Totally, 138 MB of reference transcriptome were obtained from de novo assembly of 34 GB clean bases from ten different libraries and annotated with the biological terms (GO and KEGG). A total of 10,575 transcripts exhibiting the differentially expression at least one pair of comparison and the functional annotations highlight genes related to immune response, cell adhesion, immune regulators, redox molecules and mitochondrial coding genes. Mostly, these groups of genes were dominated in hemocytes compared to other tissues. This work is a prerequisite for the identification of those physiological traits controlling H. discus hannai ability to survive against Vibrio infection.

Transcriptome Analysis Revealed Changes of Multiple Genes Involved in Haliotis discus hannai Innate Immunity during Vibrio parahemolyticus Infection

PubMed Central

Nam, Bo-Hye; Jung, Myunghee; Subramaniyam, Sathiyamoorthy; Yoo, Seung-il; Markkandan, Kesavan; Moon, Ji-Young; Kim, Young-Ok; Kim, Dong-Gyun; An, Cheul Min; Shin, Younhee; Jung, Ho-jin; Park, Jun-hyung

2016-01-01

Abalone (Haliotis discus hannai) is one of the most valuable marine aquatic species in Korea, Japan and China. Tremendous exposure to bacterial infection is common in aquaculture environment, especially by Vibrio sp. infections. It’s therefore necessary and urgent to understand the mechanism of H. discus hannai host defense against Vibrio parahemolyticus infection. However studies on its immune system are hindered by the lack of genomic resources. In the present study, we sequenced the transcriptome of control and bacterial challenged H. discus hannai tissues. Totally, 138 MB of reference transcriptome were obtained from de novo assembly of 34 GB clean bases from ten different libraries and annotated with the biological terms (GO and KEGG). A total of 10,575 transcripts exhibiting the differentially expression at least one pair of comparison and the functional annotations highlight genes related to immune response, cell adhesion, immune regulators, redox molecules and mitochondrial coding genes. Mostly, these groups of genes were dominated in hemocytes compared to other tissues. This work is a prerequisite for the identification of those physiological traits controlling H. discus hannai ability to survive against Vibrio infection. PMID:27088873

Intra-isolate genome variation in arbuscular mycorrhizal fungi persists in the transcriptome.

PubMed

Boon, E; Zimmerman, E; Lang, B F; Hijri, M

2010-07-01

Arbuscular mycorrhizal fungi (AMF) are heterokaryotes with an unusual genetic makeup. Substantial genetic variation occurs among nuclei within a single mycelium or isolate. AMF reproduce through spores that contain varying fractions of this heterogeneous population of nuclei. It is not clear whether this genetic variation on the genome level actually contributes to the AMF phenotype. To investigate the extent to which polymorphisms in nuclear genes are transcribed, we analysed the intra-isolate genomic and cDNA sequence variation of two genes, the large subunit ribosomal RNA (LSU rDNA) of Glomus sp. DAOM-197198 (previously known as G. intraradices) and the POL1-like sequence (PLS) of Glomus etunicatum. For both genes, we find high sequence variation at the genome and transcriptome level. Reconstruction of LSU rDNA secondary structure shows that all variants are functional. Patterns of PLS sequence polymorphism indicate that there is one functional gene copy, PLS2, which is preferentially transcribed, and one gene copy, PLS1, which is a pseudogene. This is the first study that investigates AMF intra-isolate variation at the transcriptome level. In conclusion, it is possible that, in AMF, multiple nuclear genomes contribute to a single phenotype.

Signatures of Rapid Evolution in Urban and Rural Transcriptomes of White-Footed Mice (Peromyscus leucopus) in the New York Metropolitan Area

PubMed Central

Harris, Stephen E.; Munshi-South, Jason; Obergfell, Craig; O’Neill, Rachel

2013-01-01

Urbanization is a major cause of ecological degradation around the world, and human settlement in large cities is accelerating. New York City (NYC) is one of the oldest and most urbanized cities in North America, but still maintains 20% vegetation cover and substantial populations of some native wildlife. The white-footed mouse, Peromyscus leucopus , is a common resident of NYC’s forest fragments and an emerging model system for examining the evolutionary consequences of urbanization. In this study, we developed transcriptomic resources for urban P . leucopus to examine evolutionary changes in protein-coding regions for an exemplar “urban adapter.” We used Roche 454 GS FLX+ high throughput sequencing to derive transcriptomes from multiple tissues from individuals across both urban and rural populations. From these data, we identified 31,015 SNPs and several candidate genes potentially experiencing positive selection in urban populations of P . leucopus . These candidate genes are involved in xenobiotic metabolism, innate immune response, demethylation activity, and other important biological phenomena in novel urban environments. This study is one of the first to report candidate genes exhibiting signatures of directional selection in divergent urban ecosystems. PMID:24015321

Informatic deconvolution of biased GPCR signaling mechanisms from in vivo pharmacological experimentation.

PubMed

Maudsley, Stuart; Martin, Bronwen; Janssens, Jonathan; Etienne, Harmonie; Jushaj, Areta; van Gastel, Jaana; Willemsen, Ann; Chen, Hongyu; Gesty-Palmer, Diane; Luttrell, Louis M

2016-01-01

Ligands possessing different physico-chemical structures productively interact with G protein-coupled receptors generating distinct downstream signaling events due to their abilities to activate/select idiosyncratic receptor entities ('receptorsomes') from the full spectrum of potential receptor partners. We have employed multiple novel informatic approaches to identify and characterize the in vivo transcriptomic signature of an arrestin-signaling biased ligand, [D-Trp(12),Tyr(34)]-bPTH(7-34), acting at the parathyroid hormone type 1 receptor (PTH1R), across six different murine tissues after chronic drug exposure. We are able to demonstrate that [D-Trp(12),Tyr(34)]-bPTH(7-34) elicits a distinctive arrestin-signaling focused transcriptomic response that is more coherently regulated, in an arrestin signaling-dependent manner, across more tissues than that of the pluripotent endogenous PTH1R ligand, hPTH(1-34). This arrestin-focused response signature is strongly linked with the transcriptional regulation of cell growth and development. Our informatic deconvolution of a conserved arrestin-dependent transcriptomic signature from wild type mice demonstrates a conceptual framework within which the in vivo outcomes of biased receptor signaling may be further investigated or predicted. Published by Elsevier Inc.

Transcriptomic data analysis and differential gene expression of antioxidant pathways in king penguin juveniles (Aptenodytes patagonicus) before and after acclimatization to marine life.

PubMed

Rey, Benjamin; Dégletagne, Cyril; Duchamp, Claude

2016-12-01

In this article, we present differentially expressed gene profiles in the pectoralis muscle of wild juvenile king penguins that were either naturally acclimated to cold marine environment or experimentally immersed in cold water as compared with penguin juveniles that never experienced cold water immersion. Transcriptomic data were obtained by hybridizing penguins total cDNA on Affymetrix GeneChip Chicken Genome arrays and analyzed using maxRS algorithm , " Transcriptome analysis in non-model species: a new method for the analysis of heterologous hybridization on microarrays " (Dégletagne et al., 2010) [1] . We focused on genes involved in multiple antioxidant pathways. For better clarity, these differentially expressed genes were clustered into six functional groups according to their role in controlling redox homeostasis. The data are related to a comprehensive research study on the ontogeny of antioxidant functions in king penguins, "Hormetic response triggers multifaceted anti-oxidant strategies in immature king penguins (Aptenodytes patagonicus)" (Rey et al., 2016) [2] . The raw microarray dataset supporting the present analyses has been deposited at the Gene Expression Omnibus (GEO) repository under accessions GEO: GSE17725 and GEO: GSE82344.

Spatial transcriptomic analysis of cryosectioned tissue samples with Geo-seq.

PubMed

Chen, Jun; Suo, Shengbao; Tam, Patrick Pl; Han, Jing-Dong J; Peng, Guangdun; Jing, Naihe

2017-03-01

Conventional gene expression studies analyze multiple cells simultaneously or single cells, for which the exact in vivo or in situ position is unknown. Although cellular heterogeneity can be discerned when analyzing single cells, any spatially defined attributes that underpin the heterogeneous nature of the cells cannot be identified. Here, we describe how to use Geo-seq, a method that combines laser capture microdissection (LCM) and single-cell RNA-seq technology. The combination of these two methods enables the elucidation of cellular heterogeneity and spatial variance simultaneously. The Geo-seq protocol allows the profiling of transcriptome information from only a small number cells and retains their native spatial information. This protocol has wide potential applications to address biological and pathological questions of cellular properties such as prospective cell fates, biological function and the gene regulatory network. Geo-seq has been applied to investigate the spatial transcriptome of mouse early embryo, mouse brain, and pathological liver and sperm tissues. The entire protocol from tissue collection and microdissection to sequencing requires ∼5 d, Data analysis takes another 1 or 2 weeks, depending on the amount of data and the speed of the processor.

Reefgenomics.Org - a repository for marine genomics data.

PubMed

Liew, Yi Jin; Aranda, Manuel; Voolstra, Christian R

2016-01-01

Over the last decade, technological advancements have substantially decreased the cost and time of obtaining large amounts of sequencing data. Paired with the exponentially increased computing power, individual labs are now able to sequence genomes or transcriptomes to investigate biological questions of interest. This has led to a significant increase in available sequence data. Although the bulk of data published in articles are stored in public sequence databases, very often, only raw sequencing data are available; miscellaneous data such as assembled transcriptomes, genome annotations etc. are not easily obtainable through the same means. Here, we introduce our website (http://reefgenomics.org) that aims to centralize genomic and transcriptomic data from marine organisms. Besides providing convenient means to download sequences, we provide (where applicable) a genome browser to explore available genomic features, and a BLAST interface to search through the hosted sequences. Through the interface, multiple datasets can be queried simultaneously, allowing for the retrieval of matching sequences from organisms of interest. The minimalistic, no-frills interface reduces visual clutter, making it convenient for end-users to search and explore processed sequence data. DATABASE URL: http://reefgenomics.org. © The Author(s) 2016. Published by Oxford University Press.

Transcriptome Analysis of Manganese-deficient Chlamydomonas reinhardtii Provides Insight on the Chlorophyll Biosynthesis Pathway

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

Lockhart, Ainsley; Zvenigorodsky, Natasha; Pedraza, Mary Ann

2011-08-11

The biosynthesis of chlorophyll and other tetrapyrroles is a vital but poorly understood process. Recent genomic advances with the unicellular green algae Chlamydomonas reinhardtii have created opportunity to more closely examine the mechanisms of the chlorophyll biosynthesis pathway via transcriptome analysis. Manganese is a nutrient of interest for complex reactions because of its multiple stable oxidation states and role in molecular oxygen coordination. C. reinhardtii was cultured in Manganese-deplete Tris-acetate-phosphate (TAP) media for 24 hours and used to create cDNA libraries for sequencing using Illumina TruSeq technology. Transcriptome analysis provided intriguing insight on possible regulatory mechanisms in the pathway. Evidencemore » supports similarities of GTR (Glutamyl-tRNA synthase) to its Chlorella vulgaris homolog in terms of Mn requirements. Data was also suggestive of Mn-related compensatory up-regulation for pathway proteins CHLH1 (Manganese Chelatase), GUN4 (Magnesium chelatase activating protein), and POR1 (Light-dependent protochlorophyllide reductase). Intriguingly, data suggests possible reciprocal expression of oxygen dependent CPX1 (coproporphyrinogen III oxidase) and oxygen independent CPX2. Further analysis using RT-PCR could provide compelling evidence for several novel regulatory mechanisms in the chlorophyll biosynthesis pathway.« less

Spliced synthetic genes as internal controls in RNA sequencing experiments.

PubMed

Hardwick, Simon A; Chen, Wendy Y; Wong, Ted; Deveson, Ira W; Blackburn, James; Andersen, Stacey B; Nielsen, Lars K; Mattick, John S; Mercer, Tim R

2016-09-01

RNA sequencing (RNA-seq) can be used to assemble spliced isoforms, quantify expressed genes and provide a global profile of the transcriptome. However, the size and diversity of the transcriptome, the wide dynamic range in gene expression and inherent technical biases confound RNA-seq analysis. We have developed a set of spike-in RNA standards, termed 'sequins' (sequencing spike-ins), that represent full-length spliced mRNA isoforms. Sequins have an entirely artificial sequence with no homology to natural reference genomes, but they align to gene loci encoded on an artificial in silico chromosome. The combination of multiple sequins across a range of concentrations emulates alternative splicing and differential gene expression, and it provides scaling factors for normalization between samples. We demonstrate the use of sequins in RNA-seq experiments to measure sample-specific biases and determine the limits of reliable transcript assembly and quantification in accompanying human RNA samples. In addition, we have designed a complementary set of sequins that represent fusion genes arising from rearrangements of the in silico chromosome to aid in cancer diagnosis. RNA sequins provide a qualitative and quantitative reference with which to navigate the complexity of the human transcriptome.

Cross-omics comparison of stress responses in mesothelial cells exposed to heat- versus filter-sterilized peritoneal dialysis fluids.

PubMed

Kratochwill, Klaus; Bender, Thorsten O; Lichtenauer, Anton M; Herzog, Rebecca; Tarantino, Silvia; Bialas, Katarzyna; Jörres, Achim; Aufricht, Christoph

2015-01-01

Recent research suggests that cytoprotective responses, such as expression of heat-shock proteins, might be inadequately induced in mesothelial cells by heat-sterilized peritoneal dialysis (PD) fluids. This study compares transcriptome data and multiple protein expression profiles for providing new insight into regulatory mechanisms. Two-dimensional difference gel electrophoresis (2D-DIGE) based proteomics and topic defined gene expression microarray-based transcriptomics techniques were used to evaluate stress responses in human omental peritoneal mesothelial cells in response to heat- or filter-sterilized PD fluids. Data from selected heat-shock proteins were validated by 2D western-blot analysis. Comparison of proteomics and transcriptomics data discriminated differentially regulated protein abundance into groups depending on correlating or noncorrelating transcripts. Inadequate abundance of several heat-shock proteins following exposure to heat-sterilized PD fluids is not reflected on the mRNA level indicating interference beyond transcriptional regulation. For the first time, this study describes evidence for posttranscriptional inadequacy of heat-shock protein expression by heat-sterilized PD fluids as a novel cytotoxic property. Cross-omics technologies introduce a novel way of understanding PDF bioincompatibility and searching for new interventions to reestablish adequate cytoprotective responses.

Transcriptome differences between enrofloxacin-resistant and enrofloxacin-susceptible strains of Aeromonas hydrophila.

PubMed

Zhu, Fengjiao; Yang, Zongying; Zhang, Yiliu; Hu, Kun; Fang, Wenhong

2017-01-01

Enrofloxacin is the most commonly used antibiotic to control diseases in aquatic animals caused by A. hydrophila. This study conducted de novo transcriptome sequencing and compared the global transcriptomes of enrofloxacin-resistant and enrofloxacin-susceptible strains. We got a total of 4,714 unigenes were assembled. Of these, 4,122 were annotated. A total of 3,280 unigenes were assigned to GO, 3,388 unigenes were classified into Cluster of Orthologous Groups of proteins (COG) using BLAST and BLAST2GO software, and 2,568 were mapped onto pathways using the Kyoto Encyclopedia of Gene and Genomes Pathway database. Furthermore, 218 unigenes were deemed to be DEGs. After enrofloxacin treatment, 135 genes were upregulated and 83 genes were downregulated. The GO terms biological process (126 genes) and metabolic process (136 genes) were the most enriched, and the terms for protein folding, response to stress, and SOS response were also significantly enriched. This study identified enrofloxacin treatment affects multiple biological functions of A. hydrophila. Enrofloxacin resistance in A. hydrophila is closely related to the reduction of intracellular drug accumulation caused by ABC transporters and increased expression of topoisomerase IV.

Transcriptome differences between enrofloxacin-resistant and enrofloxacin-susceptible strains of Aeromonas hydrophila

PubMed Central

Yang, Zongying; Zhang, Yiliu; Hu, Kun; Fang, Wenhong

2017-01-01

Enrofloxacin is the most commonly used antibiotic to control diseases in aquatic animals caused by A. hydrophila. This study conducted de novo transcriptome sequencing and compared the global transcriptomes of enrofloxacin-resistant and enrofloxacin-susceptible strains. We got a total of 4,714 unigenes were assembled. Of these, 4,122 were annotated. A total of 3,280 unigenes were assigned to GO, 3,388 unigenes were classified into Cluster of Orthologous Groups of proteins (COG) using BLAST and BLAST2GO software, and 2,568 were mapped onto pathways using the Kyoto Encyclopedia of Gene and Genomes Pathway database. Furthermore, 218 unigenes were deemed to be DEGs. After enrofloxacin treatment, 135 genes were upregulated and 83 genes were downregulated. The GO terms biological process (126 genes) and metabolic process (136 genes) were the most enriched, and the terms for protein folding, response to stress, and SOS response were also significantly enriched. This study identified enrofloxacin treatment affects multiple biological functions of A. hydrophila. Enrofloxacin resistance in A. hydrophila is closely related to the reduction of intracellular drug accumulation caused by ABC transporters and increased expression of topoisomerase IV. PMID:28708867

Antennal transcriptome analysis of the Asian longhorned beetle Anoplophora glabripennis

PubMed Central

Hu, Ping; Wang, Jingzhen; Cui, Mingming; Tao, Jing; Luo, Youqing

2016-01-01

Olfactory proteins form the basis of insect olfactory recognition, which is crucial for host identification, mating, and oviposition. Using transcriptome analysis of Anoplophora glabripennis antenna, we identified 42 odorant-binding proteins (OBPs), 12 chemosensory proteins (CSPs), 14 pheromone-degrading enzymes (PDEs), 1 odorant-degrading enzymes (ODE), 37 odorant receptors (ORs), 11 gustatory receptors (GRs), 2 sensory neuron membrane proteins (SNMPs), and 4 ionotropic receptor (IR). All CSPs and PBPs were expressed in antennae, confirming the authenticity of the transcriptome data. CSP expression profiles showed that AglaCSP3, AglaCSP6, and AglaCSP12 were expressed preferentially in maxillary palps and AglaCSP7 and AglaCSP9 were strongly expressed in antennae. The vast majority of CSPs were highly expressed in multiple chemosensory tissues, suggesting their participation in olfactory recognition in almost all olfactory tissues. Intriguingly, the PBP AglaPBP2 was preferentially expressed in antenna, indicating that it is the main protein involved in efficient and sensitive pheromone recognition. Phylogenetic analysis of olfactory proteins indicated AglaGR1 may detect CO2. This study establishes a foundation for determining the chemoreception molecular mechanisms of A. glabripennis, which would provide a new perspective for controlling pest populations, especially those of borers. PMID:27222053

Multitissue Transcriptomics Delineates the Diversity of Airway T Cell Functions in Asthma.

PubMed

Singhania, Akul; Wallington, Joshua C; Smith, Caroline G; Horowitz, Daniel; Staples, Karl J; Howarth, Peter H; Gadola, Stephan D; Djukanović, Ratko; Woelk, Christopher H; Hinks, Timothy S C

2018-02-01

Asthma arises from the complex interplay of inflammatory pathways in diverse cell types and tissues. We sought to undertake a comprehensive transcriptomic assessment of the epithelium and airway T cells that remain understudied in asthma and investigate interactions between multiple cells and tissues. Epithelial brushings and flow-sorted CD3 + T cells from sputum and BAL were obtained from healthy subjects (n = 19) and patients with asthma (mild, moderate, and severe asthma; n = 46). Gene expression was assessed using Affymetrix HT HG-U133 + PM GeneChips, and results were validated by real-time quantitative PCR. In the epithelium, IL-13 response genes (POSTN, SERPINB2, and CLCA1), mast cell mediators (CPA3 and TPSAB1), inducible nitric oxide synthase, and cystatins (CST1, CST2, and CST4) were upregulated in mild asthma, but, except for cystatins, were suppressed by corticosteroids in moderate asthma. In severe asthma-with predominantly neutrophilic phenotype-several distinct processes were upregulated, including neutrophilia (TCN1 and MMP9), mucins, and oxidative stress responses. The majority of the disease signature was evident in sputum T cells in severe asthma, where 267 genes were differentially regulated compared with health, highlighting compartmentalization of inflammation. This signature included IL-17-inducible chemokines (CXCL1, CXCL2, CXCL3, IL8, and CSF3) and chemoattractants for neutrophils (IL8, CCL3, and LGALS3), T cells, and monocytes. A protein interaction network in severe asthma highlighted signatures of responses to bacterial infections across tissues (CEACAM5, CD14, and TLR2), including Toll-like receptor signaling. In conclusion, the activation of innate immune pathways in the airways suggests that activated T cells may be driving neutrophilic inflammation and steroid-insensitive IL-17 response in severe asthma.

Transcriptomic studies reveal a key metabolic pathway contributing to a well-maintained photosynthetic system under drought stress in foxtail millet (Setaria italica L.).

PubMed

Shi, Weiping; Cheng, Jingye; Wen, Xiaojie; Wang, Jixiang; Shi, Guanyan; Yao, Jiayan; Hou, Liyuan; Sun, Qian; Xiang, Peng; Yuan, Xiangyang; Dong, Shuqi; Guo, Pingyi; Guo, Jie

2018-01-01

Drought stress is one of the most important abiotic factors limiting crop productivity. A better understanding of the effects of drought on millet ( Setaria italica L.) production, a model crop for studying drought tolerance, and the underlying molecular mechanisms responsible for drought stress responses is vital to improvement of agricultural production. In this study, we exposed the drought resistant F 1 hybrid, M79, and its parental lines E1 and H1 to drought stress. Subsequent physiological analysis demonstrated that M79 showed higher photosynthetic energy conversion efficiency and drought tolerance than its parents. A transcriptomic study using leaves collected six days after drought treatment, when the soil water content was about ∼20%, identified 3066, 1895, and 2148 differentially expressed genes (DEGs) in M79, E1 and H1 compared to the respective untreated controls, respectively. Further analysis revealed 17 Gene Ontology (GO) enrichments and 14 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways in M79, including photosystem II (PSII) oxygen-evolving complex, peroxidase (POD) activity, plant hormone signal transduction, and chlorophyll biosynthesis. Co-regulation analysis suggested that these DEGs in M79 contributed to the formation of a regulatory network involving multiple biological processes and pathways including photosynthesis, signal transduction, transcriptional regulation, redox regulation, hormonal signaling, and osmotic regulation. RNA-seq analysis also showed that some photosynthesis-related DEGs were highly expressed in M79 compared to its parental lines under drought stress. These results indicate that various molecular pathways, including photosynthesis, respond to drought stress in M79, and provide abundant molecular information for further analysis of the underlying mechanism responding to this stress.

A house finch (Haemorhous mexicanus) spleen transcriptome reveals intra- and interspecific patterns of gene expression, alternative splicing and genetic diversity in passerines

PubMed Central

2014-01-01

Background With its plumage color dimorphism and unique history in North America, including a recent population expansion and an epizootic of Mycoplasma gallisepticum (MG), the house finch (Haemorhous mexicanus) is a model species for studying sexual selection, plumage coloration and host-parasite interactions. As part of our ongoing efforts to make available genomic resources for this species, here we report a transcriptome assembly derived from genes expressed in spleen. Results We characterize transcriptomes from two populations with different histories of demography and disease exposure: a recently founded population in the eastern US that has been exposed to MG for over a decade and a native population from the western range that has never been exposed to MG. We utilize this resource to quantify conservation in gene expression in passerine birds over approximately 50 MY by comparing splenic expression profiles for 9,646 house finch transcripts and those from zebra finch and find that less than half of all genes expressed in spleen in either species are expressed in both species. Comparative gene annotations from several vertebrate species suggest that the house finch transcriptomes contain ~15 genes not yet found in previously sequenced vertebrate genomes. The house finch transcriptomes harbour ~85,000 SNPs, ~20,000 of which are non-synonymous. Although not yet validated by biological or technical replication, we identify a set of genes exhibiting differences between populations in gene expression (n = 182; 2% of all transcripts), allele frequencies (76 FST ouliers) and alternative splicing as well as genes with several fixed non-synonymous substitutions; this set includes genes with functions related to double-strand break repair and immune response. Conclusions The two house finch spleen transcriptome profiles will add to the increasing data on genome and transcriptome sequence information from natural populations. Differences in splenic expression between house finch and zebra finch imply either significant evolutionary turnover of splenic expression patterns or different physiological states of the individuals examined. The transcriptome resource will enhance the potential to annotate an eventual house finch genome, and the set of gene-based high-quality SNPs will help clarify the genetic underpinnings of host-pathogen interactions and sexual selection. PMID:24758272

The response of Isidorella newcombi to copper exposure: Using an integrated biological framework to interpret transcriptomic responses from RNA-seq analysis.

PubMed

Ubrihien, Rodney P; Ezaz, Tariq; Taylor, Anne M; Stevens, Mark M; Krikowa, Frank; Foster, Simon; Maher, William A

2017-04-01

This study describes the transcriptomic response of the Australian endemic freshwater gastropod Isidorella newcombi exposed to 80±1μg/L of copper for 3days. Analysis of copper tissue concentration, lysosomal membrane destabilisation and RNA-seq were conducted. Copper tissue concentrations confirmed that copper was bioaccumulated by the snails. Increased lysosomal membrane destabilisation in the copper-exposed snails indicated that the snails were stressed as a result of the exposure. Both copper tissue concentrations and lysosomal destabilisation were significantly greater in snails exposed to copper. In order to interpret the RNA-seq data from an ecotoxicological perspective an integrated biological response model was developed that grouped transcriptomic responses into those associated with copper transport and storage, survival mechanisms and cell death. A conceptual model of expected transcriptomic changes resulting from the copper exposure was developed as a basis to assess transcriptomic responses. Transcriptomic changes were evident at all the three levels of the integrated biological response model. Despite lacking statistical significance, increased expression of the gene encoding copper transporting ATPase provided an indication of increased internal transport of copper. Increased expression of genes associated with endocytosis are associated with increased transport of copper to the lysosome for storage in a detoxified form. Survival mechanisms included metabolic depression and processes associated with cellular repair and recycling. There was transcriptomic evidence of increased cell death by apoptosis in the copper-exposed organisms. Increased apoptosis is supported by the increase in lysosomal membrane destabilisation in the copper-exposed snails. Transcriptomic changes relating to apoptosis, phagocytosis, protein degradation and the lysosome were evident and these processes can be linked to the degradation of post-apoptotic debris. The study identified contaminant specific transcriptomic markers as well as markers of general stress. From an ecotoxicological perspective, the use of a framework to group transcriptomic responses into those associated with copper transport, survival and cell death assisted with the complex process of interpretation of RNA-seq data. The broad adoption of such a framework in ecotoxicology studies would assist in comparison between studies and the identification of reliable transcriptomic markers of contaminant exposure and response. Copyright © 2017 Elsevier B.V. All rights reserved.

Homoeolog-specific activation of genes for heat acclimation in the allopolyploid grass Brachypodium hybridum.

PubMed

Takahagi, Kotaro; Inoue, Komaki; Shimizu, Minami; Uehara-Yamaguchi, Yukiko; Onda, Yoshihiko; Mochida, Keiichi

2018-04-01

Allopolyploid plants often show wider environmental tolerances than their ancestors; this is expected to be due to the merger of multiple distinct genomes with a fixed heterozygosity. The complex homoeologous gene expression could have been evolutionarily advantageous for the adaptation of allopolyploid plants. Despite multiple previous studies reporting homoeolog-specific gene expression in allopolyploid species, there are no clear examples of homoeolog-specific function in acclimation to a long-term stress condition. We found that the allopolyploid grass Brachypodium hybridum and its ancestor Brachypodium stacei show long-term heat stress tolerance, unlike its other ancestor, Brachypodium distachyon. To understand the physiological traits of B. hybridum, we compared the transcriptome of the 3 Brachypodium species grown under normal and heat stress conditions. We found that the expression patterns of approximately 26% and approximately 38% of the homoeolog groups in B. hybridum changed toward nonadditive expression and nonancestral expression, respectively, under normal condition. Moreover, we found that B. distachyon showed similar expression patterns between normal and heat stress conditions, whereas B. hybridum and B. stacei significantly altered their transcriptome in response to heat after 3 days of stress exposure, and homoeologs that were inherited from B. stacei may have contributed to the transcriptional stress response to heat in B. hybridum. After 15 days of heat exposure, B. hybridum and B. stacei maintained transcriptional states similar to those under normal conditions. These results suggest that an earlier response to heat that was specific to homoeologs originating from B. stacei contributed to cellular homeostasis under long-term heat stress in B. hybridum. Our results provide insights into different regulatory events of the homoeo-transcriptome that are associated with stress acclimation in allopolyploid plants.

RNA-Seq Meta-analysis identifies genes in skeletal muscle associated with gain and intake across a multi-season study of crossbred beef steers.

PubMed

Keel, Brittney N; Zarek, Christina M; Keele, John W; Kuehn, Larry A; Snelling, Warren M; Oliver, William T; Freetly, Harvey C; Lindholm-Perry, Amanda K

2018-06-04

Feed intake and body weight gain are economically important inputs and outputs of beef production systems. The purpose of this study was to discover differentially expressed genes that will be robust for feed intake and gain across a large segment of the cattle industry. Transcriptomic studies often suffer from issues with reproducibility and cross-validation. One way to improve reproducibility is by integrating multiple datasets via meta-analysis. RNA sequencing (RNA-Seq) was performed on longissimus dorsi muscle from 80 steers (5 cohorts, each with 16 animals) selected from the outside fringe of a bivariate gain and feed intake distribution to understand the genes and pathways involved in feed efficiency. In each cohort, 16 steers were selected from one of four gain and feed intake phenotypes (n = 4 per phenotype) in a 2 × 2 factorial arrangement with gain and feed intake as main effect variables. Each cohort was analyzed as a single experiment using a generalized linear model and results from the 5 cohort analyses were combined in a meta-analysis to identify differentially expressed genes (DEG) across the cohorts. A total of 51 genes were differentially expressed for the main effect of gain, 109 genes for the intake main effect, and 11 genes for the gain x intake interaction (P corrected  < 0.05). A jackknife sensitivity analysis showed that, in general, the meta-analysis produced robust DEGs for the two main effects and their interaction. Pathways identified from over-represented genes included mitochondrial energy production and oxidative stress pathways for the main effect of gain due to DEG including GPD1, NDUFA6, UQCRQ, ACTC1, and MGST3. For intake, metabolic pathways including amino acid biosynthesis and degradation were identified, and for the interaction analysis the pathways identified included GADD45, pyridoxal 5'phosphate salvage, and caveolar mediated endocytosis signaling. Variation among DEG identified by cohort suggests that environment and breed may play large roles in the expression of genes associated with feed efficiency in the muscle of beef cattle. Meta-analyses of transcriptome data from groups of animals over multiple cohorts may be necessary to elucidate the genetics contributing these types of biological phenotypes.

The developmental transcriptome atlas of the spoon worm Urechis unicinctus (Echiurida: Annelida).

PubMed

Park, Chungoo; Han, Yong-Hee; Lee, Sung-Gwon; Ry, Kyoung-Bin; Oh, Jooseong; Kern, Elizabeth M A; Park, Joong-Ki; Cho, Sung-Jin

2018-03-01

Echiurida is one of the most intriguing major subgroups of annelida because, unlike most other annelids, echiurids lack metameric body segmentation as adults. For this reason, transcriptome analyses from various developmental stages of echiurid species can be of substantial value for understanding precise expression levels and the complex regulatory networks during early and larval development. A total of 914 million raw RNA-Seq reads were produced from 14 developmental stages of Urechis unicinctus and were de novo assembled into contigs spanning 63,928,225 bp with an N50 length of 2700 bp. The resulting comprehensive transcriptome database of the early developmental stages of U. unicinctus consists of 20,305 representative functional protein-coding transcripts. Approximately 66% of unigenes were assigned to superphylum-level taxa, including Lophotrochozoa (40%). The completeness of the transcriptome assembly was assessed using benchmarking universal single-copy orthologs; 75.7% of the single-copy orthologs were presented in our transcriptome database. We observed 3 distinct patterns of global transcriptome profiles from 14 developmental stages and identified 12,705 genes that showed dynamic regulation patterns during the differentiation and maturation of U. unicinctus cells. We present the first large-scale developmental transcriptome dataset of U. unicinctus and provide a general overview of the dynamics of global gene expression changes during its early developmental stages. The analysis of time-course gene expression data is a first step toward understanding the complex developmental gene regulatory networks in U. unicinctus and will furnish a valuable resource for analyzing the functions of gene repertoires in various developmental phases.

H7N9 and Other Pathogenic Avian Influenza Viruses Elicit a Three-Pronged Transcriptomic Signature That Is Reminiscent of 1918 Influenza Virus and Is Associated with Lethal Outcome in Mice

PubMed Central

Morrison, Juliet; Josset, Laurence; Tchitchek, Nicolas; Chang, Jean; Belser, Jessica A.; Swayne, David E.; Pantin-Jackwood, Mary J.; Tumpey, Terrence M.

2014-01-01

ABSTRACT Modulating the host response is a promising approach to treating influenza, caused by a virus whose pathogenesis is determined in part by the reaction it elicits within the host. Though the pathogenicity of emerging H7N9 influenza virus in several animal models has been reported, these studies have not included a detailed characterization of the host response following infection. Therefore, we characterized the transcriptomic response of BALB/c mice infected with H7N9 (A/Anhui/01/2013) virus and compared it to the responses induced by H5N1 (A/Vietnam/1203/2004), H7N7 (A/Netherlands/219/2003), and pandemic 2009 H1N1 (A/Mexico/4482/2009) influenza viruses. We found that responses to the H7 subtype viruses were intermediate to those elicited by H5N1 and pdm09H1N1 early in infection but that they evolved to resemble the H5N1 response as infection progressed. H5N1, H7N7, and H7N9 viruses were pathogenic in mice, and this pathogenicity correlated with increased transcription of cytokine response genes and decreased transcription of lipid metabolism and coagulation signaling genes. This three-pronged transcriptomic signature was observed in mice infected with pathogenic H1N1 strains such as the 1918 virus, indicating that it may be predictive of pathogenicity across multiple influenza virus strains. Finally, we used host transcriptomic profiling to computationally predict drugs that reverse the host response to H7N9 infection, and we identified six FDA-approved drugs that could potentially be repurposed to treat H7N9 and other pathogenic influenza viruses. IMPORTANCE Emerging avian influenza viruses are of global concern because the human population is immunologically naive to them. Current influenza drugs target viral molecules, but the high mutation rate of influenza viruses eventually leads to the development of antiviral resistance. As the host evolves far more slowly than the virus, and influenza pathogenesis is determined in part by the host response, targeting the host response is a promising approach to treating influenza. Here we characterize the host transcriptomic response to emerging H7N9 influenza virus and compare it with the responses to H7N7, H5N1, and pdm09H1N1. All three avian viruses were pathogenic in mice and elicited a transcriptomic signature that also occurs in response to the legendary 1918 influenza virus. Our work identifies host responses that could be targeted to treat severe H7N9 influenza and identifies six FDA-approved drugs that could potentially be repurposed as H7N9 influenza therapeutics. PMID:24991006

H7N9 and other pathogenic avian influenza viruses elicit a three-pronged transcriptomic signature that is reminiscent of 1918 influenza virus and is associated with lethal outcome in mice.

PubMed

Morrison, Juliet; Josset, Laurence; Tchitchek, Nicolas; Chang, Jean; Belser, Jessica A; Swayne, David E; Pantin-Jackwood, Mary J; Tumpey, Terrence M; Katze, Michael G

2014-09-01

Modulating the host response is a promising approach to treating influenza, caused by a virus whose pathogenesis is determined in part by the reaction it elicits within the host. Though the pathogenicity of emerging H7N9 influenza virus in several animal models has been reported, these studies have not included a detailed characterization of the host response following infection. Therefore, we characterized the transcriptomic response of BALB/c mice infected with H7N9 (A/Anhui/01/2013) virus and compared it to the responses induced by H5N1 (A/Vietnam/1203/2004), H7N7 (A/Netherlands/219/2003), and pandemic 2009 H1N1 (A/Mexico/4482/2009) influenza viruses. We found that responses to the H7 subtype viruses were intermediate to those elicited by H5N1 and pdm09H1N1 early in infection but that they evolved to resemble the H5N1 response as infection progressed. H5N1, H7N7, and H7N9 viruses were pathogenic in mice, and this pathogenicity correlated with increased transcription of cytokine response genes and decreased transcription of lipid metabolism and coagulation signaling genes. This three-pronged transcriptomic signature was observed in mice infected with pathogenic H1N1 strains such as the 1918 virus, indicating that it may be predictive of pathogenicity across multiple influenza virus strains. Finally, we used host transcriptomic profiling to computationally predict drugs that reverse the host response to H7N9 infection, and we identified six FDA-approved drugs that could potentially be repurposed to treat H7N9 and other pathogenic influenza viruses. Emerging avian influenza viruses are of global concern because the human population is immunologically naive to them. Current influenza drugs target viral molecules, but the high mutation rate of influenza viruses eventually leads to the development of antiviral resistance. As the host evolves far more slowly than the virus, and influenza pathogenesis is determined in part by the host response, targeting the host response is a promising approach to treating influenza. Here we characterize the host transcriptomic response to emerging H7N9 influenza virus and compare it with the responses to H7N7, H5N1, and pdm09H1N1. All three avian viruses were pathogenic in mice and elicited a transcriptomic signature that also occurs in response to the legendary 1918 influenza virus. Our work identifies host responses that could be targeted to treat severe H7N9 influenza and identifies six FDA-approved drugs that could potentially be repurposed as H7N9 influenza therapeutics. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Expanding frontiers in plant transcriptomics in aid of functional genomics and molecular breeding.

PubMed

Agarwal, Pinky; Parida, Swarup K; Mahto, Arunima; Das, Sweta; Mathew, Iny Elizebeth; Malik, Naveen; Tyagi, Akhilesh K

2014-12-01

The transcript pool of a plant part, under any given condition, is a collection of mRNAs that will pave the way for a biochemical reaction of the plant to stimuli. Over the past decades, transcriptome study has advanced from Northern blotting to RNA sequencing (RNA-seq), through other techniques, of which real-time quantitative polymerase chain reaction (PCR) and microarray are the most significant ones. The questions being addressed by such studies have also matured from a solitary process to expression atlas and marker-assisted genetic enhancement. Not only genes and their networks involved in various developmental processes of plant parts have been elucidated, but also stress tolerant genes have been highlighted. The transcriptome of a plant with altered expression of a target gene has given information about the downstream genes. Marker information has been used for breeding improved varieties. Fortunately, the data generated by transcriptome analysis has been made freely available for ample utilization and comparison. The review discusses this wide variety of transcriptome data being generated in plants, which includes developmental stages, abiotic and biotic stress, effect of altered gene expression, as well as comparative transcriptomics, with a special emphasis on microarray and RNA-seq. Such data can be used to determine the regulatory gene networks, which can subsequently be utilized for generating improved plant varieties. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

De Novo Transcriptome of the Hemimetabolous German Cockroach (Blattella germanica)

PubMed Central

Zhou, Xiaojie; Qian, Kun; Tong, Ying; Zhu, Junwei Jerry; Qiu, Xinghui; Zeng, Xiaopeng

2014-01-01

Background The German cockroach, Blattella germanica, is an important insect pest that transmits various pathogens mechanically and causes severe allergic diseases. This insect has long served as a model system for studies of insect biology, physiology and ecology. However, the lack of genome or transcriptome information heavily hinder our further understanding about the German cockroach in every aspect at a molecular level and on a genome-wide scale. To explore the transcriptome and identify unique sequences of interest, we subjected the B. germanica transcriptome to massively parallel pyrosequencing and generated the first reference transcriptome for B. germanica. Methodology/Principal Findings A total of 1,365,609 raw reads with an average length of 529 bp were generated via pyrosequencing the mixed cDNA library from different life stages of German cockroach including maturing oothecae, nymphs, adult females and males. The raw reads were de novo assembled to 48,800 contigs and 3,961 singletons with high-quality unique sequences. These sequences were annotated and classified functionally in terms of BLAST, GO and KEGG, and the genes putatively coding detoxification enzyme systems, insecticide targets, key components in systematic RNA interference, immunity and chemoreception pathways were identified. A total of 3,601 SSRs (Simple Sequence Repeats) loci were also predicted. Conclusions/Significance The whole transcriptome pyrosequencing data from this study provides a usable genetic resource for future identification of potential functional genes involved in various biological processes. PMID:25265537

Optimized approach for Ion Proton RNA sequencing reveals details of RNA splicing and editing features of the transcriptome.

PubMed

Brown, Roger B; Madrid, Nathaniel J; Suzuki, Hideaki; Ness, Scott A

2017-01-01

RNA-sequencing (RNA-seq) has become the standard method for unbiased analysis of gene expression but also provides access to more complex transcriptome features, including alternative RNA splicing, RNA editing, and even detection of fusion transcripts formed through chromosomal translocations. However, differences in library methods can adversely affect the ability to recover these different types of transcriptome data. For example, some methods have bias for one end of transcripts or rely on low-efficiency steps that limit the complexity of the resulting library, making detection of rare transcripts less likely. We tested several commonly used methods of RNA-seq library preparation and found vast differences in the detection of advanced transcriptome features, such as alternatively spliced isoforms and RNA editing sites. By comparing several different protocols available for the Ion Proton sequencer and by utilizing detailed bioinformatics analysis tools, we were able to develop an optimized random primer based RNA-seq technique that is reliable at uncovering rare transcript isoforms and RNA editing features, as well as fusion reads from oncogenic chromosome rearrangements. The combination of optimized libraries and rapid Ion Proton sequencing provides a powerful platform for the transcriptome analysis of research and clinical samples.

Tentacle Transcriptome and Venom Proteome of the Pacific Sea Nettle, Chrysaora fuscescens (Cnidaria: Scyphozoa).

PubMed

Ponce, Dalia; Brinkman, Diane L; Potriquet, Jeremy; Mulvenna, Jason

2016-04-05

Jellyfish venoms are rich sources of toxins designed to capture prey or deter predators, but they can also elicit harmful effects in humans. In this study, an integrated transcriptomic and proteomic approach was used to identify putative toxins and their potential role in the venom of the scyphozoan jellyfish Chrysaora fuscescens. A de novo tentacle transcriptome, containing more than 23,000 contigs, was constructed and used in proteomic analysis of C. fuscescens venom to identify potential toxins. From a total of 163 proteins identified in the venom proteome, 27 were classified as putative toxins and grouped into six protein families: proteinases, venom allergens, C-type lectins, pore-forming toxins, glycoside hydrolases and enzyme inhibitors. Other putative toxins identified in the transcriptome, but not the proteome, included additional proteinases as well as lipases and deoxyribonucleases. Sequence analysis also revealed the presence of ShKT domains in two putative venom proteins from the proteome and an additional 15 from the transcriptome, suggesting potential ion channel blockade or modulatory activities. Comparison of these potential toxins to those from other cnidarians provided insight into their possible roles in C. fuscescens venom and an overview of the diversity of potential toxin families in cnidarian venoms.

Parallel epigenomic and transcriptomic responses to viral infection in honey bees (Apis mellifera).

PubMed

Galbraith, David A; Yang, Xingyu; Niño, Elina Lastro; Yi, Soojin; Grozinger, Christina

2015-03-01

Populations of honey bees are declining throughout the world, with US beekeepers losing 30% of their colonies each winter. Though multiple factors are driving these colony losses, it is increasingly clear that viruses play a major role. However, information about the molecular mechanisms mediating antiviral immunity in honey bees is surprisingly limited. Here, we examined the transcriptional and epigenetic (DNA methylation) responses to viral infection in honey bee workers. One-day old worker honey bees were fed solutions containing Israeli Acute Paralysis Virus (IAPV), a virus which causes muscle paralysis and death and has previously been associated with colony loss. Uninfected control and infected, symptomatic bees were collected within 20-24 hours after infection. Worker fat bodies, the primary tissue involved in metabolism, detoxification and immune responses, were collected for analysis. We performed transcriptome- and bisulfite-sequencing of the worker fat bodies to identify genome-wide gene expression and DNA methylation patterns associated with viral infection. There were 753 differentially expressed genes (FDR<0.05) in infected versus control bees, including several genes involved in epigenetic and antiviral pathways. DNA methylation status of 156 genes (FDR<0.1) changed significantly as a result of the infection, including those involved in antiviral responses in humans. There was no significant overlap between the significantly differentially expressed and significantly differentially methylated genes, and indeed, the genomic characteristics of these sets of genes were quite distinct. Our results indicate that honey bees have two distinct molecular pathways, mediated by transcription and methylation, that modulate protein levels and/or function in response to viral infections.

Bioinformatic prediction of G protein-coupled receptor encoding sequences from the transcriptome of the foreleg, including the Haller’s organ, of the cattle tick, Rhipicephalus australis

PubMed Central

Munoz, Sergio; Guerrero, Felix D.; Kellogg, Anastasia; Heekin, Andrew M.

2017-01-01

The cattle tick of Australia, Rhipicephalus australis, is a vector for microbial parasites that cause serious bovine diseases. The Haller’s organ, located in the tick’s forelegs, is crucial for host detection and mating. To facilitate the development of new technologies for better control of this agricultural pest, we aimed to sequence and annotate the transcriptome of the R. australis forelegs and associated tissues, including the Haller's organ. As G protein-coupled receptors (GPCRs) are an important family of eukaryotic proteins studied as pharmaceutical targets in humans, we prioritized the identification and classification of the GPCRs expressed in the foreleg tissues. The two forelegs from adult R. australis were excised, RNA extracted, and pyrosequenced with 454 technology. Reads were assembled into unigenes and annotated by sequence similarity. Python scripts were written to find open reading frames (ORFs) from each unigene. These ORFs were analyzed by different GPCR prediction approaches based on sequence alignments, support vector machines, hidden Markov models, and principal component analysis. GPCRs consistently predicted by multiple methods were further studied by phylogenetic analysis and 3D homology modeling. From 4,782 assembled unigenes, 40,907 possible ORFs were predicted. Using Blastp, Pfam, GPCRpred, TMHMM, and PCA-GPCR, a basic set of 46 GPCR candidates were compiled and a phylogenetic tree was constructed. With further screening of tertiary structures predicted by RaptorX, 6 likely GPCRs emerged and the strongest candidate was classified by PCA-GPCR to be a GABAB receptor. PMID:28231302

RNA interference of three up-regulated transcripts associated with insecticide resistance in an imidacloprid resistant population of Leptinotarsa decemlineata.

PubMed

Clements, Justin; Schoville, Sean; Peterson, Nathan; Huseth, Anders S; Lan, Que; Groves, Russell L

2017-01-01

The Colorado potato beetle, Leptinotarsa decemlineata (Say), is a major agricultural pest of potatoes in the Central Sands production region of Wisconsin. Previous studies have shown that populations of L. decemlineata have become resistant to many classes of insecticides, including the neonicotinoid insecticide, imidacloprid. Furthermore, L. decemlineata has multiple mechanisms of resistance to deal with a pesticide insult, including enhanced metabolic detoxification by cytochrome p450s and glutathione S-transferases. With recent advances in the transcriptomic analysis of imidacloprid susceptible and resistant L. decemlineata populations, it is possible to investigate the role of candidate genes involved in imidacloprid resistance. A recently annotated transcriptome analysis of L. decemlineata was obtained from select populations of L. decemlineata collected in the Central Sands potato production region, which revealed a subset of mRNA transcripts constitutively up-regulated in resistant populations. We hypothesize that a portion of the up-regulated transcripts encoding for genes within the resistant populations also encode for pesticide resistance and can be suppressed to re-establish a susceptible phenotype. In this study, a discrete set of three up-regulated targets were selected for RNA interference experiments using a resistant L. decemlineata population. Following the successful suppression of transcripts encoding for a cytochrome p450, a cuticular protein, and a glutathione synthetase protein in a select L. decemlineata population, we observed reductions in measured resistance to imidacloprid that strongly suggest these genes control essential steps in imidacloprid metabolism in these field populations. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Bioinformatic prediction of G protein-coupled receptor encoding sequences from the transcriptome of the foreleg, including the Haller's organ, of the cattle tick, Rhipicephalus australis.

PubMed

Munoz, Sergio; Guerrero, Felix D; Kellogg, Anastasia; Heekin, Andrew M; Leung, Ming-Ying

2017-01-01

The cattle tick of Australia, Rhipicephalus australis, is a vector for microbial parasites that cause serious bovine diseases. The Haller's organ, located in the tick's forelegs, is crucial for host detection and mating. To facilitate the development of new technologies for better control of this agricultural pest, we aimed to sequence and annotate the transcriptome of the R. australis forelegs and associated tissues, including the Haller's organ. As G protein-coupled receptors (GPCRs) are an important family of eukaryotic proteins studied as pharmaceutical targets in humans, we prioritized the identification and classification of the GPCRs expressed in the foreleg tissues. The two forelegs from adult R. australis were excised, RNA extracted, and pyrosequenced with 454 technology. Reads were assembled into unigenes and annotated by sequence similarity. Python scripts were written to find open reading frames (ORFs) from each unigene. These ORFs were analyzed by different GPCR prediction approaches based on sequence alignments, support vector machines, hidden Markov models, and principal component analysis. GPCRs consistently predicted by multiple methods were further studied by phylogenetic analysis and 3D homology modeling. From 4,782 assembled unigenes, 40,907 possible ORFs were predicted. Using Blastp, Pfam, GPCRpred, TMHMM, and PCA-GPCR, a basic set of 46 GPCR candidates were compiled and a phylogenetic tree was constructed. With further screening of tertiary structures predicted by RaptorX, 6 likely GPCRs emerged and the strongest candidate was classified by PCA-GPCR to be a GABAB receptor.

Ambient temperature signalling in plants.

PubMed

Wigge, Philip A

2013-10-01

Plants are exposed to daily and seasonal fluctuations in temperature. Within the 'ambient' temperature range (about 12-27°C for Arabidopsis) temperature differences have large effects on plant growth and development, disease resistance pathways and the circadian clock without activating temperature stress pathways. It is this developmental sensing and response to non-stressful temperatures that will be covered in this review. Recent advances have revealed key players in mediating temperature signals. The bHLH transcription factor PHYTOCHROME INTERACTING FACTOR4 (PIF4) has been shown to be a hub for multiple responses to warmer temperature in Arabidopsis, including flowering and hypocotyl elongation. Changes in chromatin state are involved in transmitting temperature signals to the transcriptome. Determining the precise mechanisms of temperature perception represents an exciting goal for the field. Copyright © 2013 Elsevier Ltd. All rights reserved.

Hv 1 Proton Channels in Dinoflagellates: Not Just for Bioluminescence?

PubMed

Kigundu, Gabriel; Cooper, Jennifer L; Smith, Susan M E

2018-04-26

Bioluminescence in dinoflagellates is controlled by H V 1 proton channels. Database searches of dinoflagellate transcriptomes and genomes yielded hits with sequence features diagnostic of all confirmed H V 1, and show that H V 1 is widely distributed in the dinoflagellate phylogeny including the basal species Oxyrrhis marina. Multiple sequence alignments followed by phylogenetic analysis revealed three major subfamilies of H V 1 that do not correlate with presence of theca, autotrophy, geographic location, or bioluminescence. These data suggest that most dinoflagellates express a H V 1 which has a function separate from bioluminescence. Sequence evidence also suggests that dinoflagellates can contain more than one H V 1 gene. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Deep functional analysis of synII, a 770 kb synthetic yeast chromosome

PubMed Central

Gao, Feng; Gong, Jianhui; Abramczyk, Dariusz; Walker, Roy; Zhao, Hongcui; Chen, Shihong; Liu, Wei; Luo, Yisha; Müller, Carolin A.; Paul-Dubois-Taine, Adrien; Alver, Bonnie; Stracquadanio, Giovanni; Mitchell, Leslie A.; Luo, Zhouqing; Fan, Yanqun; Zhou, Baojin; Wen, Bo; Tan, Fengji; Wang, Yujia; Zi, Jin; Xie, Zexiong; Li, Bingzhi; Yang, Kun; Richardson, Sarah M.; Jiang, Hui; French, Christopher E.; Nieduszynski, Conrad A.; Koszul, Romain; Marston, Adele L.; Yuan, Yingjin; Wang, Jian; Bader, Joel S.; Dai, Junbiao; Boeke, Jef D.; Xu, Xun; Cai, Yizhi; Yang, Huanming

2017-01-01

Herein we report the successful design, construction and characterization of a 770 kb synthetic yeast chromosome II (synII). Our study incorporates characterization at multiple levels, including phenomics, transcriptomics, proteomics, chromosome segregation and replication analysis to provide a thorough and comprehensive analysis of a synthetic chromosome. Our “Trans-Omics” analyses reveal a modest but potentially significant pervasive up-regulation of translational machinery observed in synII is mainly caused by the deletion of 13 tRNAs. By both complementation assays and SCRaMbLE, we targeted and debuged the origin of a growth defect at 37°C in glycerol medium, which is related to misregulation of the HOG response. Despite the subtle differences, the synII strain shows highly consistent biological processes comparable to the native strain. PMID:28280153

Long-read sequencing of the coffee bean transcriptome reveals the diversity of full-length transcripts

PubMed Central

Cheng, Bing; Furtado, Agnelo

2017-01-01

Abstract Polyploidization contributes to the complexity of gene expression, resulting in numerous related but different transcripts. This study explored the transcriptome diversity and complexity of the tetraploid Arabica coffee (Coffea arabica) bean. Long-read sequencing (LRS) by Pacbio Isoform sequencing (Iso-seq) was used to obtain full-length transcripts without the difficulty and uncertainty of assembly required for reads from short-read technologies. The tetraploid transcriptome was annotated and compared with data from the sub-genome progenitors. Caffeine and sucrose genes were targeted for case analysis. An isoform-level tetraploid coffee bean reference transcriptome with 95 995 distinct transcripts (average 3236 bp) was obtained. A total of 88 715 sequences (92.42%) were annotated with BLASTx against NCBI non-redundant plant proteins, including 34 719 high-quality annotations. Further BLASTn analysis against NCBI non-redundant nucleotide sequences, Coffea canephora coding sequences with UTR, C. arabica ESTs, and Rfam resulted in 1213 sequences without hits, were potential novel genes in coffee. Longer UTRs were captured, especially in the 5΄UTRs, facilitating the identification of upstream open reading frames. The LRS also revealed more and longer transcript variants in key caffeine and sucrose metabolism genes from this polyploid genome. Long sequences (>10 kilo base) were poorly annotated. LRS technology shows the limitation of previous studies. It provides an important tool to produce a reference transcriptome including more of the diversity of full-length transcripts to help understand the biology and support the genetic improvement of polyploid species such as coffee. PMID:29048540

ChloroSeq, an optimized chloroplast RNA-Seq bioinformatic pipeline, reveals remodeling of the organellar transcriptome under heat stress

DOE PAGES

Castandet, Benoît; Hotto, Amber M.; Strickler, Susan R.; ...

2016-07-06

Although RNA-Seq has revolutionized transcript analysis, organellar transcriptomes are rarely assessed even when present in published datasets. Here, we describe the development and application of a rapid and convenient method, ChloroSeq, to delineate qualitative and quantitative features of chloroplast RNA metabolism from strand-specific RNA-Seq datasets, including processing, editing, splicing, and relative transcript abundance. The use of a single experiment to analyze systematically chloroplast transcript maturation and abundance is of particular interest due to frequent pleiotropic effects observed in mutants that affect chloroplast gene expression and/or photosynthesis. To illustrate its utility, ChloroSeq was applied to published RNA-Seq datasets derived from Arabidopsismore » thaliana grown under control and abiotic stress conditions, where the organellar transcriptome had not been examined. The most appreciable effects were found for heat stress, which induces a global reduction in splicing and editing efficiency, and leads to increased abundance of chloroplast transcripts, including genic, intergenic, and antisense transcripts. Moreover, by concomitantly analyzing nuclear transcripts that encode chloroplast gene expression regulators from the same libraries, we demonstrate the possibility of achieving a holistic understanding of the nucleus-organelle system. In conclusion, ChloroSeq thus represents a unique method for streamlining RNA-Seq data interpretation of the chloroplast transcriptome and its regulators.« less

Maternal Pre-Pregnancy Obesity Is Associated with Altered Placental Transcriptome.

PubMed

Altmäe, Signe; Segura, Maria Teresa; Esteban, Francisco J; Bartel, Sabine; Brandi, Pilar; Irmler, Martin; Beckers, Johannes; Demmelmair, Hans; López-Sabater, Carmen; Koletzko, Berthold; Krauss-Etschmann, Susanne; Campoy, Cristina

2017-01-01

Maternal obesity has a major impact on pregnancy outcomes. There is growing evidence that maternal obesity has a negative influence on placental development and function, thereby adversely influencing offspring programming and health outcomes. However, the molecular mechanisms underlying these processes are poorly understood. We analysed ten term placenta's whole transcriptomes in obese (n = 5) and normal weight women (n = 5), using the Affymetrix microarray platform. Analyses of expression data were carried out using non-parametric methods. Hierarchical clustering and principal component analysis showed a clear distinction in placental transcriptome between obese and normal weight women. We identified 72 differentially regulated genes, with most being down-regulated in obesity (n = 61). Functional analyses of the targets using DAVID and IPA confirm the dysregulation of previously identified processes and pathways in the placenta from obese women, including inflammation and immune responses, lipid metabolism, cancer pathways, and angiogenesis. In addition, we detected new molecular aspects of obesity-derived effects on the placenta, involving the glucocorticoid receptor signalling pathway and dysregulation of several genes including CCL2, FSTL3, IGFBP1, MMP12, PRG2, PRL, QSOX1, SERPINE2 and TAC3. Our global gene expression profiling approach demonstrates that maternal obesity creates a unique in utero environment that impairs the placental transcriptome.

ChloroSeq, an optimized chloroplast RNA-Seq bioinformatic pipeline, reveals remodeling of the organellar transcriptome under heat stress

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

Castandet, Benoît; Hotto, Amber M.; Strickler, Susan R.

Although RNA-Seq has revolutionized transcript analysis, organellar transcriptomes are rarely assessed even when present in published datasets. Here, we describe the development and application of a rapid and convenient method, ChloroSeq, to delineate qualitative and quantitative features of chloroplast RNA metabolism from strand-specific RNA-Seq datasets, including processing, editing, splicing, and relative transcript abundance. The use of a single experiment to analyze systematically chloroplast transcript maturation and abundance is of particular interest due to frequent pleiotropic effects observed in mutants that affect chloroplast gene expression and/or photosynthesis. To illustrate its utility, ChloroSeq was applied to published RNA-Seq datasets derived from Arabidopsismore » thaliana grown under control and abiotic stress conditions, where the organellar transcriptome had not been examined. The most appreciable effects were found for heat stress, which induces a global reduction in splicing and editing efficiency, and leads to increased abundance of chloroplast transcripts, including genic, intergenic, and antisense transcripts. Moreover, by concomitantly analyzing nuclear transcripts that encode chloroplast gene expression regulators from the same libraries, we demonstrate the possibility of achieving a holistic understanding of the nucleus-organelle system. In conclusion, ChloroSeq thus represents a unique method for streamlining RNA-Seq data interpretation of the chloroplast transcriptome and its regulators.« less

Systems and Trans-System Level Analysis Identifies Conserved Iron Deficiency Responses in the Plant Lineage[W][OA

PubMed Central

Urzica, Eugen I.; Casero, David; Yamasaki, Hiroaki; Hsieh, Scott I.; Adler, Lital N.; Karpowicz, Steven J.; Blaby-Haas, Crysten E.; Clarke, Steven G.; Loo, Joseph A.; Pellegrini, Matteo; Merchant, Sabeeha S.

2012-01-01

We surveyed the iron nutrition-responsive transcriptome of Chlamydomonas reinhardtii using RNA-Seq methodology. Presumed primary targets were identified in comparisons between visually asymptomatic iron-deficient versus iron-replete cells. This includes the known components of high-affinity iron uptake as well as candidates for distributive iron transport in C. reinhardtii. Comparison of growth-inhibited iron-limited versus iron-replete cells revealed changes in the expression of genes in chloroplastic oxidative stress response pathways, among hundreds of other genes. The output from the transcriptome was validated at multiple levels: by quantitative RT-PCR for assessing the data analysis pipeline, by quantitative proteomics for assessing the impact of changes in RNA abundance on the proteome, and by cross-species comparison for identifying conserved or universal response pathways. In addition, we assessed the functional importance of three target genes, VITAMIN C 2 (VTC2), MONODEHYDROASCORBATE REDUCTASE 1 (MDAR1), and CONSERVED IN THE GREEN LINEAGE AND DIATOMS 27 (CGLD27), by biochemistry or reverse genetics. VTC2 and MDAR1, which are key enzymes in de novo ascorbate synthesis and ascorbate recycling, respectively, are likely responsible for the 10-fold increase in ascorbate content of iron-limited cells. CGLD27/At5g67370 is a highly conserved, presumed chloroplast-localized pioneer protein and is important for growth of Arabidopsis thaliana in low iron. PMID:23043051

Transcriptome analyses of seed development in grape hybrids reveals a possible mechanism influencing seed size.

PubMed

Wang, Li; Hu, Xiaoyan; Jiao, Chen; Li, Zhi; Fei, Zhangjun; Yan, Xiaoxiao; Liu, Chonghuai; Wang, Yuejin; Wang, Xiping

2016-11-09

Seedlessness in grape (Vitis vinifera) is of considerable commercial importance for both the table grape and processing industries. Studies to date of grape seed development have been made certain progress, but many key genes have yet to be identified and characterized. In this study we analyzed the seed transcriptomes of progeny derived from the V. vinifera seeded maternal parent 'Red Globe' and the seedless paternal parent 'Centennial seedless' to identify genes associated with seedlessness. A total of 6,607 differentially expressed genes (DEGs) were identified and examined from multiple perspectives, including expression patterns, Gene Ontology (GO) annotations, pathway enrichment, inferred hormone influence and epigenetic regulation. The expression data of hormone-related genes and hormone level measurement reveals the differences during seed development between seedless and seeded progeny. Based on both our results and previous studies of A. thaliana seed development, we generated network maps of grape seed-related DEGs, with particular reference to hormone balance, seed coat and endosperm development, and seed identity complexes. In summary, the major differences identified during seed development of seedless and seeded progeny were associated with hormone and epigenetic regulation, the development of the seed coat and endosperm, and the formation of seed identity complexes. Overall the data provides insights into the possible molecular mechanism controlling grape seed size, which is of great importance for both basic research and future translation applications in the grape industry.

Comprehensive transcriptome analysis reveals distinct regulatory programs during vernalization and floral bud development of orchardgrass (Dactylis glomerata L.).

PubMed

Feng, Guangyan; Huang, Linkai; Li, Ji; Wang, Jianping; Xu, Lei; Pan, Ling; Zhao, Xinxin; Wang, Xia; Huang, Ting; Zhang, Xinquan

2017-11-22

Vernalization and the transition from vegetative to reproductive growth involve multiple pathways, vital for controlling floral organ formation and flowering time. However, little transcription information is available about the mechanisms behind environmental adaption and growth regulation. Here, we used high-throughput sequencing to analyze the comprehensive transcriptome of Dactylis glomerata L. during six different growth periods. During vernalization, 4689 differentially expressed genes (DEGs) significantly increased in abundance, while 3841 decreased. Furthermore, 12,967 DEGs were identified during booting stage and flowering stage, including 7750 up-regulated and 5219 down-regulated DEGs. Pathway analysis indicated that transcripts related to circadian rhythm, photoperiod, photosynthesis, flavonoid biosynthesis, starch, and sucrose metabolism changed significantly at different stages. Coexpression and weighted correlation network analysis (WGCNA) analysis linked different stages to transcriptional changes and provided evidence of inner relation modules associated with signal transduction, stress responses, cell division, and hormonal transport. We found enrichment in transcription factors (TFs) related to WRKY, NAC, AP2/EREBP, AUX/IAA, MADS-BOX, ABI3/VP1, bHLH, and the CCAAT family during vernalization and floral bud development. TFs expression patterns revealed intricate temporal variations, suggesting relatively separate regulatory programs of TF modules. Further study will unlock insights into the ability of the circadian rhythm and photoperiod to regulate vernalization and flowering time in perennial grass.

Tools for Genomic and Transcriptomic Analysis of Microbes at Single-Cell Level

PubMed Central

Chen, Zixi; Chen, Lei; Zhang, Weiwen

2017-01-01

Microbiologists traditionally study population rather than individual cells, as it is generally assumed that the status of individual cells will be similar to that observed in the population. However, the recent studies have shown that the individual behavior of each single cell could be quite different from that of the whole population, suggesting the importance of extending traditional microbiology studies to single-cell level. With recent technological advances, such as flow cytometry, next-generation sequencing (NGS), and microspectroscopy, single-cell microbiology has greatly enhanced the understanding of individuality and heterogeneity of microbes in many biological systems. Notably, the application of multiple ‘omics’ in single-cell analysis has shed light on how individual cells perceive, respond, and adapt to the environment, how heterogeneity arises under external stress and finally determines the fate of the whole population, and how microbes survive under natural conditions. As single-cell analysis involves no axenic cultivation of target microorganism, it has also been demonstrated as a valuable tool for dissecting the microbial ‘dark matter.’ In this review, current state-of-the-art tools and methods for genomic and transcriptomic analysis of microbes at single-cell level were critically summarized, including single-cell isolation methods and experimental strategies of single-cell analysis with NGS. In addition, perspectives on the future trends of technology development in the field of single-cell analysis was also presented. PMID:28979258

Comparative Transcriptomics to Identify Novel Genes and Pathways in Dinoflagellates

NASA Astrophysics Data System (ADS)

Ryan, D.

2016-02-01

The unarmored dinoflagellate Karenia brevis is among the most prominent harmful, bloom-forming phytoplankton species in the Gulf of Mexico. During blooms, the polyketides PbTx-1 and PbTx-2 (brevetoxins) are produced by K. brevis. Brevetoxins negatively impact human health and the Gulf shellfish harvest. However, the genes underlying brevetoxin synthesis are currently unknown. Because the K. brevis genome is extremely large ( 1 × 1011 base pairs long), and with a high proportion of repetitive, non-coding DNA, it has not been sequenced. In fact, large, repetitive genomes are common among the dinoflagellate group. High-throughput RNA sequencing technology enabled us to assemble Karenia transcriptomes de novo and investigate potential genes in the brevetoxin pathway through comparative transcriptomics. The brevetoxin profile varies among K. brevis clonal cultures. For example, well-documented Wilson-CCFWC268 typically produces 8-10 pg PbTx per cell, whereas SP1 produces < 2 pg PbTx/cell, and the mutant low-toxin Wilson clone produces undetectable to low (<0.05 pg/cell) amounts. Further, PbTx-2 has been measured in Karenia papilionacea but not Karenia mikimotoi. We compared the transcriptomes of four K. brevis clones (Wilson-CCFWC268, SP3, SP1, and mutant low-toxin Wilson) with K. papilionacea and K. mikimotoi to investigate nucleotide-level genetic variations and differences in gene expression. Of the 85,000 transcripts in the K. brevis transcriptome, 4,600 transcripts, including novel unannotated orthologs and putative polyketide synthases (PKSs), were only expressed by brevetoxin-producing K. brevis and K. papilionacea, not K. mikimotoi. Examination of gene expression between the typical- and low-toxin Wilson clones identified about 3,500 genes with significantly different expression levels, including 2 putative PKSs. One of the 2 PKSs was only found in the brevetoxin-producing Karenia species. These transcriptomes could not have been characterized without high-throughput RNA sequencing.

TCW: Transcriptome Computational Workbench

PubMed Central

Soderlund, Carol; Nelson, William; Willer, Mark; Gang, David R.

2013-01-01

Background The analysis of transcriptome data involves many steps and various programs, along with organization of large amounts of data and results. Without a methodical approach for storage, analysis and query, the resulting ad hoc analysis can lead to human error, loss of data and results, inefficient use of time, and lack of verifiability, repeatability, and extensibility. Methodology The Transcriptome Computational Workbench (TCW) provides Java graphical interfaces for methodical analysis for both single and comparative transcriptome data without the use of a reference genome (e.g. for non-model organisms). The singleTCW interface steps the user through importing transcript sequences (e.g. Illumina) or assembling long sequences (e.g. Sanger, 454, transcripts), annotating the sequences, and performing differential expression analysis using published statistical programs in R. The data, metadata, and results are stored in a MySQL database. The multiTCW interface builds a comparison database by importing sequence and annotation from one or more single TCW databases, executes the ESTscan program to translate the sequences into proteins, and then incorporates one or more clusterings, where the clustering options are to execute the orthoMCL program, compute transitive closure, or import clusters. Both singleTCW and multiTCW allow extensive query and display of the results, where singleTCW displays the alignment of annotation hits to transcript sequences, and multiTCW displays multiple transcript alignments with MUSCLE or pairwise alignments. The query programs can be executed on the desktop for fastest analysis, or from the web for sharing the results. Conclusion It is now affordable to buy a multi-processor machine, and easy to install Java and MySQL. By simply downloading the TCW, the user can interactively analyze, query and view their data. The TCW allows in-depth data mining of the results, which can lead to a better understanding of the transcriptome. TCW is freely available from www.agcol.arizona.edu/software/tcw. PMID:23874959

TCW: transcriptome computational workbench.

PubMed

Soderlund, Carol; Nelson, William; Willer, Mark; Gang, David R

2013-01-01

The analysis of transcriptome data involves many steps and various programs, along with organization of large amounts of data and results. Without a methodical approach for storage, analysis and query, the resulting ad hoc analysis can lead to human error, loss of data and results, inefficient use of time, and lack of verifiability, repeatability, and extensibility. The Transcriptome Computational Workbench (TCW) provides Java graphical interfaces for methodical analysis for both single and comparative transcriptome data without the use of a reference genome (e.g. for non-model organisms). The singleTCW interface steps the user through importing transcript sequences (e.g. Illumina) or assembling long sequences (e.g. Sanger, 454, transcripts), annotating the sequences, and performing differential expression analysis using published statistical programs in R. The data, metadata, and results are stored in a MySQL database. The multiTCW interface builds a comparison database by importing sequence and annotation from one or more single TCW databases, executes the ESTscan program to translate the sequences into proteins, and then incorporates one or more clusterings, where the clustering options are to execute the orthoMCL program, compute transitive closure, or import clusters. Both singleTCW and multiTCW allow extensive query and display of the results, where singleTCW displays the alignment of annotation hits to transcript sequences, and multiTCW displays multiple transcript alignments with MUSCLE or pairwise alignments. The query programs can be executed on the desktop for fastest analysis, or from the web for sharing the results. It is now affordable to buy a multi-processor machine, and easy to install Java and MySQL. By simply downloading the TCW, the user can interactively analyze, query and view their data. The TCW allows in-depth data mining of the results, which can lead to a better understanding of the transcriptome. TCW is freely available from www.agcol.arizona.edu/software/tcw.

Transcriptome analyses provide insights into the difference of alkaloids biosynthesis in the Chinese goldthread (Coptis chinensis Franch.) from different biotopes.

PubMed

Chen, Hanting; Deng, Cao; Nie, Hu; Fan, Gang; He, Yang

2017-01-01

Coptis chinensis Franch., the Chinese goldthread ('Weilian' in Chinese), one of the most important medicinal plants from the family Ranunculaceae, and its rhizome has been widely used in Traditional Chinese Medicine for centuries. Here, we analyzed the chemical components and the transcriptome of the Chinese goldthread from three biotopes, including Zhenping, Zunyi and Shizhu. We built comprehensive, high-quality de novo transcriptome assemblies of the Chinese goldthread from short-read RNA-Sequencing data, obtaining 155,710 transcripts and 56,071 unigenes. More than 98.39% and 95.97% of core eukaryotic genes were found in the transcripts and unigenes respectively, indicating that this unigene set capture the majority of the coding genes. A total of 520,462, 493,718, and 507,247 heterozygous SNPs were identified in the three accessions from Zhenping, Zunyi, and Shizhu respectively, indicating high polymorphism in coding regions of the Chinese goldthread (∼1%). Chemical analyses of the rhizome identified six major components, including berberine, palmatine, coptisine, epiberberine, columbamine, and jatrorrhizine. Berberine has the highest concentrations, followed by coptisine, palmatine, and epiberberine sequentially for all the three accessions. The drug quality of the accession from Shizhu may be the highest among these accessions. Differential analyses of the transcriptome identified four pivotal candidate enzymes, including aspartate aminotransferaseprotein, polyphenol oxidase, primary-amine oxidase, and tyrosine decarboxylase, were significantly differentially expressed and may be responsible for the difference of alkaloids contents in the accessions from different biotopes.

Transcriptome analysis of sika deer in China.

PubMed

Jia, Bo-Yin; Ba, Heng-Xing; Wang, Gui-Wu; Yang, Ying; Cui, Xue-Zhe; Peng, Ying-Hua; Zheng, Jun-Jun; Xing, Xiu-Mei; Yang, Fu-He

2016-10-01

Sika deer is of great commercial value because their antlers are used in tonics and alternative medicine and their meat is healthy and delicious. The goal of this study was to generate transcript sequences from sika deer for functional genomic analyses and to identify the transcripts that demonstrate tissue-specific, age-dependent differential expression patterns. These sequences could enhance our understanding of the molecular mechanisms underlying sika deer growth and development. In the present study, we performed de novo transcriptome assembly and profiling analysis across ten tissue types and four developmental stages (juvenile, adolescent, adult, and aged) of sika deer, using Illumina paired-end tag (PET) sequencing technology. A total of 1,752,253 contigs with an average length of 799 bp were generated, from which 1,348,618 unigenes with an average length of 590 bp were defined. Approximately 33.2 % of these (447,931 unigenes) were then annotated in public protein databases. Many sika deer tissue-specific, age-dependent unigenes were identified. The testes have the largest number of tissue-enriched unigenes, and some of them were prone to develop new functions for other tissues. Additionally, our transcriptome revealed that the juvenile-adolescent transition was the most complex and important stage of the sika deer life cycle. The present work represents the first multiple tissue transcriptome analysis of sika deer across four developmental stages. The generated data not only provide a functional genomics resource for future biological research on sika deer but also guide the selection and manipulation of genes controlling growth and development.

De novo transcriptome of the muga silkworm, Antheraea assamensis (Helfer).

PubMed

Chetia, Hasnahana; Kabiraj, Debajyoti; Singh, Deepika; Mosahari, Ponnala Vimal; Das, Suradip; Sharma, Pragya; Neog, Kartik; Sharma, Swagata; Jayaprakash, P; Bora, Utpal

2017-05-05

Antheraea assamensis (Lepidoptera: Saturniidae), is a semi-domesticated silkworm known to be endemic to Assam and the adjoining hilly areas of Northeast India. It is the only producer of a unique, commercially important variety of golden silk called "muga silk". Herein, we report the de novo transcriptome of A. assamensis reared on Machilus bombycina leaves for the first time. Short reads generated by high throughput sequencing of cDNA libraries from multiple tissues, viz. alimentary canal, silk gland and residual body of the 5 th instar of muga silkworm were assembled into transcripts via a de novo assembly pipeline followed by functional annotation and classification. A total of 1,21,433 transcripts were generated from ~231 million raw reads of which ~74% (89,583) were either allocated a functional annotation or categorized under Pfam/COG/KEGG categories. Identification of differentially expressed transcripts and their comparative sequence analysis revealed candidate genes related to silk synthesis, viz. silk gland factor-1 and 3, sericin-like transcript, etc. with conserved forkhead, homeo- and POU domains. Several candidate anti-microbial peptides which may have potential anti-bacterial, anti-fungal or anti-parasitic activity in A. assamensis were also identified. T/A and AT/TA were predicted to be the most abundant mono- and di-nucleotide simple sequence repeat markers in the transcriptome. Transcriptome validation was carried out by quantitative real-time PCR (qPCR) amplification of eight transcripts. The resources generated by this study will expand the periphery of existing genomic data on A. assamensis facilitating future in-depth studies on its unknown aspects. Copyright © 2017 Elsevier B.V. All rights reserved.

Sympatric speciation of spiny mice, Acomys, unfolded transcriptomically at Evolution Canyon, Israel

PubMed Central

Li, Kexin; Wang, Huihua; Cai, Zhenyuan; Wang, Liuyang; Xu, Qinqin; Lövy, Matěj; Wang, Zhenlong; Nevo, Eviatar

2016-01-01

Spiny mice, Acomys cahirinus, colonized Israel 30,000 y ago from dry tropical Africa and inhabited rocky habitats across Israel. Earlier, we had shown by mtDNA that A. cahirinus incipiently sympatrically speciates at Evolution Canyon I (EC I) in Mount Carmel, Israel because of microclimatic interslope divergence. The EC I microsite consists of a dry and hot savannoid “African” slope (AS) and an abutting humid and cool-forested “European” slope (ES). Here, we substantiate incipient SS in A. cahirinus at EC I based on the entire transcriptome, showing that multiple slope-specific adaptive complexes across the transcriptome result in two divergent clusters. Tajima’s D distribution of the abutting Acomys interslope populations shows that the ES population is under stronger positive selection, whereas the AS population is under balancing selection, harboring higher genetic polymorphisms. Considerable sites of the two populations were differentiated with a coefficient of FST = 0.25–0.75. Remarkably, 24 and 37 putatively adaptively selected genes were detected in the AS and ES populations, respectively. The AS genes involved DNA repair, growth arrest, neural cell differentiation, and heat-shock proteins adapting to the local AS stresses of high solar radiation, drought, and high temperature. In contrast, the ES genes involved high ATP associated with energetics stress. The sharp ecological interslope divergence led to strong slope-specific selection overruling the interslope gene flow. Earlier tests suggested slope-specific mate choice. Habitat interslope-adaptive selection across the transcriptome and mate choice substantiate sympatric speciation (SS), suggesting its prevalence at EC I and commonality in nature. PMID:27370801

Analyses of transcriptome sequences reveal multiple ancient large-scale duplication events in the ancestor of Sphagnopsida (Bryophyta)

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

Devos, Nicolas; Szövényi, Péter; Weston, David J.

In this study, the goal of this research was to investigate whether there has been a whole-genome duplication (WGD) in the ancestry of Sphagnum (peatmoss) or the class Sphagnopsida, and to determine if the timing of any such duplication(s) and patterns of paralog retention could help explain the rapid radiation and current ecological dominance of peatmosses.

Pan-cancer genome and transcriptome analyses of 1,699 paediatric leukaemias and solid tumours | Office of Cancer Genomics

Cancer.gov

Analysis of molecular aberrations across multiple cancer types, known as pan-cancer analysis, identifies commonalities and differences in key biological processes that are dysregulated in cancer cells from diverse lineages. Pan-cancer analyses have been performed for adult1–4 but not paediatric cancers, which commonly occur in developing mesodermic rather than adult epithelial tissues5.

Analyses of transcriptome sequences reveal multiple ancient large-scale duplication events in the ancestor of Sphagnopsida (Bryophyta)

DOE PAGES

Devos, Nicolas; Szövényi, Péter; Weston, David J.; ...

2016-02-22

In this study, the goal of this research was to investigate whether there has been a whole-genome duplication (WGD) in the ancestry of Sphagnum (peatmoss) or the class Sphagnopsida, and to determine if the timing of any such duplication(s) and patterns of paralog retention could help explain the rapid radiation and current ecological dominance of peatmosses.

The transcriptional landscape of age in human peripheral blood

PubMed Central

Peters, Marjolein J.; Joehanes, Roby; Pilling, Luke C.; Schurmann, Claudia; Conneely, Karen N.; Powell, Joseph; Reinmaa, Eva; Sutphin, George L.; Zhernakova, Alexandra; Schramm, Katharina; Wilson, Yana A.; Kobes, Sayuko; Tukiainen, Taru; Nalls, Michael A.; Hernandez, Dena G.; Cookson, Mark R.; Gibbs, Raphael J.; Hardy, John; Ramasamy, Adaikalavan; Zonderman, Alan B.; Dillman, Allissa; Traynor, Bryan; Smith, Colin; Longo, Dan L.; Trabzuni, Daniah; Troncoso, Juan; van der Brug, Marcel; Weale, Michael E.; O'Brien, Richard; Johnson, Robert; Walker, Robert; Zielke, Ronald H.; Arepalli, Sampath; Ryten, Mina; Singleton, Andrew B.; Ramos, Yolande F.; Göring, Harald H. H.; Fornage, Myriam; Liu, Yongmei; Gharib, Sina A.; Stranger, Barbara E.; De Jager, Philip L.; Aviv, Abraham; Levy, Daniel; Murabito, Joanne M.; Munson, Peter J.; Huan, Tianxiao; Hofman, Albert; Uitterlinden, André G.; Rivadeneira, Fernando; van Rooij, Jeroen; Stolk, Lisette; Broer, Linda; Verbiest, Michael M. P. J.; Jhamai, Mila; Arp, Pascal; Metspalu, Andres; Tserel, Liina; Milani, Lili; Samani, Nilesh J.; Peterson, Pärt; Kasela, Silva; Codd, Veryan; Peters, Annette; Ward-Caviness, Cavin K.; Herder, Christian; Waldenberger, Melanie; Roden, Michael; Singmann, Paula; Zeilinger, Sonja; Illig, Thomas; Homuth, Georg; Grabe, Hans-Jörgen; Völzke, Henry; Steil, Leif; Kocher, Thomas; Murray, Anna; Melzer, David; Yaghootkar, Hanieh; Bandinelli, Stefania; Moses, Eric K.; Kent, Jack W.; Curran, Joanne E.; Johnson, Matthew P.; Williams-Blangero, Sarah; Westra, Harm-Jan; McRae, Allan F.; Smith, Jennifer A.; Kardia, Sharon L. R.; Hovatta, Iiris; Perola, Markus; Ripatti, Samuli; Salomaa, Veikko; Henders, Anjali K.; Martin, Nicholas G.; Smith, Alicia K.; Mehta, Divya; Binder, Elisabeth B.; Nylocks, K Maria; Kennedy, Elizabeth M.; Klengel, Torsten; Ding, Jingzhong; Suchy-Dicey, Astrid M.; Enquobahrie, Daniel A.; Brody, Jennifer; Rotter, Jerome I.; Chen, Yii-Der I.; Houwing-Duistermaat, Jeanine; Kloppenburg, Margreet; Slagboom, P. Eline; Helmer, Quinta; den Hollander, Wouter; Bean, Shannon; Raj, Towfique; Bakhshi, Noman; Wang, Qiao Ping; Oyston, Lisa J.; Psaty, Bruce M.; Tracy, Russell P.; Montgomery, Grant W.; Turner, Stephen T.; Blangero, John; Meulenbelt, Ingrid; Ressler, Kerry J.; Yang, Jian; Franke, Lude; Kettunen, Johannes; Visscher, Peter M.; Neely, G. Gregory; Korstanje, Ron; Hanson, Robert L.; Prokisch, Holger; Ferrucci, Luigi; Esko, Tonu; Teumer, Alexander; van Meurs, Joyce B. J.; Johnson, Andrew D.

2015-01-01

Disease incidences increase with age, but the molecular characteristics of ageing that lead to increased disease susceptibility remain inadequately understood. Here we perform a whole-blood gene expression meta-analysis in 14,983 individuals of European ancestry (including replication) and identify 1,497 genes that are differentially expressed with chronological age. The age-associated genes do not harbor more age-associated CpG-methylation sites than other genes, but are instead enriched for the presence of potentially functional CpG-methylation sites in enhancer and insulator regions that associate with both chronological age and gene expression levels. We further used the gene expression profiles to calculate the ‘transcriptomic age' of an individual, and show that differences between transcriptomic age and chronological age are associated with biological features linked to ageing, such as blood pressure, cholesterol levels, fasting glucose, and body mass index. The transcriptomic prediction model adds biological relevance and complements existing epigenetic prediction models, and can be used by others to calculate transcriptomic age in external cohorts. PMID:26490707

Using single nuclei for RNA-seq to capture the transcriptome of postmortem neurons

PubMed Central

Krishnaswami, Suguna Rani; Grindberg, Rashel V; Novotny, Mark; Venepally, Pratap; Lacar, Benjamin; Bhutani, Kunal; Linker, Sara B; Pham, Son; Erwin, Jennifer A; Miller, Jeremy A; Hodge, Rebecca; McCarthy, James K; Kelder, Martin; McCorrison, Jamison; Aevermann, Brian D; Fuertes, Francisco Diez; Scheuermann, Richard H; Lee, Jun; Lein, Ed S; Schork, Nicholas; McConnell, Michael J; Gage, Fred H; Lasken, Roger S

2016-01-01

A protocol is described for sequencing the transcriptome of a cell nucleus. Nuclei are isolated from specimens and sorted by FACS, cDNA libraries are constructed and RNA-seq is performed, followed by data analysis. Some steps follow published methods (Smart-seq2 for cDNA synthesis and Nextera XT barcoded library preparation) and are not described in detail here. Previous single-cell approaches for RNA-seq from tissues include cell dissociation using protease treatment at 30 °C, which is known to alter the transcriptome. We isolate nuclei at 4 °C from tissue homogenates, which cause minimal damage. Nuclear transcriptomes can be obtained from postmortem human brain tissue stored at −80 °C, making brain archives accessible for RNA-seq from individual neurons. The method also allows investigation of biological features unique to nuclei, such as enrichment of certain transcripts and precursors of some noncoding RNAs. By following this procedure, it takes about 4 d to construct cDNA libraries that are ready for sequencing. PMID:26890679

Insecticide resistance is mediated by multiple mechanisms in recently introduced Aedes aegypti from Madeira Island (Portugal)

PubMed Central

Seixas, Gonçalo; Grigoraki, Linda; Weetman, David; Vicente, José Luís; Silva, Ana Clara; Pinto, João; Vontas, John

2017-01-01

Background Aedes aegypti is a major mosquito vector of arboviruses, including dengue, chikungunya and Zika. In 2005, Ae. aegypti was identified for the first time in Madeira Island. Despite an initial insecticide-based vector control program, the species expanded throughout the Southern coast of the island, suggesting the presence of insecticide resistance. Here, we characterized the insecticide resistance status and the underlying mechanisms of two populations of Ae. aegypti from Madeira Island, Funchal and Paúl do Mar. Methodology/Principal findings WHO susceptibility bioassays indicated resistance to cyfluthrin, permethrin, fenitrothion and bendiocarb. Use of synergists significantly increased mortality rates, and biochemical assays indicated elevated activities of detoxification enzymes, suggesting the importance of metabolic resistance. Microarray-based transcriptome analysis detected significant upregulation in both populations of nine cytochrome P450 oxidase genes (including four known pyrethroid metabolizing enzymes), the organophosphate metabolizer CCEae3a, Glutathione-S-transferases, and multiple putative cuticle proteins. Genotyping of knockdown resistance loci linked to pyrethroid resistance revealed fixation of the 1534C mutation, and presence with moderate frequencies of the V1016I mutation in each population. Conclusions/Significance Significant resistance to three major insecticide classes (pyrethroid, carbamate and organophosphate) is present in Ae. aegypti from Madeira Island, and appears to be mediated by multiple mechanisms. Implementation of appropriate resistance management strategies including rotation of insecticides with alternative modes of action, and methods other than chemical-based vector control are strongly advised to delay or reverse the spread of resistance and achieve efficient control. PMID:28742096

Multiple Polyploidization Events across Asteraceae with Two Nested Events in the Early History Revealed by Nuclear Phylogenomics

PubMed Central

Huang, Chien-Hsun; Zhang, Caifei; Liu, Mian; Hu, Yi; Gao, Tiangang; Qi, Ji; Ma, Hong

2016-01-01

Biodiversity results from multiple evolutionary mechanisms, including genetic variation and natural selection. Whole-genome duplications (WGDs), or polyploidizations, provide opportunities for large-scale genetic modifications. Many evolutionarily successful lineages, including angiosperms and vertebrates, are ancient polyploids, suggesting that WGDs are a driving force in evolution. However, this hypothesis is challenged by the observed lower speciation and higher extinction rates of recently formed polyploids than diploids. Asteraceae includes about 10% of angiosperm species, is thus undoubtedly one of the most successful lineages and paleopolyploidization was suggested early in this family using a small number of datasets. Here, we used genes from 64 new transcriptome datasets and others to reconstruct a robust Asteraceae phylogeny, covering 73 species from 18 tribes in six subfamilies. We estimated their divergence times and further identified multiple potential ancient WGDs within several tribes and shared by the Heliantheae alliance, core Asteraceae (Asteroideae–Mutisioideae), and also with the sister family Calyceraceae. For two of the WGD events, there were subsequent great increases in biodiversity; the older one proceeded the divergence of at least 10 subfamilies within 10 My, with great variation in morphology and physiology, whereas the other was followed by extremely high species richness in the Heliantheae alliance clade. Our results provide different evidence for several WGDs in Asteraceae and reveal distinct association among WGD events, dramatic changes in environment and species radiations, providing a possible scenario for polyploids to overcome the disadvantages of WGDs and to evolve into lineages with high biodiversity. PMID:27604225

Comparative analyses of two Geraniaceae transcriptomes using next-generation sequencing.

PubMed

Zhang, Jin; Ruhlman, Tracey A; Mower, Jeffrey P; Jansen, Robert K

2013-12-29

Organelle genomes of Geraniaceae exhibit several unusual evolutionary phenomena compared to other angiosperm families including accelerated nucleotide substitution rates, widespread gene loss, reduced RNA editing, and extensive genomic rearrangements. Since most organelle-encoded proteins function in multi-subunit complexes that also contain nuclear-encoded proteins, it is likely that the atypical organellar phenomena affect the evolution of nuclear genes encoding organellar proteins. To begin to unravel the complex co-evolutionary interplay between organellar and nuclear genomes in this family, we sequenced nuclear transcriptomes of two species, Geranium maderense and Pelargonium x hortorum. Normalized cDNA libraries of G. maderense and P. x hortorum were used for transcriptome sequencing. Five assemblers (MIRA, Newbler, SOAPdenovo, SOAPdenovo-trans [SOAPtrans], Trinity) and two next-generation technologies (454 and Illumina) were compared to determine the optimal transcriptome sequencing approach. Trinity provided the highest quality assembly of Illumina data with the deepest transcriptome coverage. An analysis to determine the amount of sequencing needed for de novo assembly revealed diminishing returns of coverage and quality with data sets larger than sixty million Illumina paired end reads for both species. The G. maderense and P. x hortorum transcriptomes contained fewer transcripts encoding the PLS subclass of PPR proteins relative to other angiosperms, consistent with reduced mitochondrial RNA editing activity in Geraniaceae. In addition, transcripts for all six plastid targeted sigma factors were identified in both transcriptomes, suggesting that one of the highly divergent rpoA-like ORFs in the P. x hortorum plastid genome is functional. The findings support the use of the Illumina platform and assemblers optimized for transcriptome assembly, such as Trinity or SOAPtrans, to generate high-quality de novo transcriptomes with broad coverage. In addition, results indicated no major improvements in breadth of coverage with data sets larger than six billion nucleotides or when sampling RNA from four tissue types rather than from a single tissue. Finally, this work demonstrates the power of cross-compartmental genomic analyses to deepen our understanding of the correlated evolution of the nuclear, plastid, and mitochondrial genomes in plants.

Comparative analyses of two Geraniaceae transcriptomes using next-generation sequencing

PubMed Central

2013-01-01

Background Organelle genomes of Geraniaceae exhibit several unusual evolutionary phenomena compared to other angiosperm families including accelerated nucleotide substitution rates, widespread gene loss, reduced RNA editing, and extensive genomic rearrangements. Since most organelle-encoded proteins function in multi-subunit complexes that also contain nuclear-encoded proteins, it is likely that the atypical organellar phenomena affect the evolution of nuclear genes encoding organellar proteins. To begin to unravel the complex co-evolutionary interplay between organellar and nuclear genomes in this family, we sequenced nuclear transcriptomes of two species, Geranium maderense and Pelargonium x hortorum. Results Normalized cDNA libraries of G. maderense and P. x hortorum were used for transcriptome sequencing. Five assemblers (MIRA, Newbler, SOAPdenovo, SOAPdenovo-trans [SOAPtrans], Trinity) and two next-generation technologies (454 and Illumina) were compared to determine the optimal transcriptome sequencing approach. Trinity provided the highest quality assembly of Illumina data with the deepest transcriptome coverage. An analysis to determine the amount of sequencing needed for de novo assembly revealed diminishing returns of coverage and quality with data sets larger than sixty million Illumina paired end reads for both species. The G. maderense and P. x hortorum transcriptomes contained fewer transcripts encoding the PLS subclass of PPR proteins relative to other angiosperms, consistent with reduced mitochondrial RNA editing activity in Geraniaceae. In addition, transcripts for all six plastid targeted sigma factors were identified in both transcriptomes, suggesting that one of the highly divergent rpoA-like ORFs in the P. x hortorum plastid genome is functional. Conclusions The findings support the use of the Illumina platform and assemblers optimized for transcriptome assembly, such as Trinity or SOAPtrans, to generate high-quality de novo transcriptomes with broad coverage. In addition, results indicated no major improvements in breadth of coverage with data sets larger than six billion nucleotides or when sampling RNA from four tissue types rather than from a single tissue. Finally, this work demonstrates the power of cross-compartmental genomic analyses to deepen our understanding of the correlated evolution of the nuclear, plastid, and mitochondrial genomes in plants. PMID:24373163

De novo transcriptome of Ischnura elegans provides insights into sensory biology, colour and vision genes.

PubMed

Chauhan, Pallavi; Hansson, Bengt; Kraaijeveld, Ken; de Knijff, Peter; Svensson, Erik I; Wellenreuther, Maren

2014-09-22

There is growing interest in odonates (damselflies and dragonflies) as model organisms in ecology and evolutionary biology but the development of genomic resources has been slow. So far only one draft genome (Ladona fulva) and one transcriptome assembly (Enallagma hageni) have been published. Odonates have some of the most advanced visual systems among insects and several species are colour polymorphic, and genomic and transcriptomic data would allow studying the genomic architecture of these interesting traits and make detailed comparative studies between related species possible. Here, we present a comprehensive de novo transcriptome assembly for the blue-tailed damselfly Ischnura elegans (Odonata: Coenagrionidae) built from short-read RNA-seq data. The transcriptome analysis in this paper provides a first step towards identifying genes and pathways underlying the visual and colour systems in this insect group. Illumina RNA sequencing performed on tissues from the head, thorax and abdomen generated 428,744,100 paired-ends reads amounting to 110 Gb of sequence data, which was assembled de novo with Trinity. A transcriptome was produced after filtering and quality checking yielding a final set of 60,232 high quality transcripts for analysis. CEGMA software identified 247 out of 248 ultra-conserved core proteins as 'complete' in the transcriptome assembly, yielding a completeness of 99.6%. BLASTX and InterProScan annotated 55% of the assembled transcripts and showed that the three tissue types differed both qualitatively and quantitatively in I. elegans. Differential expression identified 8,625 transcripts to be differentially expressed in head, thorax and abdomen. Targeted analyses of vision and colour functional pathways identified the presence of four different opsin types and three pigmentation pathways. We also identified transcripts involved in temperature sensitivity, thermoregulation and olfaction. All these traits and their associated transcripts are of considerable ecological and evolutionary interest for this and other insect orders. Our work presents a comprehensive transcriptome resource for the ancient insect order Odonata and provides insight into their biology and physiology. The transcriptomic resource can provide a foundation for future investigations into this diverse group, including the evolution of colour, vision, olfaction and thermal adaptation.

Singlet oxygen signatures are detected independent of light or chloroplasts in response to multiple stresses.

PubMed

Mor, Avishai; Koh, Eugene; Weiner, Lev; Rosenwasser, Shilo; Sibony-Benyamini, Hadas; Fluhr, Robert

2014-05-01

The production of singlet oxygen is typically associated with inefficient dissipation of photosynthetic energy or can arise from light reactions as a result of accumulation of chlorophyll precursors as observed in fluorescent (flu)-like mutants. Such photodynamic production of singlet oxygen is thought to be involved in stress signaling and programmed cell death. Here we show that transcriptomes of multiple stresses, whether from light or dark treatments, were correlated with the transcriptome of the flu mutant. A core gene set of 118 genes, common to singlet oxygen, biotic and abiotic stresses was defined and confirmed to be activated photodynamically by the photosensitizer Rose Bengal. In addition, induction of the core gene set by abiotic and biotic selected stresses was shown to occur in the dark and in nonphotosynthetic tissue. Furthermore, when subjected to various biotic and abiotic stresses in the dark, the singlet oxygen-specific probe Singlet Oxygen Sensor Green detected rapid production of singlet oxygen in the Arabidopsis (Arabidopsis thaliana) root. Subcellular localization of Singlet Oxygen Sensor Green fluorescence showed its accumulation in mitochondria, peroxisomes, and the nucleus, suggesting several compartments as the possible origins or targets for singlet oxygen. Collectively, the results show that singlet oxygen can be produced by multiple stress pathways and can emanate from compartments other than the chloroplast in a light-independent manner. The results imply that the role of singlet oxygen in plant stress regulation and response is more ubiquitous than previously thought.

Singlet Oxygen Signatures Are Detected Independent of Light or Chloroplasts in Response to Multiple Stresses1[C][W

PubMed Central

Mor, Avishai; Koh, Eugene; Weiner, Lev; Rosenwasser, Shilo; Sibony-Benyamini, Hadas; Fluhr, Robert

2014-01-01

The production of singlet oxygen is typically associated with inefficient dissipation of photosynthetic energy or can arise from light reactions as a result of accumulation of chlorophyll precursors as observed in fluorescent (flu)-like mutants. Such photodynamic production of singlet oxygen is thought to be involved in stress signaling and programmed cell death. Here we show that transcriptomes of multiple stresses, whether from light or dark treatments, were correlated with the transcriptome of the flu mutant. A core gene set of 118 genes, common to singlet oxygen, biotic and abiotic stresses was defined and confirmed to be activated photodynamically by the photosensitizer Rose Bengal. In addition, induction of the core gene set by abiotic and biotic selected stresses was shown to occur in the dark and in nonphotosynthetic tissue. Furthermore, when subjected to various biotic and abiotic stresses in the dark, the singlet oxygen-specific probe Singlet Oxygen Sensor Green detected rapid production of singlet oxygen in the Arabidopsis (Arabidopsis thaliana) root. Subcellular localization of Singlet Oxygen Sensor Green fluorescence showed its accumulation in mitochondria, peroxisomes, and the nucleus, suggesting several compartments as the possible origins or targets for singlet oxygen. Collectively, the results show that singlet oxygen can be produced by multiple stress pathways and can emanate from compartments other than the chloroplast in a light-independent manner. The results imply that the role of singlet oxygen in plant stress regulation and response is more ubiquitous than previously thought. PMID:24599491

A regulation probability model-based meta-analysis of multiple transcriptomics data sets for cancer biomarker identification.

PubMed

Xie, Xin-Ping; Xie, Yu-Feng; Wang, Hong-Qiang

2017-08-23

Large-scale accumulation of omics data poses a pressing challenge of integrative analysis of multiple data sets in bioinformatics. An open question of such integrative analysis is how to pinpoint consistent but subtle gene activity patterns across studies. Study heterogeneity needs to be addressed carefully for this goal. This paper proposes a regulation probability model-based meta-analysis, jGRP, for identifying differentially expressed genes (DEGs). The method integrates multiple transcriptomics data sets in a gene regulatory space instead of in a gene expression space, which makes it easy to capture and manage data heterogeneity across studies from different laboratories or platforms. Specifically, we transform gene expression profiles into a united gene regulation profile across studies by mathematically defining two gene regulation events between two conditions and estimating their occurring probabilities in a sample. Finally, a novel differential expression statistic is established based on the gene regulation profiles, realizing accurate and flexible identification of DEGs in gene regulation space. We evaluated the proposed method on simulation data and real-world cancer datasets and showed the effectiveness and efficiency of jGRP in identifying DEGs identification in the context of meta-analysis. Data heterogeneity largely influences the performance of meta-analysis of DEGs identification. Existing different meta-analysis methods were revealed to exhibit very different degrees of sensitivity to study heterogeneity. The proposed method, jGRP, can be a standalone tool due to its united framework and controllable way to deal with study heterogeneity.

Transcriptomics of cortical gray matter thickness decline during normal aging

PubMed Central

Kochunov, P; Charlesworth, J; Winkler, A; Hong, LE; Nichols, T; Curran, JE; Sprooten, E; Jahanshad, N; Thompson, PM; Johnson, MP; Kent, JW; Landman, BA; Mitchell, B; Cole, SA; Dyer, TD; Moses, EK; Goring, HHH; Almasy, L; Duggirala, R; Olvera, RL; Glahn, DC; Blangero, J

2013-01-01

Introduction We performed a whole-transcriptome correlation analysis, followed by the pathway enrichment and testing of innate immune response pathways analyses to evaluate the hypothesis that transcriptional activity can predict cortical gray matter thickness (GMT) variability during normal cerebral aging Methods Transcriptome and GMT data were availabe for 379 individuals (age range=28–85) community-dwelling members of large extended Mexican-American families. Collection of transcriptome data preceded that of neuroimaging data by 17 years. Genome-wide gene transcriptome data consisted of 20,413 heritable lymphocytes-based transcripts. GMT measurements were performed from high-resolution (isotropic 800µm) T1-weighted MRI. Transcriptome-wide and pathway enrichment analysis was used to classify genes correlated with GMT. Transcripts for sixty genes from seven innate immune pathways were tested as specific predictors of GMT variability. Results Transcripts for eight genes (IGFBP3, LRRN3, CRIP2, SCD, IDS, TCF4, GATA3, HN1) passed the transcriptome-wide significance threshold. Four orthogonal factors extracted from this set predicted 31.9% of the variability in the whole-brain and between 23.4 and 35% of regional GMT measurements. Pathway enrichment analysis identified six functional categories including cellular proliferation, aggregation, differentiation, viral infection, and metabolism. The integrin signaling pathway was significantly (p<10−6) enriched with GMT. Finally, three innate immune pathways (complement signaling, toll-receptors and scavenger and immunoglobulins) were significantly associated with GMT. Conclusion Expression activity for the genes that regulate cellular proliferation, adhesion, differentiation and inflammation can explain a significant proportion of individual variability in cortical GMT. Our findings suggest that normal cerebral aging is the product of a progressive decline in regenerative capacity and increased neuroinflammation. PMID:23707588

Transcriptomics of cortical gray matter thickness decline during normal aging.

PubMed

Kochunov, P; Charlesworth, J; Winkler, A; Hong, L E; Nichols, T E; Curran, J E; Sprooten, E; Jahanshad, N; Thompson, P M; Johnson, M P; Kent, J W; Landman, B A; Mitchell, B; Cole, S A; Dyer, T D; Moses, E K; Goring, H H H; Almasy, L; Duggirala, R; Olvera, R L; Glahn, D C; Blangero, J

2013-11-15

We performed a whole-transcriptome correlation analysis, followed by the pathway enrichment and testing of innate immune response pathway analyses to evaluate the hypothesis that transcriptional activity can predict cortical gray matter thickness (GMT) variability during normal cerebral aging. Transcriptome and GMT data were available for 379 individuals (age range=28-85) community-dwelling members of large extended Mexican American families. Collection of transcriptome data preceded that of neuroimaging data by 17 years. Genome-wide gene transcriptome data consisted of 20,413 heritable lymphocytes-based transcripts. GMT measurements were performed from high-resolution (isotropic 800 μm) T1-weighted MRI. Transcriptome-wide and pathway enrichment analysis was used to classify genes correlated with GMT. Transcripts for sixty genes from seven innate immune pathways were tested as specific predictors of GMT variability. Transcripts for eight genes (IGFBP3, LRRN3, CRIP2, SCD, IDS, TCF4, GATA3, and HN1) passed the transcriptome-wide significance threshold. Four orthogonal factors extracted from this set predicted 31.9% of the variability in the whole-brain and between 23.4 and 35% of regional GMT measurements. Pathway enrichment analysis identified six functional categories including cellular proliferation, aggregation, differentiation, viral infection, and metabolism. The integrin signaling pathway was significantly (p<10(-6)) enriched with GMT. Finally, three innate immune pathways (complement signaling, toll-receptors and scavenger and immunoglobulins) were significantly associated with GMT. Expression activity for the genes that regulate cellular proliferation, adhesion, differentiation and inflammation can explain a significant proportion of individual variability in cortical GMT. Our findings suggest that normal cerebral aging is the product of a progressive decline in regenerative capacity and increased neuroinflammation. Copyright © 2013 Elsevier Inc. All rights reserved.

GigaTON: an extensive publicly searchable database providing a new reference transcriptome in the pacific oyster Crassostrea gigas.

PubMed

Riviere, Guillaume; Klopp, Christophe; Ibouniyamine, Nabihoudine; Huvet, Arnaud; Boudry, Pierre; Favrel, Pascal

2015-12-02

The Pacific oyster, Crassostrea gigas, is one of the most important aquaculture shellfish resources worldwide. Important efforts have been undertaken towards a better knowledge of its genome and transcriptome, which makes now C. gigas becoming a model organism among lophotrochozoans, the under-described sister clade of ecdysozoans within protostomes. These massive sequencing efforts offer the opportunity to assemble gene expression data and make such resource accessible and exploitable for the scientific community. Therefore, we undertook this assembly into an up-to-date publicly available transcriptome database: the GigaTON (Gigas TranscriptOme pipeliNe) database. We assembled 2204 million sequences obtained from 114 publicly available RNA-seq libraries that were realized using all embryo-larval development stages, adult organs, different environmental stressors including heavy metals, temperature, salinity and exposure to air, which were mostly performed as part of the Crassostrea gigas genome project. This data was analyzed in silico and resulted into 56621 newly assembled contigs that were deposited into a publicly available database, the GigaTON database. This database also provides powerful and user-friendly request tools to browse and retrieve information about annotation, expression level, UTRs, splice and polymorphism, and gene ontology associated to all the contigs into each, and between all libraries. The GigaTON database provides a convenient, potent and versatile interface to browse, retrieve, confront and compare massive transcriptomic information in an extensive range of conditions, tissues and developmental stages in Crassostrea gigas. To our knowledge, the GigaTON database constitutes the most extensive transcriptomic database to date in marine invertebrates, thereby a new reference transcriptome in the oyster, a highly valuable resource to physiologists and evolutionary biologists.

Exploring the host transcriptome for mechanisms underlying protective immunity and resistance to nematode infections in ruminants.

PubMed

Li, Robert W; Choudhary, Ratan K; Capuco, Anthony V; Urban, Joseph F

2012-11-23

Nematode infections in ruminants are a major impediment to the profitable production of meat and dairy products, especially for small farms. Gastrointestinal parasitism not only negatively impacts weight gain and milk yield, but is also a major cause of mortality in small ruminants. The current parasite control strategy involves heavy use of anthelmintics that has resulted in the emergence of drug-resistant parasite strains. This, in addition to increasing consumer demand for animal products that are free of drug residues has stimulated development of alternative strategies, including selective breeding of parasite resistant ruminants. The development of protective immunity and manifestations of resistance to nematode infections relies upon the precise expression of the host genome that is often confounded by mechanisms simultaneously required to control multiple nematode species as well as ecto- and protozoan parasites, and microbial and viral pathogens. Understanding the molecular mechanisms underlying these processes represents a key step toward development of effective new parasite control strategies. Recent progress in characterizing the transcriptome of both hosts and parasites, utilizing high-throughput microarrays and RNA-seq technology, has led to the recognition of unique interactions and the identification of genes and biological pathways involved in the response to parasitism. Innovative use of the knowledge gained by these technologies should provide a basis for enhancing innate immunity while limiting the polarization of acquired immunity can negatively affect optimal responses to co-infection. Strategies for parasite control that use diet and vaccine/adjuvant combination could be evaluated by monitoring the host transcriptome for induction of appropriate mechanisms for imparting parasite resistance. Knowledge of different mechanisms of host immunity and the critical regulation of parasite development, physiology, and virulence can also selectively identify targets for parasite control. Comparative transcriptome analysis, in concert with genome-wide association (GWS) studies to identify quantitative trait loci (QTLs) affecting host resistance, represents a promising molecular technology to evaluate integrated control strategies that involve breed and environmental factors that contribute to parasite resistance and improved performance. Tailoring these factors to control parasitism without severely affecting production qualities, management efficiencies, and responses to pathogenic co-infection will remain a challenge. This review summarizes recent progress and limitations of understanding regulatory genetic networks and biological pathways that affect host resistance and susceptibility to nematode infection in ruminants. Published by Elsevier B.V.

Comparison of next generation sequencing technologies for transcriptome characterization

PubMed Central

2009-01-01

Background We have developed a simulation approach to help determine the optimal mixture of sequencing methods for most complete and cost effective transcriptome sequencing. We compared simulation results for traditional capillary sequencing with "Next Generation" (NG) ultra high-throughput technologies. The simulation model was parameterized using mappings of 130,000 cDNA sequence reads to the Arabidopsis genome (NCBI Accession SRA008180.19). We also generated 454-GS20 sequences and de novo assemblies for the basal eudicot California poppy (Eschscholzia californica) and the magnoliid avocado (Persea americana) using a variety of methods for cDNA synthesis. Results The Arabidopsis reads tagged more than 15,000 genes, including new splice variants and extended UTR regions. Of the total 134,791 reads (13.8 MB), 119,518 (88.7%) mapped exactly to known exons, while 1,117 (0.8%) mapped to introns, 11,524 (8.6%) spanned annotated intron/exon boundaries, and 3,066 (2.3%) extended beyond the end of annotated UTRs. Sequence-based inference of relative gene expression levels correlated significantly with microarray data. As expected, NG sequencing of normalized libraries tagged more genes than non-normalized libraries, although non-normalized libraries yielded more full-length cDNA sequences. The Arabidopsis data were used to simulate additional rounds of NG and traditional EST sequencing, and various combinations of each. Our simulations suggest a combination of FLX and Solexa sequencing for optimal transcriptome coverage at modest cost. We have also developed ESTcalc http://fgp.huck.psu.edu/NG_Sims/ngsim.pl, an online webtool, which allows users to explore the results of this study by specifying individualized costs and sequencing characteristics. Conclusion NG sequencing technologies are a highly flexible set of platforms that can be scaled to suit different project goals. In terms of sequence coverage alone, the NG sequencing is a dramatic advance over capillary-based sequencing, but NG sequencing also presents significant challenges in assembly and sequence accuracy due to short read lengths, method-specific sequencing errors, and the absence of physical clones. These problems may be overcome by hybrid sequencing strategies using a mixture of sequencing methodologies, by new assemblers, and by sequencing more deeply. Sequencing and microarray outcomes from multiple experiments suggest that our simulator will be useful for guiding NG transcriptome sequencing projects in a wide range of organisms. PMID:19646272

Polyphenism in social insects: insights from a transcriptome-wide analysis of gene expression in the life stages of the key pollinator, Bombus terrestris

PubMed Central

2011-01-01

Background Understanding polyphenism, the ability of a single genome to express multiple morphologically and behaviourally distinct phenotypes, is an important goal for evolutionary and developmental biology. Polyphenism has been key to the evolution of the Hymenoptera, and particularly the social Hymenoptera where the genome of a single species regulates distinct larval stages, sexual dimorphism and physical castes within the female sex. Transcriptomic analyses of social Hymenoptera will therefore provide unique insights into how changes in gene expression underlie such complexity. Here we describe gene expression in individual specimens of the pre-adult stages, sexes and castes of the key pollinator, the buff-tailed bumblebee Bombus terrestris. Results cDNA was prepared from mRNA from five life cycle stages (one larva, one pupa, one male, one gyne and two workers) and a total of 1,610,742 expressed sequence tags (ESTs) were generated using Roche 454 technology, substantially increasing the sequence data available for this important species. Overlapping ESTs were assembled into 36,354 B. terrestris putative transcripts, and functionally annotated. A preliminary assessment of differences in gene expression across non-replicated specimens from the pre-adult stages, castes and sexes was performed using R-STAT analysis. Individual samples from the life cycle stages of the bumblebee differed in the expression of a wide array of genes, including genes involved in amino acid storage, metabolism, immunity and olfaction. Conclusions Detailed analyses of immune and olfaction gene expression across phenotypes demonstrated how transcriptomic analyses can inform our understanding of processes central to the biology of B. terrestris and the social Hymenoptera in general. For example, examination of immunity-related genes identified high conservation of important immunity pathway components across individual specimens from the life cycle stages while olfactory-related genes exhibited differential expression with a wider repertoire of gene expression within adults, especially sexuals, in comparison to immature stages. As there is an absence of replication across the samples, the results of this study are preliminary but provide a number of candidate genes which may be related to distinct phenotypic stage expression. This comprehensive transcriptome catalogue will provide an important gene discovery resource for directed programmes in ecology, evolution and conservation of a key pollinator. PMID:22185240

Comparative high-throughput transcriptome sequencing and development of SiESTa, the Silene EST annotation database

PubMed Central

2011-01-01

Background The genus Silene is widely used as a model system for addressing ecological and evolutionary questions in plants, but advances in using the genus as a model system are impeded by the lack of available resources for studying its genome. Massively parallel sequencing cDNA has recently developed into an efficient method for characterizing the transcriptomes of non-model organisms, generating massive amounts of data that enable the study of multiple species in a comparative framework. The sequences generated provide an excellent resource for identifying expressed genes, characterizing functional variation and developing molecular markers, thereby laying the foundations for future studies on gene sequence and gene expression divergence. Here, we report the results of a comparative transcriptome sequencing study of eight individuals representing four Silene and one Dianthus species as outgroup. All sequences and annotations have been deposited in a newly developed and publicly available database called SiESTa, the Silene EST annotation database. Results A total of 1,041,122 EST reads were generated in two runs on a Roche GS-FLX 454 pyrosequencing platform. EST reads were analyzed separately for all eight individuals sequenced and were assembled into contigs using TGICL. These were annotated with results from BLASTX searches and Gene Ontology (GO) terms, and thousands of single-nucleotide polymorphisms (SNPs) were characterized. Unassembled reads were kept as singletons and together with the contigs contributed to the unigenes characterized in each individual. The high quality of unigenes is evidenced by the proportion (49%) that have significant hits in similarity searches with the A. thaliana proteome. The SiESTa database is accessible at http://www.siesta.ethz.ch. Conclusion The sequence collections established in the present study provide an important genomic resource for four Silene and one Dianthus species and will help to further develop Silene as a plant model system. The genes characterized will be useful for future research not only in the species included in the present study, but also in related species for which no genomic resources are yet available. Our results demonstrate the efficiency of massively parallel transcriptome sequencing in a comparative framework as an approach for developing genomic resources in diverse groups of non-model organisms. PMID:21791039

Sex and tissue specific gene expression patterns identified following de novo transcriptomic analysis of the Norway lobster, Nephrops norvegicus.

PubMed

Rotllant, Guiomar; Nguyen, Tuan Viet; Sbragaglia, Valerio; Rahi, Lifat; Dudley, Kevin J; Hurwood, David; Ventura, Tomer; Company, Joan B; Chand, Vincent; Aguzzi, Jacopo; Mather, Peter B

2017-08-16

The Norway lobster, Nephrops norvegicus, is economically important in European fisheries and is a key organism in local marine ecosystems. Despite multi-faceted scientific interest in this species, our current knowledge of genetic resources in this species remains very limited. Here, we generated a reference de novo transcriptome for N. norvegicus from multiple tissues in both sexes. Bioinformatic analyses were conducted to detect transcripts that were expressed exclusively in either males or females. Patterns were validated via RT-PCR. Sixteen N. norvegicus libraries were sequenced from immature and mature ovary, testis and vas deferens (including the masculinizing androgenic gland). In addition, eyestalk, brain, thoracic ganglia and hepatopancreas tissues were screened in males and both immature and mature females. RNA-Sequencing resulted in >600 million reads. De novo assembly that combined the current dataset with two previously published libraries from eyestalk tissue, yielded a reference transcriptome of 333,225 transcripts with an average size of 708 base pairs (bp), with an N50 of 1272 bp. Sex-specific transcripts were detected primarily in gonads followed by hepatopancreas, brain, thoracic ganglia, and eyestalk, respectively. Candidate transcripts that were expressed exclusively either in males or females were highlighted and the 10 most abundant ones were validated via RT-PCR. Among the most highly expressed genes were Serine threonine protein kinase in testis and Vitellogenin in female hepatopancreas. These results align closely with gene annotation results. Moreover, a differential expression heatmap showed that the majority of differentially expressed transcripts were identified in gonad and eyestalk tissues. Results indicate that sex-specific gene expression patterns in Norway lobster are controlled by differences in gene regulation pattern between males and females in somatic tissues. The current study presents the first multi-tissue reference transcriptome for the Norway lobster that can be applied to future biological, wild restocking and fisheries studies. Sex-specific markers were mainly expressed in males implying that males may experience stronger selection than females. It is apparent that differential expression is due to sex-specific gene regulatory pathways that are present in somatic tissues and not from effects of genes located on heterogametic sex chromosomes. The N. norvegicus data provide a foundation for future gene-based reproductive studies.

A Transcriptomic Network Underlies Microstructural and Physiological Responses to Cadmium in Populus × canescens1[C][W

PubMed Central

He, Jiali; Li, Hong; Luo, Jie; Ma, Chaofeng; Li, Shaojun; Qu, Long; Gai, Ying; Jiang, Xiangning; Janz, Dennis; Polle, Andrea; Tyree, Melvin; Luo, Zhi-Bin

2013-01-01

Bark tissue of Populus × canescens can hyperaccumulate cadmium, but microstructural, transcriptomic, and physiological response mechanisms are poorly understood. Histochemical assays, transmission electron microscopic observations, energy-dispersive x-ray microanalysis, and transcriptomic and physiological analyses have been performed to enhance our understanding of cadmium accumulation and detoxification in P. × canescens. Cadmium was allocated to the phloem of the bark, and subcellular cadmium compartmentalization occurred mainly in vacuoles of phloem cells. Transcripts involved in microstructural alteration, changes in nutrition and primary metabolism, and stimulation of stress responses showed significantly differential expression in the bark of P. × canescens exposed to cadmium. About 48% of the differentially regulated transcripts formed a coregulation network in which 43 hub genes played a central role both in cross talk among distinct biological processes and in coordinating the transcriptomic regulation in the bark of P. × canescens in response to cadmium. The cadmium transcriptome in the bark of P. × canescens was mirrored by physiological readouts. Cadmium accumulation led to decreased total nitrogen, phosphorus, and calcium and increased sulfur in the bark. Cadmium inhibited photosynthesis, resulting in decreased carbohydrate levels. Cadmium induced oxidative stress and antioxidants, including free proline, soluble phenolics, ascorbate, and thiol compounds. These results suggest that orchestrated microstructural, transcriptomic, and physiological regulation may sustain cadmium hyperaccumulation in P. × canescens bark and provide new insights into engineering woody plants for phytoremediation. PMID:23530184

Deep sequencing reveals cell-type-specific patterns of single-cell transcriptome variation.

PubMed

Dueck, Hannah; Khaladkar, Mugdha; Kim, Tae Kyung; Spaethling, Jennifer M; Francis, Chantal; Suresh, Sangita; Fisher, Stephen A; Seale, Patrick; Beck, Sheryl G; Bartfai, Tamas; Kuhn, Bernhard; Eberwine, James; Kim, Junhyong

2015-06-09

Differentiation of metazoan cells requires execution of different gene expression programs but recent single-cell transcriptome profiling has revealed considerable variation within cells of seeming identical phenotype. This brings into question the relationship between transcriptome states and cell phenotypes. Additionally, single-cell transcriptomics presents unique analysis challenges that need to be addressed to answer this question. We present high quality deep read-depth single-cell RNA sequencing for 91 cells from five mouse tissues and 18 cells from two rat tissues, along with 30 control samples of bulk RNA diluted to single-cell levels. We find that transcriptomes differ globally across tissues with regard to the number of genes expressed, the average expression patterns, and within-cell-type variation patterns. We develop methods to filter genes for reliable quantification and to calibrate biological variation. All cell types include genes with high variability in expression, in a tissue-specific manner. We also find evidence that single-cell variability of neuronal genes in mice is correlated with that in rats consistent with the hypothesis that levels of variation may be conserved. Single-cell RNA-sequencing data provide a unique view of transcriptome function; however, careful analysis is required in order to use single-cell RNA-sequencing measurements for this purpose. Technical variation must be considered in single-cell RNA-sequencing studies of expression variation. For a subset of genes, biological variability within each cell type appears to be regulated in order to perform dynamic functions, rather than solely molecular noise.

Human and feline adipose-derived mesenchymal stem cells have comparable phenotype, immunomodulatory functions, and transcriptome.

PubMed

Clark, Kaitlin C; Fierro, Fernando A; Ko, Emily Mills; Walker, Naomi J; Arzi, Boaz; Tepper, Clifford G; Dahlenburg, Heather; Cicchetto, Andrew; Kol, Amir; Marsh, Lyndsey; Murphy, William J; Fazel, Nasim; Borjesson, Dori L

2017-03-20

Adipose-derived mesenchymal stem cells (ASCs) are a promising cell therapy to treat inflammatory and immune-mediated diseases. Development of appropriate pre-clinical animal models is critical to determine safety and attain early efficacy data for the most promising therapeutic candidates. Naturally occurring diseases in cats already serve as valuable models to inform human clinical trials in oncologic, cardiovascular, and genetic diseases. The objective of this study was to complete a comprehensive side-by-side comparison of human and feline ASCs, with an emphasis on their immunomodulatory capacity and transcriptome. Human and feline ASCs were evaluated for phenotype, immunomodulatory profile, and transcriptome. Additionally, transwells were used to determine the role of cell-cell contact in ASC-mediated inhibition of lymphocyte proliferation in both humans and cats. Similar to human ASCs, feline ASCs were highly proliferative at low passages and fit the minimal criteria of multipotent stem cells including a compatible surface protein phenotype, osteogenic capacity, and normal karyotype. Like ASCs from all species, feline ASCs inhibited mitogen-activated lymphocyte proliferation in vitro, with or without direct ASC-lymphocyte contact. Feline ASCs mimic human ASCs in their mediator secretion pattern, including prostaglandin E2, indoleamine 2,3 dioxygenase, transforming growth factor beta, and interleukin-6, all augmented by interferon gamma secretion by lymphocytes. The transcriptome of three unactivated feline ASC lines were highly similar. Functional analysis of the most highly expressed genes highlighted processes including: 1) the regulation of apoptosis; 2) cell adhesion; 3) response to oxidative stress; and 4) regulation of cell differentiation. Finally, feline ASCs had a similar gene expression profile to noninduced human ASCs. Findings suggest that feline ASCs modulate lymphocyte proliferation using soluble mediators that mirror the human ASC secretion pattern. Uninduced feline ASCs have similar gene expression profiles to uninduced human ASCs, as revealed by transcriptome analysis. These data will help inform clinical trials using cats with naturally occurring diseases as surrogate models for human clinical trials in the regenerative medicine arena.

Transcriptomic and epigenomic characterization of the developing bat wing

PubMed Central

Eckalbar, Walter L.; Schlebusch, Stephen A.; Mason, Mandy K.; Gill, Zoe; Parker, Ash V.; Booker, Betty M.; Nishizaki, Sierra; Muswamba-Nday, Christiane; Terhune, Elizabeth; Nevonen, Kimberly; Makki, Nadja; Friedrich, Tara; VanderMeer, Julia E.; Pollard, Katherine S.; Carbone, Lucia; Wall, Jeff D.; Illing, Nicola; Ahituv, Nadav

2016-01-01

Bats are the only mammals capable of powered flight, but little is known about the genetic determinants that shape their wings. Here, we generated a genome for Miniopterus natalensis and performed RNA-seq and ChIP-seq (H3K27ac, H3K27me3) on its developing forelimb and hindlimb autopods at sequential embryonic stages to decipher the molecular events that underlie bat wing development. Over 7,000 genes and several lncRNAs, including Tbx5-as1 and Hottip, were differentially expressed between forelimb, hindlimb and different stages. ChIP-seq identified thousands of regions that are differentially modified in forelimb versus hindlimb. Comparative genomics found 2,796 bat-accelerated regions within H3K27ac peaks, several of which cluster near limb-associated genes. Pathway analyses revealed multiple ribosomal proteins and known limb patterning signaling pathways as differentially regulated, and implicated increased forelimb mesenchymal condensations with differential growth. Combined, our work outlines multiple genetic components that contribute to bat wing formation, providing a genomic blueprint for this morphological innovation. PMID:27019111

Survival, gene and metabolite responses of Litoria verreauxii alpina frogs to fungal disease chytridiomycosis

NASA Astrophysics Data System (ADS)

Grogan, Laura F.; Mulvenna, Jason; Gummer, Joel P. A.; Scheele, Ben C.; Berger, Lee; Cashins, Scott D.; McFadden, Michael S.; Harlow, Peter; Hunter, David A.; Trengove, Robert D.; Skerratt, Lee F.

2018-03-01

The fungal skin disease chytridiomycosis has caused the devastating decline and extinction of hundreds of amphibian species globally, yet the potential for evolving resistance, and the underlying pathophysiological mechanisms remain poorly understood. We exposed 406 naïve, captive-raised alpine tree frogs (Litoria verreauxii alpina) from multiple populations (one evolutionarily naïve to chytridiomycosis) to the aetiological agent Batrachochytrium dendrobatidis in two concurrent and controlled infection experiments. We investigated (A) survival outcomes and clinical pathogen burdens between populations and clutches, and (B) individual host tissue responses to chytridiomycosis. Here we present multiple interrelated datasets associated with these exposure experiments, including animal signalment, survival and pathogen burden of 355 animals from Experiment A, and the following datasets related to 61 animals from Experiment B: animal signalment and pathogen burden; raw RNA-Seq reads from skin, liver and spleen tissues; de novo assembled transcriptomes for each tissue type; raw gene expression data; annotation data for each gene; and raw metabolite expression data from skin and liver tissues. These data provide an extensive baseline for future analyses.

Single-cell transcriptome of early embryos and cultured embryonic stem cells of cynomolgus monkeys

PubMed Central

Nakamura, Tomonori; Yabuta, Yukihiro; Okamoto, Ikuhiro; Sasaki, Kotaro; Iwatani, Chizuru; Tsuchiya, Hideaki; Saitou, Mitinori

2017-01-01

In mammals, the development of pluripotency and specification of primordial germ cells (PGCs) have been studied predominantly using mice as a model organism. However, divergences among mammalian species for such processes have begun to be recognized. Between humans and mice, pre-implantation development appears relatively similar, but the manner and morphology of post-implantation development are significantly different. Nevertheless, the embryogenesis just after implantation in primates, including the specification of PGCs, has been unexplored due to the difficulties in analyzing the embryos at relevant developmental stages. Here, we present a comprehensive single-cell transcriptome dataset of pre- and early post-implantation embryo cells, PGCs and embryonic stem cells (ESCs) of cynomolgus monkeys as a model of higher primates. The identities of each transcriptome were also validated rigorously by other way such as immunofluorescent analysis. The information reported here will serve as a foundation for our understanding of a wide range of processes in the developmental biology of primates, including humans. PMID:28649393

Enabling large-scale next-generation sequence assembly with Blacklight

PubMed Central

Couger, M. Brian; Pipes, Lenore; Squina, Fabio; Prade, Rolf; Siepel, Adam; Palermo, Robert; Katze, Michael G.; Mason, Christopher E.; Blood, Philip D.

2014-01-01

Summary A variety of extremely challenging biological sequence analyses were conducted on the XSEDE large shared memory resource Blacklight, using current bioinformatics tools and encompassing a wide range of scientific applications. These include genomic sequence assembly, very large metagenomic sequence assembly, transcriptome assembly, and sequencing error correction. The data sets used in these analyses included uncategorized fungal species, reference microbial data, very large soil and human gut microbiome sequence data, and primate transcriptomes, composed of both short-read and long-read sequence data. A new parallel command execution program was developed on the Blacklight resource to handle some of these analyses. These results, initially reported previously at XSEDE13 and expanded here, represent significant advances for their respective scientific communities. The breadth and depth of the results achieved demonstrate the ease of use, versatility, and unique capabilities of the Blacklight XSEDE resource for scientific analysis of genomic and transcriptomic sequence data, and the power of these resources, together with XSEDE support, in meeting the most challenging scientific problems. PMID:25294974

Transcriptomic Impacts of Rumen Epithelium Induced by Butyrate Infusion in Dairy Cattle in Dry Period

PubMed Central

Baldwin, Ransom L; Li, Robert W; Jia, Yankai; Li, Cong-Jun

2018-01-01

The purpose of this study was to evaluate the effects of butyrate infusion on rumen epithelial transcriptome. Next-generation sequencing (NGS) and bioinformatics are used to accelerate our understanding of regulation in rumen epithelial transcriptome of cattle in the dry period induced by butyrate infusion at the level of the whole transcriptome. Butyrate, as an essential element of nutrients, is a histone deacetylase (HDAC) inhibitor that can alter histone acetylation and methylation, and plays a prominent role in regulating genomic activities influencing rumen nutrition utilization and function. Ruminal infusion of butyrate was following 0-hour sampling (baseline controls) and continued for 168 hours at a rate of 5.0 L/day of a 2.5 M solution as a continuous infusion. Following the 168-hour infusion, the infusion was stopped, and cows were maintained on the basal lactation ration for an additional 168 hours for sampling. Rumen epithelial samples were serially collected via biopsy through rumen fistulae at 0-, 24-, 72-, and 168-hour (D1, D3, D7) and 168-hour post-infusion (D14). In comparison with pre-infusion at 0 hours, a total of 3513 genes were identified to be impacted in the rumen epithelium by butyrate infusion at least once at different sampling time points at a stringent cutoff of false discovery rate (FDR) < 0.01. The maximal effect of butyrate was observed at day 7. Among these impacted genes, 117 genes were responsive consistently from day 1 to day 14, and another 42 genes were lasting through day 7. Temporal effects induced by butyrate infusion indicate that the transcriptomic alterations are very dynamic. Gene ontology (GO) enrichment analysis revealed that in the early stage of rumen butyrate infusion (on day 1 and day 3 of butyrate infusion), the transcriptomic effects in the rumen epithelium were involved with mitotic cell cycle process, cell cycle process, and regulation of cell cycle. Bioinformatic analysis of cellular functions, canonical pathways, and upstream regulator of impacted genes underlie the potential mechanisms of butyrate-induced gene expression regulation in rumen epithelium. The introduction of transcriptomic and bioinformatic technologies to study nutrigenomics in the farm animal presented a new prospect to study multiple levels of biological information to better apprehend the whole animal response to nutrition, physiological state, and their interactions. The nutrigenomics approach may eventually lead to more precise management of utilization of feed resources in a more effective approach. PMID:29785087

SPARTA: Simple Program for Automated reference-based bacterial RNA-seq Transcriptome Analysis.

PubMed

Johnson, Benjamin K; Scholz, Matthew B; Teal, Tracy K; Abramovitch, Robert B

2016-02-04

Many tools exist in the analysis of bacterial RNA sequencing (RNA-seq) transcriptional profiling experiments to identify differentially expressed genes between experimental conditions. Generally, the workflow includes quality control of reads, mapping to a reference, counting transcript abundance, and statistical tests for differentially expressed genes. In spite of the numerous tools developed for each component of an RNA-seq analysis workflow, easy-to-use bacterially oriented workflow applications to combine multiple tools and automate the process are lacking. With many tools to choose from for each step, the task of identifying a specific tool, adapting the input/output options to the specific use-case, and integrating the tools into a coherent analysis pipeline is not a trivial endeavor, particularly for microbiologists with limited bioinformatics experience. To make bacterial RNA-seq data analysis more accessible, we developed a Simple Program for Automated reference-based bacterial RNA-seq Transcriptome Analysis (SPARTA). SPARTA is a reference-based bacterial RNA-seq analysis workflow application for single-end Illumina reads. SPARTA is turnkey software that simplifies the process of analyzing RNA-seq data sets, making bacterial RNA-seq analysis a routine process that can be undertaken on a personal computer or in the classroom. The easy-to-install, complete workflow processes whole transcriptome shotgun sequencing data files by trimming reads and removing adapters, mapping reads to a reference, counting gene features, calculating differential gene expression, and, importantly, checking for potential batch effects within the data set. SPARTA outputs quality analysis reports, gene feature counts and differential gene expression tables and scatterplots. SPARTA provides an easy-to-use bacterial RNA-seq transcriptional profiling workflow to identify differentially expressed genes between experimental conditions. This software will enable microbiologists with limited bioinformatics experience to analyze their data and integrate next generation sequencing (NGS) technologies into the classroom. The SPARTA software and tutorial are available at sparta.readthedocs.org.

Circadian oscillatory transcriptional programs in grapevine ripening fruits

PubMed Central

2014-01-01

Background Temperature and solar radiation influence Vitis vinifera L. berry ripening. Both environmental conditions fluctuate cyclically on a daily period basis and the strength of this fluctuation affects grape ripening too. Additionally, a molecular circadian clock regulates daily cyclic expression in a large proportion of the plant transcriptome modulating multiple developmental processes in diverse plant organs and developmental phases. Circadian cycling of fruit transcriptomes has not been characterized in detail despite their putative relevance in the final composition of the fruit. Thus, in this study, gene expression throughout 24 h periods in pre-ripe berries of Tempranillo and Verdejo grapevine cultivars was followed to determine whether different ripening transcriptional programs are activated during certain times of day in different grape tissues and genotypes. Results Microarray analyses identified oscillatory transcriptional profiles following circadian variations in the photocycle and the thermocycle. A higher number of expression oscillating transcripts were detected in samples carrying exocarp tissue including biotic stress-responsive transcripts activated around dawn. Thermotolerance-like responses and regulation of circadian clock-related genes were observed in all studied samples. Indeed, homologs of core clock genes were identified in the grapevine genome and, among them, VvREVEILLE1 (VvRVE1), showed a consistent circadian expression rhythm in every grape berry tissue analysed. Light signalling components and terpenoid biosynthetic transcripts were specifically induced during the daytime in Verdejo, a cultivar bearing white-skinned and aromatic berries, whereas transcripts involved in phenylpropanoid biosynthesis were more prominently regulated in Tempranillo, a cultivar bearing black-skinned berries. Conclusions The transcriptome of ripening fruits varies in response to daily environmental changes, which might partially be under the control of circadian clock components. Certain cultivar and berry tissue features could rely on specific circadian oscillatory expression profiles. These findings may help to a better understanding of the progress of berry ripening in short term time scales. PMID:24666982

Genomic and Transcriptomic Analyses of Indole-3-Acetic Acid Biosynthesis in Diatoms

NASA Astrophysics Data System (ADS)

Lim, R.; Armbrust, V.

2016-02-01

Indole-3-acetic acid (IAA) is a major plant growth hormone and a common mediator of plant-bacterial interactions. Recently, IAA has also been found to play a role in interactions between diatoms and bacteria, with IAA production by an associated Sulfitobacter leading to increased growth rates in the marine diatom Pseudo-nitzschia multiseries. It is unclear, however, if diatoms themselves are able to synthesize IAA and whether this capability is widespread throughout Bacillariophyta. Four major tryptophan-dependent IAA biosynthesis pathways have been identified in plants and bacteria, each denoted by the first intermediate downstream of tryptophan: the indole-3-pyruvate (IPyA), tryptamine (TAM), indole-3-acetaldoxime (IAOx) and indole-3-acetamide (IAM) pathways. To investigate the possibility of IAA biosynthesis in diatoms, we first analyzed publicly available genomes of raphid pennates P. multiseries, Phaeodactylum tricornutum, Fragilariopsis cylindrus and centric Thalassiosira pseudonana for potential homologs to plant and bacterial IAA biosynthesis genes. The P. multiseries, F. cylindrus and P. tricornutum genomes encode downstream enzymes for bacterial TAM and IAM and plant IPyA pathways. The more evolutionarily ancient T. pseudonana encodes one TAM enzyme in its genome. To investigate the potential distribution of these pathways more broadly, we surveyed the transcriptomes of 11 diatom species that include representatives from all four Bacillariophyta classes. Datasets used were sequenced as part of the Marine Microbial Eukaryote Transcriptome Sequencing Project (MMETSP) and obtained from cultures maintained axenically. Transcripts associated with the TAM pathway were most frequently detected, with potential homologs to required enzymes identified in 10 of the 11 species examined. Transcripts homologous to rate-limiting IPyA enzymes were detected in six species. Only two centric and araphid pennate species expressed transcripts associated with enzymes in the IAM and IAOx pathways. This pattern suggests multiple events of gene loss as the phylum expanded and diversified. Mass spectrometry analyses will be conducted to confirm the production of IAA in axenic cultures of P. pungens, P. multistriata, Skeletonema marinoi and F. cylindrus.

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.

PubMed

Chi, Wei; Gao, Yu; Hu, Qing; Guo, Wei; Li, Dapeng

2017-01-01

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.

CLIP-seq analysis of multi-mapped reads discovers novel functional RNA regulatory sites in the human transcriptome.

PubMed

Zhang, Zijun; Xing, Yi

2017-09-19

Crosslinking or RNA immunoprecipitation followed by sequencing (CLIP-seq or RIP-seq) allows transcriptome-wide discovery of RNA regulatory sites. As CLIP-seq/RIP-seq reads are short, existing computational tools focus on uniquely mapped reads, while reads mapped to multiple loci are discarded. We present CLAM (CLIP-seq Analysis of Multi-mapped reads). CLAM uses an expectation-maximization algorithm to assign multi-mapped reads and calls peaks combining uniquely and multi-mapped reads. To demonstrate the utility of CLAM, we applied it to a wide range of public CLIP-seq/RIP-seq datasets involving numerous splicing factors, microRNAs and m6A RNA methylation. CLAM recovered a large number of novel RNA regulatory sites inaccessible by uniquely mapped reads. The functional significance of these sites was demonstrated by consensus motif patterns and association with alternative splicing (splicing factors), transcript abundance (AGO2) and mRNA half-life (m6A). CLAM provides a useful tool to discover novel protein-RNA interactions and RNA modification sites from CLIP-seq and RIP-seq data, and reveals the significant contribution of repetitive elements to the RNA regulatory landscape of the human transcriptome. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Comparative Transcriptomic Analysis of the Response of Dunaliella acidophila (Chlorophyta) to Short-Term Cadmium and Chronic Natural Metal-Rich Water Exposures.

PubMed

Puente-Sánchez, Fernando; Olsson, Sanna; Aguilera, Angeles

2016-10-01

Heavy metals are toxic compounds known to cause multiple and severe cellular damage. However, acidophilic extremophiles are able to cope with very high concentrations of heavy metals. This study investigated the stress response under natural environmental heavy metal concentrations in an acidophilic Dunaliella acidophila. We employed Illumina sequencing for a de novo transcriptome assembly and to identify changes in response to high cadmium concentrations and natural metal-rich water. The photosynthetic performance was also estimated by pulse amplitude-modulated (PAM) fluorescence. Transcriptomic analysis highlights a number of processes mainly related to a high constitutive expression of genes involved in oxidative stress and response to reactive oxygen species (ROS), even in the absence of heavy metals. Photosynthetic activity seems to be unaltered under short-term exposition to Cd and chronic exposure to natural metal-rich water, probably due to an increase in the synthesis of structural photosynthetic components preserving their functional integrity. An overrepresentation of Gene Ontology (GO) terms related to metabolic activities, transcription, and proteosomal catabolic process was observed when D. acidophila grew under chronic exposure to natural metal-rich water. GO terms involved in carbohydrate metabolic process, reticulum endoplasmic and Golgi bodies, were also specifically overrepresented in natural metal-rich water library suggesting an endoplasmic reticulum stress response.

SEASTAR: systematic evaluation of alternative transcription start sites in RNA.

PubMed

Qin, Zhiyi; Stoilov, Peter; Zhang, Xuegong; Xing, Yi

2018-05-04

Alternative first exons diversify the transcriptomes of eukaryotes by producing variants of the 5' Untranslated Regions (5'UTRs) and N-terminal coding sequences. Accurate transcriptome-wide detection of alternative first exons typically requires specialized experimental approaches that are designed to identify the 5' ends of transcripts. We developed a computational pipeline SEASTAR that identifies first exons from RNA-seq data alone then quantifies and compares alternative first exon usage across multiple biological conditions. The exons inferred by SEASTAR coincide with transcription start sites identified directly by CAGE experiments and bear epigenetic hallmarks of active promoters. To determine if differential usage of alternative first exons can yield insights into the mechanism controlling gene expression, we applied SEASTAR to an RNA-seq dataset that tracked the reprogramming of mouse fibroblasts into induced pluripotent stem cells. We observed dynamic temporal changes in the usage of alternative first exons, along with correlated changes in transcription factor expression. Using a combined sequence motif and gene set enrichment analysis we identified N-Myc as a regulator of alternative first exon usage in the pluripotent state. Our results demonstrate that SEASTAR can leverage the available RNA-seq data to gain insights into the control of gene expression and alternative transcript variation in eukaryotic transcriptomes.

Seasonal and Regional Differences in Gene Expression in the Brain of a Hibernating Mammal

PubMed Central

Schwartz, Christine; Hampton, Marshall; Andrews, Matthew T.

2013-01-01

Mammalian hibernation presents a unique opportunity to study naturally occurring neuroprotection. Hibernating ground squirrels undergo rapid and extreme physiological changes in body temperature, oxygen consumption, and heart rate without suffering neurological damage from ischemia and reperfusion injury. Different brain regions show markedly different activity during the torpor/arousal cycle: the cerebral cortex shows activity only during the periodic returns to normothermia, while the hypothalamus is active over the entire temperature range. Therefore, region-specific neuroprotective strategies must exist to permit this compartmentalized spectrum of activity. In this study, we use the Illumina HiSeq platform to compare the transcriptomes of these two brain regions at four collection points across the hibernation season: April Active, October Active, Torpor, and IBA. In the cerebral cortex, 1,085 genes were found to be differentially expressed across collection points, while 1,063 genes were differentially expressed in the hypothalamus. Comparison of these transcripts indicates that the cerebral cortex and hypothalamus implement very different strategies during hibernation, showing less than 20% of these differentially expressed genes in common. The cerebral cortex transcriptome shows evidence of remodeling and plasticity during hibernation, including transcripts for the presynaptic cytomatrix proteins bassoon and piccolo, and extracellular matrix components, including laminins and collagens. Conversely, the hypothalamic transcriptome displays upregulation of transcripts involved in damage response signaling and protein turnover during hibernation, including the DNA damage repair gene RAD50 and ubiquitin E3 ligases UBR1 and UBR5. Additionally, the hypothalamus transcriptome also provides evidence of potential mechanisms underlying the hibernation phenotype, including feeding and satiety signaling, seasonal timing mechanisms, and fuel utilization. This study provides insight into potential neuroprotective strategies and hibernation control mechanisms, and also specifically shows that the hibernator brain exhibits both seasonal and regional differences in mRNA expression. PMID:23526982

De novo transcriptome assembly of drought tolerant CAM plants, Agave deserti and Agave tequilana.

PubMed

Gross, Stephen M; Martin, Jeffrey A; Simpson, June; Abraham-Juarez, María Jazmín; Wang, Zhong; Visel, Axel

2013-08-19

Agaves are succulent monocotyledonous plants native to xeric environments of North America. Because of their adaptations to their environment, including crassulacean acid metabolism (CAM, a water-efficient form of photosynthesis), and existing technologies for ethanol production, agaves have gained attention both as potential lignocellulosic bioenergy feedstocks and models for exploring plant responses to abiotic stress. However, the lack of comprehensive Agave sequence datasets limits the scope of investigations into the molecular-genetic basis of Agave traits. Here, we present comprehensive, high quality de novo transcriptome assemblies of two Agave species, A. tequilana and A. deserti, built from short-read RNA-seq data. Our analyses support completeness and accuracy of the de novo transcriptome assemblies, with each species having a minimum of approximately 35,000 protein-coding genes. Comparison of agave proteomes to those of additional plant species identifies biological functions of gene families displaying sequence divergence in agave species. Additionally, a focus on the transcriptomics of the A. deserti juvenile leaf confirms evolutionary conservation of monocotyledonous leaf physiology and development along the proximal-distal axis. Our work presents a comprehensive transcriptome resource for two Agave species and provides insight into their biology and physiology. These resources are a foundation for further investigation of agave biology and their improvement for bioenergy development.

De novo transcriptome assembly of drought tolerant CAM plants, Agave deserti and Agave tequilana

PubMed Central

2013-01-01

Background Agaves are succulent monocotyledonous plants native to xeric environments of North America. Because of their adaptations to their environment, including crassulacean acid metabolism (CAM, a water-efficient form of photosynthesis), and existing technologies for ethanol production, agaves have gained attention both as potential lignocellulosic bioenergy feedstocks and models for exploring plant responses to abiotic stress. However, the lack of comprehensive Agave sequence datasets limits the scope of investigations into the molecular-genetic basis of Agave traits. Results Here, we present comprehensive, high quality de novo transcriptome assemblies of two Agave species, A. tequilana and A. deserti, built from short-read RNA-seq data. Our analyses support completeness and accuracy of the de novo transcriptome assemblies, with each species having a minimum of approximately 35,000 protein-coding genes. Comparison of agave proteomes to those of additional plant species identifies biological functions of gene families displaying sequence divergence in agave species. Additionally, a focus on the transcriptomics of the A. deserti juvenile leaf confirms evolutionary conservation of monocotyledonous leaf physiology and development along the proximal-distal axis. Conclusions Our work presents a comprehensive transcriptome resource for two Agave species and provides insight into their biology and physiology. These resources are a foundation for further investigation of agave biology and their improvement for bioenergy development. PMID:23957668

Mycobacterium tuberculosis Transcriptome Profiling in Mice with Genetically Different Susceptibility to Tuberculosis.

PubMed

Skvortsov, T A; Ignatov, D V; Majorov, K B; Apt, A S; Azhikina, T L

2013-04-01

Whole transcriptome profiling is now almost routinely used in various fields of biology, including microbiology. In vivo transcriptome studies usually provide relevant information about the biological processes in the organism and thus are indispensable for the formulation of hypotheses, testing, and correcting. In this study, we describe the results of genome-wide transcriptional profiling of the major human bacterial pathogen M. tuberculosis during its persistence in lungs. Two mouse strains differing in their susceptibility to tuberculosis were used for experimental infection with M. tuberculosis. Mycobacterial transcriptomes obtained from the infected tissues of the mice at two different time points were analyzed by deep sequencing and compared. It was hypothesized that the changes in the M. tuberculosis transcriptome may attest to the activation of the metabolism of lipids and amino acids, transition to anaerobic respiration, and increased expression of the factors modulating the immune response. A total of 209 genes were determined whose expression increased with disease progression in both host strains (commonly upregulated genes, CUG). Among them, the genes related to the functional categories of lipid metabolism, cell wall, and cell processes are of great interest. It was assumed that the products of these genes are involved in M. tuberculosis adaptation to the host immune system defense, thus being potential targets for drug development.

Transcriptome analysis in non-model species: a new method for the analysis of heterologous hybridization on microarrays

PubMed Central

2010-01-01

Background Recent developments in high-throughput methods of analyzing transcriptomic profiles are promising for many areas of biology, including ecophysiology. However, although commercial microarrays are available for most common laboratory models, transcriptome analysis in non-traditional model species still remains a challenge. Indeed, the signal resulting from heterologous hybridization is low and difficult to interpret because of the weak complementarity between probe and target sequences, especially when no microarray dedicated to a genetically close species is available. Results We show here that transcriptome analysis in a species genetically distant from laboratory models is made possible by using MAXRS, a new method of analyzing heterologous hybridization on microarrays. This method takes advantage of the design of several commercial microarrays, with different probes targeting the same transcript. To illustrate and test this method, we analyzed the transcriptome of king penguin pectoralis muscle hybridized to Affymetrix chicken microarrays, two organisms separated by an evolutionary distance of approximately 100 million years. The differential gene expression observed between different physiological situations computed by MAXRS was confirmed by real-time PCR on 10 genes out of 11 tested. Conclusions MAXRS appears to be an appropriate method for gene expression analysis under heterologous hybridization conditions. PMID:20509979

Tentacle Transcriptome and Venom Proteome of the Pacific Sea Nettle, Chrysaora fuscescens (Cnidaria: Scyphozoa)

PubMed Central

Ponce, Dalia; Brinkman, Diane L.; Potriquet, Jeremy; Mulvenna, Jason

2016-01-01

Jellyfish venoms are rich sources of toxins designed to capture prey or deter predators, but they can also elicit harmful effects in humans. In this study, an integrated transcriptomic and proteomic approach was used to identify putative toxins and their potential role in the venom of the scyphozoan jellyfish Chrysaora fuscescens. A de novo tentacle transcriptome, containing more than 23,000 contigs, was constructed and used in proteomic analysis of C. fuscescens venom to identify potential toxins. From a total of 163 proteins identified in the venom proteome, 27 were classified as putative toxins and grouped into six protein families: proteinases, venom allergens, C-type lectins, pore-forming toxins, glycoside hydrolases and enzyme inhibitors. Other putative toxins identified in the transcriptome, but not the proteome, included additional proteinases as well as lipases and deoxyribonucleases. Sequence analysis also revealed the presence of ShKT domains in two putative venom proteins from the proteome and an additional 15 from the transcriptome, suggesting potential ion channel blockade or modulatory activities. Comparison of these potential toxins to those from other cnidarians provided insight into their possible roles in C. fuscescens venom and an overview of the diversity of potential toxin families in cnidarian venoms. PMID:27058558

Dynamic evolution of Geranium mitochondrial genomes through multiple horizontal and intracellular gene transfers.

PubMed

Park, Seongjun; Grewe, Felix; Zhu, Andan; Ruhlman, Tracey A; Sabir, Jamal; Mower, Jeffrey P; Jansen, Robert K

2015-10-01

The exchange of genetic material between cellular organelles through intracellular gene transfer (IGT) or between species by horizontal gene transfer (HGT) has played an important role in plant mitochondrial genome evolution. The mitochondrial genomes of Geraniaceae display a number of unusual phenomena including highly accelerated rates of synonymous substitutions, extensive gene loss and reduction in RNA editing. Mitochondrial DNA sequences assembled for 17 species of Geranium revealed substantial reduction in gene and intron content relative to the ancestor of the Geranium lineage. Comparative analyses of nuclear transcriptome data suggest that a number of these sequences have been functionally relocated to the nucleus via IGT. Evidence for rampant HGT was detected in several Geranium species containing foreign organellar DNA from diverse eudicots, including many transfers from parasitic plants. One lineage has experienced multiple, independent HGT episodes, many of which occurred within the past 5.5 Myr. Both duplicative and recapture HGT were documented in Geranium lineages. The mitochondrial genome of Geranium brycei contains at least four independent HGT tracts that are absent in its nearest relative. Furthermore, G. brycei mitochondria carry two copies of the cox1 gene that differ in intron content, providing insight into contrasting hypotheses on cox1 intron evolution. © 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.

Transcriptional profiling suggests that multiple metabolic adaptations are required for effective proliferation of Pseudomonas aeruginosa in jet fuel.

PubMed

Gunasekera, Thusitha S; Striebich, Richard C; Mueller, Susan S; Strobel, Ellen M; Ruiz, Oscar N

2013-01-01

Fuel is a harsh environment for microbial growth. However, some bacteria can grow well due to their adaptive mechanisms. Our goal was to characterize the adaptations required for Pseudomonas aeruginosa proliferation in fuel. We have used DNA-microarrays and RT-PCR to characterize the transcriptional response of P. aeruginosa to fuel. Transcriptomics revealed that genes essential for medium- and long-chain n-alkane degradation including alkB1 and alkB2 were transcriptionally induced. Gas chromatography confirmed that P. aeruginosa possesses pathways to degrade different length n-alkanes, favoring the use of n-C11-18. Furthermore, a gamut of synergistic metabolic pathways, including porins, efflux pumps, biofilm formation, and iron transport, were transcriptionally regulated. Bioassays confirmed that efflux pumps and biofilm formation were required for growth in jet fuel. Furthermore, cell homeostasis appeared to be carefully maintained by the regulation of porins and efflux pumps. The Mex RND efflux pumps were required for fuel tolerance; blockage of these pumps precluded growth in fuel. This study provides a global understanding of the multiple metabolic adaptations required by bacteria for survival and proliferation in fuel-containing environments. This information can be applied to improve the fuel bioremediation properties of bacteria.

Insights into the innate immunome of actiniarians using a comparative genomic approach.

PubMed

van der Burg, Chloé A; Prentis, Peter J; Surm, Joachim M; Pavasovic, Ana

2016-11-02

Innate immune genes tend to be highly conserved in metazoans, even in early divergent lineages such as Cnidaria (jellyfish, corals, hydroids and sea anemones) and Porifera (sponges). However, constant and diverse selection pressures on the immune system have driven the expansion and diversification of different immune gene families in a lineage-specific manner. To investigate how the innate immune system has evolved in a subset of sea anemone species (Order: Actiniaria), we performed a comprehensive and comparative study using 10 newly sequenced transcriptomes, as well as three publically available transcriptomes, to identify the origins, expansions and contractions of candidate and novel immune gene families. We characterised five conserved genes and gene families, as well as multiple novel innate immune genes, including the newly recognised putative pattern recognition receptor CniFL. Single copies of TLR, MyD88 and NF-κB were found in most species, and several copies of IL-1R-like, NLR and CniFL were found in almost all species. Multiple novel immune genes were identified with domain architectures including the Toll/interleukin-1 receptor (TIR) homology domain, which is well documented as functioning in protein-protein interactions and signal transduction in immune pathways. We hypothesise that these genes may interact as novel proteins in immune pathways of cnidarian species. Novelty in the actiniarian immunome is not restricted to only TIR-domain-containing proteins, as we identify a subset of NLRs which have undergone neofunctionalisation and contain 3-5 N-terminal transmembrane domains, which have so far only been identified in two anthozoan species. This research has significance in understanding the evolution and origin of the core eumetazoan gene set, including how novel innate immune genes evolve. For example, the evolution of transmembrane domain containing NLRs indicates that these NLRs may be membrane-bound, while all other metazoan and plant NLRs are exclusively cytosolic receptors. This is one example of how species without an adaptive immune system may evolve innovative solutions to detect pathogens or interact with native microbiota. Overall, these results provide an insight into the evolution of the innate immune system, and show that early divergent lineages, such as actiniarians, have a diverse repertoire of conserved and novel innate immune genes.

Transcriptome landscape of Lactococcus lactis reveals many novel RNAs including a small regulatory RNA involved in carbon uptake and metabolism.

PubMed

van der Meulen, Sjoerd B; de Jong, Anne; Kok, Jan

2016-01-01

RNA sequencing has revolutionized genome-wide transcriptome analyses, and the identification of non-coding regulatory RNAs in bacteria has thus increased concurrently. Here we reveal the transcriptome map of the lactic acid bacterial paradigm Lactococcus lactis MG1363 by employing differential RNA sequencing (dRNA-seq) and a combination of manual and automated transcriptome mining. This resulted in a high-resolution genome annotation of L. lactis and the identification of 60 cis-encoded antisense RNAs (asRNAs), 186 trans-encoded putative regulatory RNAs (sRNAs) and 134 novel small ORFs. Based on the putative targets of asRNAs, a novel classification is proposed. Several transcription factor DNA binding motifs were identified in the promoter sequences of (a)sRNAs, providing insight in the interplay between lactococcal regulatory RNAs and transcription factors. The presence and lengths of 14 putative sRNAs were experimentally confirmed by differential Northern hybridization, including the abundant RNA 6S that is differentially expressed depending on the available carbon source. For another sRNA, LLMGnc_147, functional analysis revealed that it is involved in carbon uptake and metabolism. L. lactis contains 13% leaderless mRNAs (lmRNAs) that, from an analysis of overrepresentation in GO classes, seem predominantly involved in nucleotide metabolism and DNA/RNA binding. Moreover, an A-rich sequence motif immediately following the start codon was uncovered, which could provide novel insight in the translation of lmRNAs. Altogether, this first experimental genome-wide assessment of the transcriptome landscape of L. lactis and subsequent sRNA studies provide an extensive basis for the investigation of regulatory RNAs in L. lactis and related lactococcal species.

The immune gene repertoire of an important viral reservoir, the Australian black flying fox.

PubMed

Papenfuss, Anthony T; Baker, Michelle L; Feng, Zhi-Ping; Tachedjian, Mary; Crameri, Gary; Cowled, Chris; Ng, Justin; Janardhana, Vijaya; Field, Hume E; Wang, Lin-Fa

2012-06-20

Bats are the natural reservoir host for a range of emerging and re-emerging viruses, including SARS-like coronaviruses, Ebola viruses, henipaviruses and Rabies viruses. However, the mechanisms responsible for the control of viral replication in bats are not understood and there is little information available on any aspect of antiviral immunity in bats. Massively parallel sequencing of the bat transcriptome provides the opportunity for rapid gene discovery. Although the genomes of one megabat and one microbat have now been sequenced to low coverage, no transcriptomic datasets have been reported from any bat species. In this study, we describe the immune transcriptome of the Australian flying fox, Pteropus alecto, providing an important resource for identification of genes involved in a range of activities including antiviral immunity. Towards understanding the adaptations that have allowed bats to coexist with viruses, we have de novo assembled transcriptome sequence from immune tissues and stimulated cells from P. alecto. We identified about 18,600 genes involved in a broad range of activities with the most highly expressed genes involved in cell growth and maintenance, enzyme activity, cellular components and metabolism and energy pathways. 3.5% of the bat transcribed genes corresponded to immune genes and a total of about 500 immune genes were identified, providing an overview of both innate and adaptive immunity. A small proportion of transcripts found no match with annotated sequences in any of the public databases and may represent bat-specific transcripts. This study represents the first reported bat transcriptome dataset and provides a survey of expressed bat genes that complement existing bat genomic data. In addition, these data provide insight into genes relevant to the antiviral responses of bats, and form a basis for examining the roles of these molecules in immune response to viral infection.

Analyses of transcriptome sequences reveal multiple ancient large-scale duplication events in the ancestor of Sphagnopsida (Bryophyta).

PubMed

Devos, Nicolas; Szövényi, Péter; Weston, David J; Rothfels, Carl J; Johnson, Matthew G; Shaw, A Jonathan

2016-07-01

The goal of this research was to investigate whether there has been a whole-genome duplication (WGD) in the ancestry of Sphagnum (peatmoss) or the class Sphagnopsida, and to determine if the timing of any such duplication(s) and patterns of paralog retention could help explain the rapid radiation and current ecological dominance of peatmosses. RNA sequencing (RNA-seq) data were generated for nine taxa in Sphagnopsida (Bryophyta). Analyses of frequency plots for synonymous substitutions per synonymous site (Ks ) between paralogous gene pairs and reconciliation of 578 gene trees were conducted to assess evidence of large-scale or genome-wide duplication events in each transcriptome. Both Ks frequency plots and gene tree-based analyses indicate multiple duplication events in the history of the Sphagnopsida. The most recent WGD event predates divergence of Sphagnum from the two other genera of Sphagnopsida. Duplicate retention is highly variable across species, which might be best explained by local adaptation. Our analyses indicate that the last WGD could have been an important factor underlying the diversification of peatmosses and facilitated their rise to ecological dominance in peatlands. The timing of the duplication events and their significance in the evolutionary history of peat mosses are discussed. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

SncRNA (microRNA & snoRNA) opposite expression pattern found in multiple sclerosis relapse and remission is sex dependent

PubMed Central

Muñoz-Culla, Maider; Irizar, Haritz; Sáenz-Cuesta, Matías; Castillo-Triviño, Tamara; Osorio-Querejeta, Iñaki; Sepúlveda, Lucía; López de Munain, Adolfo; Olascoaga, Javier; Otaegui, David

2016-01-01

Multiple sclerosis (MS) is a common inflammatory and degenerative disease that causes neurological disability. It affects young adults and its prevalence is higher in women. The most common form is manifested as a series of acute episodes of neurological disability (relapses) followed by a recovery phase (remission). Recently, non-coding RNAs have emerged as new players in transcriptome regulation, and in turn, they could have a significant role in MS pathogenesis. In this context, our aim was to investigate the involvement of microRNAs and snoRNAs in the relapse-remission dynamics of MS in peripheral blood leucocytes, to shed light on the molecular and regulatory mechanisms that underlie this complex process. With this approach, we found that a subset of small non-coding RNAs (sncRNA) is altered in relapse and remission, revealing unexpected opposite changes that are sex dependent. Furthermore, we found that a relapse-related miRNA signature regulated general metabolism processes in leucocytes, and miRNA altered in remission are involved in the regulation of innate immunity. We observed that sncRNA dysregulation is different in relapse and remission leading to differences in transcriptome regulation, and that this process is sex dependent. In conclusion, relapse and remission have a different molecular background in men and women. PMID:26831009

Transcriptome changes associated with Tomato spotted wilt virus infection in various life stages of its thrips vector, Frankliniella fusca (Hinds).

PubMed

Shrestha, Anita; Champagne, Donald E; Culbreath, Albert K; Rotenberg, Dorith; Whitfield, Anna E; Srinivasan, Rajagopalbabu

2017-08-01

Persistent propagative viruses maintain intricate interactions with their arthropod vectors. In this study, we investigated the transcriptome-level responses associated with a persistent propagative phytovirus infection in various life stages of its vector using an Illumina HiSeq sequencing platform. The pathosystem components included a Tospovirus, Tomato spotted wilt virus (TSWV), its insect vector, Frankliniella fusca (Hinds), and a plant host, Arachis hypogaea (L.). We assembled (de novo) reads from three developmental stage groups of virus-exposed and non-virus-exposed F. fusca into one transcriptome consisting of 72 366 contigs and identified 1161 differentially expressed (DE) contigs. The number of DE contigs was greatest in adults (female) (562) when compared with larvae (first and second instars) (395) and pupae (pre- and pupae) (204). Upregulated contigs in virus-exposed thrips had blastx annotations associated with intracellular transport and virus replication. Upregulated contigs were also assigned blastx annotations associated with immune responses, including apoptosis and phagocytosis. In virus-exposed larvae, Blast2GO analysis identified functional groups, such as multicellular development with downregulated contigs, while reproduction, embryo development and growth were identified with upregulated contigs in virus-exposed adults. This study provides insights into differences in transcriptome-level responses modulated by TSWV in various life stages of an important vector, F. fusca.

Concurrent Host-Pathogen Transcriptional Responses in a Clostridium perfringens Murine Myonecrosis Infection

PubMed Central

2018-01-01

ABSTRACT To obtain an insight into host-pathogen interactions in clostridial myonecrosis, we carried out comparative transcriptome analysis of both the bacterium and the host in a murine Clostridium perfringens infection model, which is the first time that such an investigation has been conducted. Analysis of the host transcriptome from infected muscle tissues indicated that many genes were upregulated compared to the results seen with mock-infected mice. These genes were enriched for host defense pathways, including Toll-like receptor (TLR) and Nod-like receptor (NLR) signaling components. Real-time PCR confirmed that host TLR2 and NLRP3 inflammasome genes were induced in response to C. perfringens infection. Comparison of the transcriptome of C. perfringens cells from the infected tissues with that from broth cultures showed that host selective pressure induced a global change in C. perfringens gene expression. A total of 33% (923) of C. perfringens genes were differentially regulated, including 10 potential virulence genes that were upregulated relative to their expression in vitro. These genes encoded putative proteins that may be involved in the synthesis of cell wall-associated macromolecules, in adhesion to host cells, or in protection from host cationic antimicrobial peptides. This report presents the first successful expression profiling of coregulated transcriptomes of bacterial and host genes during a clostridial myonecrosis infection and provides new insights into disease pathogenesis and host-pathogen interactions. PMID:29588405

Transcriptome Dynamics during Maize Endosperm Development

PubMed Central

Feng, Jiaojiao; Xu, Shutu; Wang, Lei; Li, Feifei; Li, Yibo; Zhang, Renhe; Zhang, Xinghua; Xue, Jiquan; Guo, Dongwei

2016-01-01

The endosperm is a major organ of the seed that plays vital roles in determining seed weight and quality. However, genome-wide transcriptome patterns throughout maize endosperm development have not been comprehensively investigated to date. Accordingly, we performed a high-throughput RNA sequencing (RNA-seq) analysis of the maize endosperm transcriptome at 5, 10, 15 and 20 days after pollination (DAP). We found that more than 11,000 protein-coding genes underwent alternative splicing (AS) events during the four developmental stages studied. These genes were mainly involved in intracellular protein transport, signal transmission, cellular carbohydrate metabolism, cellular lipid metabolism, lipid biosynthesis, protein modification, histone modification, cellular amino acid metabolism, and DNA repair. Additionally, 7,633 genes, including 473 transcription factors (TFs), were differentially expressed among the four developmental stages. The differentially expressed TFs were from 50 families, including the bZIP, WRKY, GeBP and ARF families. Further analysis of the stage-specific TFs showed that binding, nucleus and ligand-dependent nuclear receptor activities might be important at 5 DAP, that immune responses, signalling, binding and lumen development are involved at 10 DAP, that protein metabolic processes and the cytoplasm might be important at 15 DAP, and that the responses to various stimuli are different at 20 DAP compared with the other developmental stages. This RNA-seq analysis provides novel, comprehensive insights into the transcriptome dynamics during early endosperm development in maize. PMID:27695101

Antennal Transcriptome Analysis and Comparison of Chemosensory Gene Families in Two Closely Related Noctuidae Moths, Helicoverpa armigera and H. assulta

PubMed Central

Zhang, Jin; Wang, Bing; Dong, Shuanglin; Cao, Depan; Dong, Junfeng; Walker, William B.; Liu, Yang; Wang, Guirong

2015-01-01

To better understand the olfactory mechanisms in the two lepidopteran pest model species, the Helicoverpa armigera and H. assulta, we conducted transcriptome analysis of the adult antennae using Illumina sequencing technology and compared the chemosensory genes between these two related species. Combined with the chemosensory genes we had identified previously in H. armigera by 454 sequencing, we identified 133 putative chemosensory unigenes in H. armigera including 60 odorant receptors (ORs), 19 ionotropic receptors (IRs), 34 odorant binding proteins (OBPs), 18 chemosensory proteins (CSPs), and 2 sensory neuron membrane proteins (SNMPs). Consistent with these results, 131 putative chemosensory genes including 64 ORs, 19 IRs, 29 OBPs, 17 CSPs, and 2 SNMPs were identified through male and female antennal transcriptome analysis in H. assulta. Reverse Transcription-PCR (RT-PCR) was conducted in H. assulta to examine the accuracy of the assembly and annotation of the transcriptome and the expression profile of these unigenes in different tissues. Most of the ORs, IRs and OBPs were enriched in adult antennae, while almost all the CSPs were expressed in antennae as well as legs. We compared the differences of the chemosensory genes between these two species in detail. Our work will surely provide valuable information for further functional studies of pheromones and host volatile recognition genes in these two related species. PMID:25659090

Transcriptomics and molecular evolutionary rate analysis of the bladderwort (Utricularia), a carnivorous plant with a minimal genome

PubMed Central

2011-01-01

Background The carnivorous plant Utricularia gibba (bladderwort) is remarkable in having a minute genome, which at ca. 80 megabases is approximately half that of Arabidopsis. Bladderworts show an incredible diversity of forms surrounding a defined theme: tiny, bladder-like suction traps on terrestrial, epiphytic, or aquatic plants with a diversity of unusual vegetative forms. Utricularia plants, which are rootless, are also anomalous in physiological features (respiration and carbon distribution), and highly enhanced molecular evolutionary rates in chloroplast, mitochondrial and nuclear ribosomal sequences. Despite great interest in the genus, no genomic resources exist for Utricularia, and the substitution rate increase has received limited study. Results Here we describe the sequencing and analysis of the Utricularia gibba transcriptome. Three different organs were surveyed, the traps, the vegetative shoot bodies, and the inflorescence stems. We also examined the bladderwort transcriptome under diverse stress conditions. We detail aspects of functional classification, tissue similarity, nitrogen and phosphorus metabolism, respiration, DNA repair, and detoxification of reactive oxygen species (ROS). Long contigs of plastid and mitochondrial genomes, as well as sequences for 100 individual nuclear genes, were compared with those of other plants to better establish information on molecular evolutionary rates. Conclusion The Utricularia transcriptome provides a detailed genomic window into processes occurring in a carnivorous plant. It contains a deep representation of the complex metabolic pathways that characterize a putative minimal plant genome, permitting its use as a source of genomic information to explore the structural, functional, and evolutionary diversity of the genus. Vegetative shoots and traps are the most similar organs by functional classification of their transcriptome, the traps expressing hydrolytic enzymes for prey digestion that were previously thought to be encoded by bacteria. Supporting physiological data, global gene expression analysis shows that traps significantly over-express genes involved in respiration and that phosphate uptake might occur mainly in traps, whereas nitrogen uptake could in part take place in vegetative parts. Expression of DNA repair and ROS detoxification enzymes may be indicative of a response to increased respiration. Finally, evidence from the bladderwort transcriptome, direct measurement of ROS in situ, and cross-species comparisons of organellar genomes and multiple nuclear genes supports the hypothesis that increased nucleotide substitution rates throughout the plant may be due to the mutagenic action of amplified ROS production. PMID:21639913

Systems biology of human atherosclerosis.

PubMed

Shalhoub, Joseph; Sikkel, Markus B; Davies, Kerry J; Vorkas, Panagiotis A; Want, Elizabeth J; Davies, Alun H

2014-01-01

Systems biology describes a holistic and integrative approach to understand physiology and pathology. The "omic" disciplines include genomics, transcriptomics, proteomics, and metabolic profiling (metabonomics and metabolomics). By adopting a stance, which is opposing (yet complimentary) to conventional research techniques, systems biology offers an overview by assessing the "net" biological effect imposed by a disease or nondisease state. There are a number of different organizational levels to be understood, from DNA to protein, metabolites, cells, organs and organisms, even beyond this to an organism's context. Systems biology relies on the existence of "nodes" and "edges." Nodes are the constituent part of the system being studied (eg, proteins in the proteome), while the edges are the way these constituents interact. In future, it will be increasingly important to collaborate, collating data from multiple studies to improve data sets, making them freely available and undertaking integrative analyses.

ABMapper: a suffix array-based tool for multi-location searching and splice-junction mapping.

PubMed

Lou, Shao-Ke; Ni, Bing; Lo, Leung-Yau; Tsui, Stephen Kwok-Wing; Chan, Ting-Fung; Leung, Kwong-Sak

2011-02-01

Sequencing reads generated by RNA-sequencing (RNA-seq) must first be mapped back to the genome through alignment before they can be further analyzed. Current fast and memory-saving short-read mappers could give us a quick view of the transcriptome. However, they are neither designed for reads that span across splice junctions nor for repetitive reads, which can be mapped to multiple locations in the genome (multi-reads). Here, we describe a new software package: ABMapper, which is specifically designed for exploring all putative locations of reads that are mapped to splice junctions or repetitive in nature. The software is freely available at: http://abmapper.sourceforge.net/. The software is written in C++ and PERL. It runs on all major platforms and operating systems including Windows, Mac OS X and LINUX.

Transcriptome analysis of Petunia axillaris flowers reveals genes involved in morphological differentiation and metabolite transport

PubMed Central

Amano, Ikuko; Kitajima, Sakihito; Suzuki, Hideyuki; Koeduka, Takao

2018-01-01

The biosynthesis of plant secondary metabolites is associated with morphological and metabolic differentiation. As a consequence, gene expression profiles can change drastically, and primary and secondary metabolites, including intermediate and end-products, move dynamically within and between cells. However, little is known about the molecular mechanisms underlying differentiation and transport mechanisms. In this study, we performed a transcriptome analysis of Petunia axillaris subsp. parodii, which produces various volatiles in its corolla limbs and emits metabolites to attract pollinators. RNA-sequencing from leaves, buds, and limbs identified 53,243 unigenes. Analysis of differentially expressed genes, combined with gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses, showed that many biological processes were highly enriched in limbs. These included catabolic processes and signaling pathways of hormones, such as gibberellins, and metabolic pathways, including phenylpropanoids and fatty acids. Moreover, we identified five transporter genes that showed high expression in limbs, and we performed spatiotemporal expression analyses and homology searches to infer their putative functions. Our systematic analysis provides comprehensive transcriptomic information regarding morphological differentiation and metabolite transport in the Petunia flower and lays the foundation for establishing the specific mechanisms that control secondary metabolite biosynthesis in plants. PMID:29902274

Hyperexpansion of RNA Bacteriophage Diversity

PubMed Central

Krishnamurthy, Siddharth R.; Janowski, Andrew B.; Zhao, Guoyan; Barouch, Dan; Wang, David

2016-01-01

Bacteriophage modulation of microbial populations impacts critical processes in ocean, soil, and animal ecosystems. However, the role of bacteriophages with RNA genomes (RNA bacteriophages) in these processes is poorly understood, in part because of the limited number of known RNA bacteriophage species. Here, we identify partial genome sequences of 122 RNA bacteriophage phylotypes that are highly divergent from each other and from previously described RNA bacteriophages. These novel RNA bacteriophage sequences were present in samples collected from a range of ecological niches worldwide, including invertebrates and extreme microbial sediment, demonstrating that they are more widely distributed than previously recognized. Genomic analyses of these novel bacteriophages yielded multiple novel genome organizations. Furthermore, one RNA bacteriophage was detected in the transcriptome of a pure culture of Streptomyces avermitilis, suggesting for the first time that the known tropism of RNA bacteriophages may include gram-positive bacteria. Finally, reverse transcription PCR (RT-PCR)-based screening for two specific RNA bacteriophages in stool samples from a longitudinal cohort of macaques suggested that they are generally acutely present rather than persistent. PMID:27010970

Coexpression network analysis identifies transcriptional modules associated with genomic alterations in neuroblastoma.

PubMed

Yang, Liulin; Li, Yun; Wei, Zhi; Chang, Xiao

2018-06-01

Neuroblastoma is a highly complex and heterogeneous cancer in children. Acquired genomic alterations including MYCN amplification, 1p deletion and 11q deletion are important risk factors and biomarkers in neuroblastoma. Here, we performed a co-expression-based gene network analysis to study the intrinsic association between specific genomic changes and transcriptome organization. We identified multiple gene coexpression modules which are recurrent in two independent datasets and associated with functional pathways including nervous system development, cell cycle, immune system process and extracellular matrix/space. Our results also indicated that modules involved in nervous system development and cell cycle are highly associated with MYCN amplification and 1p deletion, while modules responding to immune system process are associated with MYCN amplification only. In summary, this integrated analysis provides novel insights into molecular heterogeneity and pathogenesis of neuroblastoma. This article is part of a Special Issue entitled: Accelerating Precision Medicine through Genetic and Genomic Big Data Analysis edited by Yudong Cai & Tao Huang. Copyright © 2017. Published by Elsevier B.V.

An Update on ToxCast™ | Science Inventory | US EPA

EPA Pesticide Factsheets

In its first phase, ToxCast™ is profiling over 300 well-characterized chemicals (primarily pesticides) in over 400 HTS endpoints. These endpoints include biochemical assays of protein function, cell-based transcriptional reporter assays, multi-cell interaction assays, transcriptomics on primary cell cultures, and developmental assays in zebrafish embryos. Almost all of the compounds being examined in Phase 1 of ToxCast™ have been tested in traditional toxicology tests, including developmental toxicity, multi-generation studies, and sub-chronic and chronic rodent bioassays Lessons learned to date for ToxCast: Large amounts of quality HTS data can be economically obtained. Large scale data sets will be required to understand potential for biological activity. Value in having multiple assays with overlapping coverage of biological pathways and a variety of methodologies Concentration-response will be important for ultimate interpretation Data transparency will be important for acceptance. Metabolic capabilities and coverage of developmental toxicity pathways will need additional attention. Need to define the gold standard Partnerships are needed to bring critical mass and expertise.

Assessing the hodgepodge of non-mapped reads in bacterial transcriptomes: real or artifactual RNA chimeras?

PubMed

Lloréns-Rico, Verónica; Serrano, Luis; Lluch-Senar, Maria

2014-07-29

RNA sequencing methods have already altered our view of the extent and complexity of bacterial and eukaryotic transcriptomes, revealing rare transcript isoforms (circular RNAs, RNA chimeras) that could play an important role in their biology. We performed an analysis of chimera formation by four different computational approaches, including a custom designed pipeline, to study the transcriptomes of M. pneumoniae and P. aeruginosa, as well as mixtures of both. We found that rare transcript isoforms detected by conventional pipelines of analysis could be artifacts of the experimental procedure used in the library preparation, and that they are protocol-dependent. By using a customized pipeline we show that optimal library preparation protocol and the pipeline to analyze the results are crucial to identify real chimeric RNAs.

An RNA-binding protein, Qki5, regulates embryonic neural stem cells through pre-mRNA processing in cell adhesion signaling.

PubMed

Hayakawa-Yano, Yoshika; Suyama, Satoshi; Nogami, Masahiro; Yugami, Masato; Koya, Ikuko; Furukawa, Takako; Zhou, Li; Abe, Manabu; Sakimura, Kenji; Takebayashi, Hirohide; Nakanishi, Atsushi; Okano, Hideyuki; Yano, Masato

2017-09-15

Cell type-specific transcriptomes are enabled by the action of multiple regulators, which are frequently expressed within restricted tissue regions. In the present study, we identify one such regulator, Quaking 5 (Qki5), as an RNA-binding protein (RNABP) that is expressed in early embryonic neural stem cells and subsequently down-regulated during neurogenesis. mRNA sequencing analysis in neural stem cell culture indicates that Qki proteins play supporting roles in the neural stem cell transcriptome and various forms of mRNA processing that may result from regionally restricted expression and subcellular localization. Also, our in utero electroporation gain-of-function study suggests that the nuclear-type Qki isoform Qki5 supports the neural stem cell state. We next performed in vivo transcriptome-wide protein-RNA interaction mapping to search for direct targets of Qki5 and elucidate how Qki5 regulates neural stem cell function. Combined with our transcriptome analysis, this mapping analysis yielded a bona fide map of Qki5-RNA interaction at single-nucleotide resolution, the identification of 892 Qki5 direct target genes, and an accurate Qki5-dependent alternative splicing rule in the developing brain. Last, our target gene list provides the first compelling evidence that Qki5 is associated with specific biological events; namely, cell-cell adhesion. This prediction was confirmed by histological analysis of mice in which Qki proteins were genetically ablated, which revealed disruption of the apical surface of the lateral wall in the developing brain. These data collectively indicate that Qki5 regulates communication between neural stem cells by mediating numerous RNA processing events and suggest new links between splicing regulation and neural stem cell states. © 2017 Hayakawa-Yano et al.; Published by Cold Spring Harbor Laboratory Press.

CrossQuery: a web tool for easy associative querying of transcriptome data.

PubMed

Wagner, Toni U; Fischer, Andreas; Thoma, Eva C; Schartl, Manfred

2011-01-01

Enormous amounts of data are being generated by modern methods such as transcriptome or exome sequencing and microarray profiling. Primary analyses such as quality control, normalization, statistics and mapping are highly complex and need to be performed by specialists. Thereafter, results are handed back to biomedical researchers, who are then confronted with complicated data lists. For rather simple tasks like data filtering, sorting and cross-association there is a need for new tools which can be used by non-specialists. Here, we describe CrossQuery, a web tool that enables straight forward, simple syntax queries to be executed on transcriptome sequencing and microarray datasets. We provide deep-sequencing data sets of stem cell lines derived from the model fish Medaka and microarray data of human endothelial cells. In the example datasets provided, mRNA expression levels, gene, transcript and sample identification numbers, GO-terms and gene descriptions can be freely correlated, filtered and sorted. Queries can be saved for later reuse and results can be exported to standard formats that allow copy-and-paste to all widespread data visualization tools such as Microsoft Excel. CrossQuery enables researchers to quickly and freely work with transcriptome and microarray data sets requiring only minimal computer skills. Furthermore, CrossQuery allows growing association of multiple datasets as long as at least one common point of correlated information, such as transcript identification numbers or GO-terms, is shared between samples. For advanced users, the object-oriented plug-in and event-driven code design of both server-side and client-side scripts allow easy addition of new features, data sources and data types.

Hierarchical cortical transcriptome disorganization in autism.

PubMed

Lombardo, Michael V; Courchesne, Eric; Lewis, Nathan E; Pramparo, Tiziano

2017-01-01

Autism spectrum disorders (ASD) are etiologically heterogeneous and complex. Functional genomics work has begun to identify a diverse array of dysregulated transcriptomic programs (e.g., synaptic, immune, cell cycle, DNA damage, WNT signaling, cortical patterning and differentiation) potentially involved in ASD brain abnormalities during childhood and adulthood. However, it remains unclear whether such diverse dysregulated pathways are independent of each other or instead reflect coordinated hierarchical systems-level pathology. Two ASD cortical transcriptome datasets were re-analyzed using consensus weighted gene co-expression network analysis (WGCNA) to identify common co-expression modules across datasets. Linear mixed-effect models and Bayesian replication statistics were used to identify replicable differentially expressed modules. Eigengene network analysis was then utilized to identify between-group differences in how co-expression modules interact and cluster into hierarchical meta-modular organization. Protein-protein interaction analyses were also used to determine whether dysregulated co-expression modules show enhanced interactions. We find replicable evidence for 10 gene co-expression modules that are differentially expressed in ASD cortex. Rather than being independent non-interacting sources of pathology, these dysregulated co-expression modules work in synergy and physically interact at the protein level. These systems-level transcriptional signals are characterized by downregulation of synaptic processes coordinated with upregulation of immune/inflammation, response to other organism, catabolism, viral processes, translation, protein targeting and localization, cell proliferation, and vasculature development. Hierarchical organization of meta-modules (clusters of highly correlated modules) is also highly affected in ASD. These findings highlight that dysregulation of the ASD cortical transcriptome is characterized by the dysregulation of multiple coordinated transcriptional programs producing synergistic systems-level effects that cannot be fully appreciated by studying the individual component biological processes in isolation.

Major transcriptome re-organisation and abrupt changes in signalling, cell cycle and chromatin regulation at neural differentiation in vivo.

PubMed

Olivera-Martinez, Isabel; Schurch, Nick; Li, Roman A; Song, Junfang; Halley, Pamela A; Das, Raman M; Burt, Dave W; Barton, Geoffrey J; Storey, Kate G

2014-08-01

Here, we exploit the spatial separation of temporal events of neural differentiation in the elongating chick body axis to provide the first analysis of transcriptome change in progressively more differentiated neural cell populations in vivo. Microarray data, validated against direct RNA sequencing, identified: (1) a gene cohort characteristic of the multi-potent stem zone epiblast, which contains neuro-mesodermal progenitors that progressively generate the spinal cord; (2) a major transcriptome re-organisation as cells then adopt a neural fate; and (3) increasing diversity as neural patterning and neuron production begin. Focussing on the transition from multi-potent to neural state cells, we capture changes in major signalling pathways, uncover novel Wnt and Notch signalling dynamics, and implicate new pathways (mevalonate pathway/steroid biogenesis and TGFβ). This analysis further predicts changes in cellular processes, cell cycle, RNA-processing and protein turnover as cells acquire neural fate. We show that these changes are conserved across species and provide biological evidence for reduced proteasome efficiency and a novel lengthening of S phase. This latter step may provide time for epigenetic events to mediate large-scale transcriptome re-organisation; consistent with this, we uncover simultaneous downregulation of major chromatin modifiers as the neural programme is established. We further demonstrate that transcription of one such gene, HDAC1, is dependent on FGF signalling, making a novel link between signals that control neural differentiation and transcription of a core regulator of chromatin organisation. Our work implicates new signalling pathways and dynamics, cellular processes and epigenetic modifiers in neural differentiation in vivo, identifying multiple new potential cellular and molecular mechanisms that direct differentiation. © 2014. Published by The Company of Biologists Ltd.

Improving transcriptome de novo assembly by using a reference genome of a related species: Translational genomics from oil palm to coconut.

PubMed

Armero, Alix; Baudouin, Luc; Bocs, Stéphanie; This, Dominique

2017-01-01

The palms are a family of tropical origin and one of the main constituents of the ecosystems of these regions around the world. The two main species of palm represent different challenges: coconut (Cocos nucifera L.) is a source of multiple goods and services in tropical communities, while oil palm (Elaeis guineensis Jacq) is the main protagonist of the oil market. In this study, we present a workflow that exploits the comparative genomics between a target species (coconut) and a reference species (oil palm) to improve the transcriptomic data, providing a proteome useful to answer functional or evolutionary questions. This workflow reduces redundancy and fragmentation, two inherent problems of transcriptomic data, while preserving the functional representation of the target species. Our approach was validated in Arabidopsis thaliana using Arabidopsis lyrata and Capsella rubella as references species. This analysis showed the high sensitivity and specificity of our strategy, relatively independent of the reference proteome. The workflow increased the length of proteins products in A. thaliana by 13%, allowing, often, to recover 100% of the protein sequence length. In addition redundancy was reduced by a factor greater than 3. In coconut, the approach generated 29,366 proteins, 1,246 of these proteins deriving from new contigs obtained with the BRANCH software. The coconut proteome presented a functional profile similar to that observed in rice and an important number of metabolic pathways related to secondary metabolism. The new sequences found with BRANCH software were enriched in functions related to biotic stress. Our strategy can be used as a complementary step to de novo transcriptome assembly to get a representative proteome of a target species. The results of the current analysis are available on the website PalmComparomics (http://palm-comparomics.southgreen.fr/).

De Novo Assembly and Characterization of Two Transcriptomes Reveal Multiple Light-Mediated Functions in the Scallop Eye (Bivalvia: Pectinidae)

PubMed Central

Pairett, Autum N.; Serb, Jeanne M.

2013-01-01

Background The eye has evolved across 13 separate lineages of molluscs. Yet, there have been very few studies examining the molecular machinary underlying eye function of this group, which is due, in part, to a lack of genomic resources. The scallop (Bivalvia: Pectinidae) represents a compeling molluscan model to study photoreception due to its morphologically novel and separately evolved mirror-type eye. We sequenced the adult eye transcriptome of two scallop species to: 1) identify the phototransduction pathway components; 2) identify any additional light detection functions; and 3) test the hypothesis that molluscs possess genes not found in other animal lineages. Results A total of 3,039 contigs from the bay scallop, Argopecten irradians and 26,395 contigs from the sea scallop, Placopecten magellanicus were produced by 454 sequencing. Targeted BLAST searches and functional annotation using Gene Ontology (GO) terms and KEGG pathways identified transcripts from three light detection systems: two phototransduction pathways and the circadian clock, a previously unrecognized function of the scallop eye. By comparing the scallop transcriptomes to molluscan and non-molluscan genomes, we discovered that a large proportion of the transcripts (7,776 sequences) may be specific to the scallop lineage. Nearly one-third of these contain transmembrane protein domains, suggesting these unannotated transcripts may be sensory receptors. Conclusions Our data provide the most comprehensive transcriptomic resource currently available from a single molluscan eye type. Candidate genes potentially involved in sensory reception were identified, and are worthy of further investigation. This resource, combined with recent phylogenetic and genomic data, provides a strong foundation for future investigations of the function and evolution of molluscan photosensory systems in this morphologically and taxonomically diverse phylum. PMID:23922823

Deregulation of SYCP2 predicts early stage human papillomavirus-positive oropharyngeal carcinoma: A prospective whole transcriptome analysis.

PubMed

Masterson, Liam; Sorgeloos, Frederic; Winder, David; Lechner, Matt; Marker, Alison; Malhotra, Shalini; Sudhoff, Holger; Jani, Piyush; Goon, Peter; Sterling, Jane

2015-11-01

This study was designed to identify significant differences in gene expression profiles of human papillomavirus (HPV)-positive and HPV-negative oropharyngeal squamous cell carcinomas (OPSCC) and to better understand the functional and biological effects of HPV infection in the premalignant pathway. Twenty-four consecutive patients with locally advanced primary OPSCC were included in a prospective clinical trial. Fresh tissue samples (tumor vs. matched normal epithelium) were subjected to whole transcriptome analysis and the results validated on the same cohort with RT-quantitative real-time PCR. In a separate retrospective cohort of 27 OPSCC patients, laser capture microdissection of formalin-fixed, paraffin-embedded tissue allowed RNA extraction from adjacent regions of normal epithelium, carcinoma in situ (premalignant) and invasive SCC tissue. The majority of patients showed evidence of high-risk HPV16 positivity (80.4%). Predictable fold changes of RNA expression in HPV-associated disease included multiple transcripts within the p53 oncogenic pathway (e.g. CDKN2A/CCND1). Other candidate transcripts found to have altered levels of expression in this study have not previously been established (SFRP1, CRCT1, DLG2, SYCP2, and CRNN). Of these, SYCP2 showed the most consistent fold change from baseline in premalignant tissue; aberrant expression of this protein may contribute to genetic instability during HPV-associated cancer development. If further corroborated, this data may contribute to the development of a non-invasive screening tool. This study is registered with the UK Clinical Research Network (ref.: 11945). © 2015 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

Developmental Transcriptomics of the Hawaiian Anchialine Shrimp Halocaridina rubra Holthuis, 1963 (Crustacea: Atyidae).

PubMed

Havird, Justin C; Santos, Scott R

2016-12-01

Many crustacean species progress through a series of metamorphoses during the developmental transition from embryo to adult. The molecular genetic basis of this transition, however, is not well characterized for a large number of crustaceans. Here, we employ multiple RNA-Seq methodologies to identify differentially expressed genes (DEGs) between "early" (i.e., Z 1 - Z 2 ) as well as "late" (i.e., Z 3 - Z 4 ) larval and adult developmental stages of Halocaridina rubra Holthuis (1963), an atyid shrimp endemic to the environmentally variable anchialine ecosystem of the Hawaiian Islands. Given the differences in salinity tolerance (narrow vs. wide range), energy acquisition (maternal yolk-bearing vs. microphagous grazing), and behavior (positively phototactic vs. not) between larvae and adults, respectively, of this species, we hypothesized the recovery of numerous DEGs belonging to functional categories relating to these characteristics. Consistent with this and regardless of methodology, hundreds of DEGs were identified, including upregulation of opsins and other light/stimulus detection genes and downregulation of genes related to ion transport, digestion, and reproduction in larvae relative to adults. Furthermore, isoform-switching, which has been largely unexplored in crustacean development, appears to be pervasive between H. rubra larvae and adults, especially among structural and oxygen-transport genes. Finally, by comparing RNA-Seq methodologies, we provide recommendations for future crustacean transcriptomic studies, including a demonstration of the pitfalls associated with identifying DEGs from single replicate samples as well as the utility of leveraging "prepackaged" bioinformatics pipelines. © The Author 2016. Published by Oxford University Press on behalf of the Society for Integrative and Comparative Biology. All rights reserved. For permissions please email: journals.permissions@oup.com.

MicroRNA-31 is a positive modulator of endothelial-mesenchymal transition and associated secretory phenotype induced by TGF-β.

PubMed

Katsura, Akihiro; Suzuki, Hiroshi I; Ueno, Toshihide; Mihira, Hajime; Yamazaki, Tomoko; Yasuda, Takahiko; Watabe, Tetsuro; Mano, Hiroyuki; Yamada, Yoshitsugu; Miyazono, Kohei

2016-01-01

Transforming growth factor-β (TGF-β) plays central roles in endothelial-mesenchymal transition (EndMT) involved in development and pathogenesis. Although EndMT and epithelial-mesenchymal transition are similar processes, roles of microRNAs in EndMT are largely unknown. Here, we report that constitutively active microRNA-31 (miR-31) is a positive regulator of TGF-β-induced EndMT. Although the expression is not induced by TGF-β, miR-31 is required for induction of mesenchymal genes including α-SMA, actin reorganization and MRTF-A activation during EndMT. We identified VAV3, a regulator of actin remodeling and MRTF-A activity, as a miR-31 target. Global transcriptome analysis further showed that miR-31 positively regulates EndMT-associated unique secretory phenotype (EndMT-SP) characterized by induction of multiple inflammatory chemokines and cytokines including CCL17, CX3CL1, CXCL16, IL-6 and Angptl2. As a mechanism for this phenomenon, TGF-β and miR-31 suppress Stk40, a negative regulator of NF-κB pathway. Interestingly, TGF-β induces alternative polyadenylation (APA)-coupled miR-31-dependent Stk40 suppression without concomitant miR-31 induction, and APA-mediated exclusion of internal poly(A) sequence in Stk40 3'UTR enhances target efficiency of Stk40. Finally, miR-31 functions as a molecular hub to integrate TGF-β and TNF-α signaling to enhance EndMT. These data confirm that constitutively active microRNAs, as well as inducible microRNAs, serve as phenotypic modifiers interconnected with transcriptome dynamics during EndMT. © 2015 The Molecular Biology Society of Japan and Wiley Publishing Asia Pty Ltd.

Personalized oncogenomic analysis of metastatic adenoid cystic carcinoma: using whole-genome sequencing to inform clinical decision-making

PubMed Central

Chahal, Manik; Pleasance, Erin; Grewal, Jasleen; Zhao, Eric; Ng, Tony; Chapman, Erin; Jones, Martin R.; Shen, Yaoqing; Mungall, Karen L.; Bonakdar, Melika; Taylor, Gregory A.; Ma, Yussanne; Mungall, Andrew J.; Moore, Richard A.; Lim, Howard; Renouf, Daniel; Yip, Stephen; Jones, Steven J.M.; Marra, Marco A.; Laskin, Janessa

2018-01-01

Metastatic adenoid cystic carcinomas (ACCs) can cause significant morbidity and mortality. Because of their slow growth and relative rarity, there is limited evidence for systemic therapy regimens. Recently, molecular profiling studies have begun to reveal the genetic landscape of these poorly understood cancers, and new treatment possibilities are beginning to emerge. The objective is to use whole-genome and transcriptome sequencing and analysis to better understand the genetic alterations underlying the pathology of metastatic and rare ACCs and determine potentially actionable therapeutic targets. We report five cases of metastatic ACC, not originating in the salivary glands, in patients enrolled in the Personalized Oncogenomics (POG) Program at the BC Cancer Agency. Genomic workup included whole-genome and transcriptome sequencing, detailed analysis of tumor alterations, and integration with existing knowledge of drug–target combinations to identify potential therapeutic targets. Analysis reveals low mutational burden in these five ACC cases, and mutation signatures that are commonly observed in multiple cancer types. Notably, the only recurrent structural aberration identified was the well-described MYB-NFIB fusion that was present in four of five cases, and one case exhibited a closely related MYBL1-NFIB fusion. Recurrent mutations were also identified in BAP1 and BCOR, with additional mutations in individual samples affecting NOTCH1 and the epigenetic regulators ARID2, SMARCA2, and SMARCB1. Copy changes were rare, and they included amplification of MYC and homozygous loss of CDKN2A in individual samples. Genomic analysis revealed therapeutic targets in all five cases and served to inform a therapeutic choice in three of the cases to date. PMID:29610392

Genome-wide Annotation, Identification, and Global Transcriptomic Analysis of Regulatory or Small RNA Gene Expression in Staphylococcus aureus.

PubMed

Carroll, Ronan K; Weiss, Andy; Broach, William H; Wiemels, Richard E; Mogen, Austin B; Rice, Kelly C; Shaw, Lindsey N

2016-02-09

In Staphylococcus aureus, hundreds of small regulatory or small RNAs (sRNAs) have been identified, yet this class of molecule remains poorly understood and severely understudied. sRNA genes are typically absent from genome annotation files, and as a consequence, their existence is often overlooked, particularly in global transcriptomic studies. To facilitate improved detection and analysis of sRNAs in S. aureus, we generated updated GenBank files for three commonly used S. aureus strains (MRSA252, NCTC 8325, and USA300), in which we added annotations for >260 previously identified sRNAs. These files, the first to include genome-wide annotation of sRNAs in S. aureus, were then used as a foundation to identify novel sRNAs in the community-associated methicillin-resistant strain USA300. This analysis led to the discovery of 39 previously unidentified sRNAs. Investigating the genomic loci of the newly identified sRNAs revealed a surprising degree of inconsistency in genome annotation in S. aureus, which may be hindering the analysis and functional exploration of these elements. Finally, using our newly created annotation files as a reference, we perform a global analysis of sRNA gene expression in S. aureus and demonstrate that the newly identified tsr25 is the most highly upregulated sRNA in human serum. This study provides an invaluable resource to the S. aureus research community in the form of our newly generated annotation files, while at the same time presenting the first examination of differential sRNA expression in pathophysiologically relevant conditions. Despite a large number of studies identifying regulatory or small RNA (sRNA) genes in Staphylococcus aureus, their annotation is notably lacking in available genome files. In addition to this, there has been a considerable lack of cross-referencing in the wealth of studies identifying these elements, often leading to the same sRNA being identified multiple times and bearing multiple names. In this work, we have consolidated and curated known sRNA genes from the literature and mapped them to their position on the S. aureus genome, creating new genome annotation files. These files can now be used by the scientific community at large in experiments to search for previously undiscovered sRNA genes and to monitor sRNA gene expression by transcriptome sequencing (RNA-seq). We demonstrate this application, identifying 39 new sRNAs and studying their expression during S. aureus growth in human serum. Copyright © 2016 Carroll et al.

MetaKTSP: a meta-analytic top scoring pair method for robust cross-study validation of omics prediction analysis.

PubMed

Kim, SungHwan; Lin, Chien-Wei; Tseng, George C

2016-07-01

Supervised machine learning is widely applied to transcriptomic data to predict disease diagnosis, prognosis or survival. Robust and interpretable classifiers with high accuracy are usually favored for their clinical and translational potential. The top scoring pair (TSP) algorithm is an example that applies a simple rank-based algorithm to identify rank-altered gene pairs for classifier construction. Although many classification methods perform well in cross-validation of single expression profile, the performance usually greatly reduces in cross-study validation (i.e. the prediction model is established in the training study and applied to an independent test study) for all machine learning methods, including TSP. The failure of cross-study validation has largely diminished the potential translational and clinical values of the models. The purpose of this article is to develop a meta-analytic top scoring pair (MetaKTSP) framework that combines multiple transcriptomic studies and generates a robust prediction model applicable to independent test studies. We proposed two frameworks, by averaging TSP scores or by combining P-values from individual studies, to select the top gene pairs for model construction. We applied the proposed methods in simulated data sets and three large-scale real applications in breast cancer, idiopathic pulmonary fibrosis and pan-cancer methylation. The result showed superior performance of cross-study validation accuracy and biomarker selection for the new meta-analytic framework. In conclusion, combining multiple omics data sets in the public domain increases robustness and accuracy of the classification model that will ultimately improve disease understanding and clinical treatment decisions to benefit patients. An R package MetaKTSP is available online. (http://tsenglab.biostat.pitt.edu/software.htm). ctseng@pitt.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.

Circadian Enhancers Coordinate Multiple Phases of Rhythmic Gene Transcription In Vivo

PubMed Central

Fang, Bin; Everett, Logan J.; Jager, Jennifer; Briggs, Erika; Armour, Sean M.; Feng, Dan; Roy, Ankur; Gerhart-Hines, Zachary; Sun, Zheng; Lazar, Mitchell A.

2014-01-01

SUMMARY Mammalian transcriptomes display complex circadian rhythms with multiple phases of gene expression that cannot be accounted for by current models of the molecular clock. We have determined the underlying mechanisms by measuring nascent RNA transcription around the clock in mouse liver. Unbiased examination of eRNAs that cluster in specific circadian phases identified functional enhancers driven by distinct transcription factors (TFs). We further identify on a global scale the components of the TF cistromes that function to orchestrate circadian gene expression. Integrated genomic analyses also revealed novel mechanisms by which a single circadian factor controls opposing transcriptional phases. These findings shed new light on the diversity and specificity of TF function in the generation of multiple phases of circadian gene transcription in a mammalian organ. PMID:25416951

Circadian enhancers coordinate multiple phases of rhythmic gene transcription in vivo.

PubMed

Fang, Bin; Everett, Logan J; Jager, Jennifer; Briggs, Erika; Armour, Sean M; Feng, Dan; Roy, Ankur; Gerhart-Hines, Zachary; Sun, Zheng; Lazar, Mitchell A

2014-11-20

Mammalian transcriptomes display complex circadian rhythms with multiple phases of gene expression that cannot be accounted for by current models of the molecular clock. We have determined the underlying mechanisms by measuring nascent RNA transcription around the clock in mouse liver. Unbiased examination of enhancer RNAs (eRNAs) that cluster in specific circadian phases identified functional enhancers driven by distinct transcription factors (TFs). We further identify on a global scale the components of the TF cistromes that function to orchestrate circadian gene expression. Integrated genomic analyses also revealed mechanisms by which a single circadian factor controls opposing transcriptional phases. These findings shed light on the diversity and specificity of TF function in the generation of multiple phases of circadian gene transcription in a mammalian organ.

Consensus-phenotype integration of transcriptomic and metabolomic data implies a role for metabolism in the chemosensitivity of tumour cells.

PubMed

Cavill, Rachel; Kamburov, Atanas; Ellis, James K; Athersuch, Toby J; Blagrove, Marcus S C; Herwig, Ralf; Ebbels, Timothy M D; Keun, Hector C

2011-03-01

Using transcriptomic and metabolomic measurements from the NCI60 cell line panel, together with a novel approach to integration of molecular profile data, we show that the biochemical pathways associated with tumour cell chemosensitivity to platinum-based drugs are highly coincident, i.e. they describe a consensus phenotype. Direct integration of metabolome and transcriptome data at the point of pathway analysis improved the detection of consensus pathways by 76%, and revealed associations between platinum sensitivity and several metabolic pathways that were not visible from transcriptome analysis alone. These pathways included the TCA cycle and pyruvate metabolism, lipoprotein uptake and nucleotide synthesis by both salvage and de novo pathways. Extending the approach across a wide panel of chemotherapeutics, we confirmed the specificity of the metabolic pathway associations to platinum sensitivity. We conclude that metabolic phenotyping could play a role in predicting response to platinum chemotherapy and that consensus-phenotype integration of molecular profiling data is a powerful and versatile tool for both biomarker discovery and for exploring the complex relationships between biological pathways and drug response.

A synthesis of transcriptomic surveys to dissect the genetic basis of C 4 photosynthesis

DOE PAGES

Huang, Pu; Brutnell, Thomas P.

2016-04-11

C 4 photosynthesis is used by only three percent of all flowering plants, but explains a quarter of global primary production, including some of the worlds’ most important cereals and bioenergy grasses. Recent advances in our understanding of C 4 development can be attributed to the application of comparative transcriptomics approaches that has been fueled by high throughput sequencing. Global surveys of gene expression conducted between different developmental stages or on phylogenetically closely related C 3 and C 4 species are providing new insights into C 4 function, development and evolution. Importantly, through co-expression analysis and comparative genomics, these studiesmore » help define novel candidate genes that transcend traditional genetic screens. In this review, we briefly summarize the major findings from recent transcriptomic studies, compare and contrast these studies to summarize emerging consensus, and suggest new approaches to exploit the data. Lastly, we suggest using Setaria viridis as a model system to relieve a major bottleneck in genetic studies of C 4 photosynthesis, and discuss the challenges and new opportunities for future comparative transcriptomic studies.« less

Comparative transcriptome analysis by RNAseq of necrotic enteritis Clostridium perfringens during in vivo colonization and in vitro conditions.

PubMed

Parreira, Valeria R; Russell, Kay; Athanasiadou, Spiridoula; Prescott, John F

2016-08-12

Necrotic enteritis (NE) caused by netB-positive type A Clostridium perfringens is an important bacterial disease of poultry. Through its complex regulatory system, C. perfringens orchestrates the expression of a collection of toxins and extracellular enzymes that are crucial for the development of the disease; environmental conditions play an important role in their regulation. In this study, and for the first time, global transcriptomic analysis was performed on ligated intestinal loops in chickens colonized with a netB-positive C. perfringens strain, as well as the same strain propagated in vitro under various nutritional and environmental conditions. Analysis of the respective pathogen transcriptomes revealed up to 673 genes that were significantly expressed in vivo. Gene expression profiles in vivo were most similar to those of C. perfringens grown in nutritionally-deprived conditions. Taken together, our results suggest a bacterial transcriptome responses to the early stages of adaptation, and colonization of, the chicken intestine. Our work also reveals how netB-positive C. perfringens reacts to different environmental conditions including those in the chicken intestine.

First Insights into the Subterranean Crustacean Bathynellacea Transcriptome: Transcriptionally Reduced Opsin Repertoire and Evidence of Conserved Homeostasis Regulatory Mechanisms

PubMed Central

Kim, Bo-Mi; Kang, Seunghyun; Ahn, Do-Hwan; Kim, Jin-Hyoung; Ahn, Inhye; Lee, Chi-Woo; Cho, Joo-Lae; Min, Gi-Sik; Park, Hyun

2017-01-01

Bathynellacea (Crustacea, Syncarida, Parabathynellidae) are subterranean aquatic crustaceans that typically inhabit freshwater interstitial spaces (e.g., groundwater) and are occasionally found in caves and even hot springs. In this study, we sequenced the whole transcriptome of Allobathynella bangokensis using RNA-seq. De novo sequence assembly produced 74,866 contigs including 28,934 BLAST hits. Overall, the gene sequences were most similar to those of the waterflea Daphnia pulex. In the A. bangokensis transcriptome, no opsin or related sequences were identified, and no contig aligned to the crustacean visual opsins and non-visual opsins (i.e. arthropsins, peropsins, and melaopsins), suggesting potential regressive adaptation to the dark environment. However, A. bangokensis expressed conserved gene family sets, such as heat shock proteins and those related to key innate immunity pathways and antioxidant defense systems, at the transcriptional level, suggesting that this species has evolved adaptations involving molecular mechanisms of homeostasis. The transcriptomic information of A. bangokensis will be useful for investigating molecular adaptations and response mechanisms to subterranean environmental conditions. PMID:28107438

Comparative genomics reveals conservative evolution of the xylem transcriptome in vascular plants.

PubMed

Li, Xinguo; Wu, Harry X; Southerton, Simon G

2010-06-21

Wood is a valuable natural resource and a major carbon sink. Wood formation is an important developmental process in vascular plants which played a crucial role in plant evolution. Although genes involved in xylem formation have been investigated, the molecular mechanisms of xylem evolution are not well understood. We use comparative genomics to examine evolution of the xylem transcriptome to gain insights into xylem evolution. The xylem transcriptome is highly conserved in conifers, but considerably divergent in angiosperms. The functional domains of genes in the xylem transcriptome are moderately to highly conserved in vascular plants, suggesting the existence of a common ancestral xylem transcriptome. Compared to the total transcriptome derived from a range of tissues, the xylem transcriptome is relatively conserved in vascular plants. Of the xylem transcriptome, cell wall genes, ancestral xylem genes, known proteins and transcription factors are relatively more conserved in vascular plants. A total of 527 putative xylem orthologs were identified, which are unevenly distributed across the Arabidopsis chromosomes with eight hot spots observed. Phylogenetic analysis revealed that evolution of the xylem transcriptome has paralleled plant evolution. We also identified 274 conifer-specific xylem unigenes, all of which are of unknown function. These xylem orthologs and conifer-specific unigenes are likely to have played a crucial role in xylem evolution. Conifers have highly conserved xylem transcriptomes, while angiosperm xylem transcriptomes are relatively diversified. Vascular plants share a common ancestral xylem transcriptome. The xylem transcriptomes of vascular plants are more conserved than the total transcriptomes. Evolution of the xylem transcriptome has largely followed the trend of plant evolution.

Comparative genomics reveals conservative evolution of the xylem transcriptome in vascular plants

PubMed Central

2010-01-01

Background Wood is a valuable natural resource and a major carbon sink. Wood formation is an important developmental process in vascular plants which played a crucial role in plant evolution. Although genes involved in xylem formation have been investigated, the molecular mechanisms of xylem evolution are not well understood. We use comparative genomics to examine evolution of the xylem transcriptome to gain insights into xylem evolution. Results The xylem transcriptome is highly conserved in conifers, but considerably divergent in angiosperms. The functional domains of genes in the xylem transcriptome are moderately to highly conserved in vascular plants, suggesting the existence of a common ancestral xylem transcriptome. Compared to the total transcriptome derived from a range of tissues, the xylem transcriptome is relatively conserved in vascular plants. Of the xylem transcriptome, cell wall genes, ancestral xylem genes, known proteins and transcription factors are relatively more conserved in vascular plants. A total of 527 putative xylem orthologs were identified, which are unevenly distributed across the Arabidopsis chromosomes with eight hot spots observed. Phylogenetic analysis revealed that evolution of the xylem transcriptome has paralleled plant evolution. We also identified 274 conifer-specific xylem unigenes, all of which are of unknown function. These xylem orthologs and conifer-specific unigenes are likely to have played a crucial role in xylem evolution. Conclusions Conifers have highly conserved xylem transcriptomes, while angiosperm xylem transcriptomes are relatively diversified. Vascular plants share a common ancestral xylem transcriptome. The xylem transcriptomes of vascular plants are more conserved than the total transcriptomes. Evolution of the xylem transcriptome has largely followed the trend of plant evolution. PMID:20565927

Microbiome and ecotypic adaption of Holcus lanatus (L.) to extremes of its soil pH range, investigated through transcriptome sequencing.

PubMed

Young, Ellen; Carey, Manus; Meharg, Andrew A; Meharg, Caroline

2018-03-20

Plants can adapt to edaphic stress, such as nutrient deficiency, toxicity and biotic challenges, by controlled transcriptomic responses, including microbiome interactions. Traditionally studied in model plant species with controlled microbiota inoculation treatments, molecular plant-microbiome interactions can be functionally investigated via RNA-Seq. Complex, natural plant-microbiome studies are limited, typically focusing on microbial rRNA and omitting functional microbiome investigations, presenting a fundamental knowledge gap. Here, root and shoot meta-transcriptome analyses, in tandem with shoot elemental content and root staining, were employed to investigate transcriptome responses in the wild grass Holcus lanatus and its associated natural multi-species eukaryotic microbiome. A full factorial reciprocal soil transplant experiment was employed, using plant ecotypes from two widely contrasting natural habitats, acid bog and limestone quarry soil, to investigate naturally occurring, and ecologically meaningful, edaphically driven molecular plant-microbiome interactions. Arbuscular mycorrhizal (AM) and non-AM fungal colonization was detected in roots in both soils. Staining showed greater levels of non-AM fungi, and transcriptomics indicated a predominance of Ascomycota-annotated genes. Roots in acid bog soil were dominated by Phialocephala-annotated transcripts, a putative growth-promoting endophyte, potentially involved in N nutrition and ion homeostasis. Limestone roots in acid bog soil had greater expression of other Ascomycete genera and Oomycetes and lower expression of Phialocephala-annotated transcripts compared to acid ecotype roots, which corresponded with reduced induction of pathogen defense processes, particularly lignin biosynthesis in limestone ecotypes. Ascomycota dominated in shoots and limestone soil roots, but Phialocephala-annotated transcripts were insignificant, and no single Ascomycete genus dominated. Fusarium-annotated transcripts were the most common genus in shoots, with Colletotrichum and Rhizophagus (AM fungi) most numerous in limestone soil roots. The latter coincided with upregulation of plant genes involved in AM symbiosis initiation and AM-based P acquisition in an environment where P availability is low. Meta-transcriptome analyses provided novel insights into H. lanatus transcriptome responses, associated eukaryotic microbiota functions and taxonomic community composition. Significant edaphic and plant ecotype effects were identified, demonstrating that meta-transcriptome-based functional analysis is a powerful tool for the study of natural plant-microbiome interactions.

Multiple Polyploidization Events across Asteraceae with Two Nested Events in the Early History Revealed by Nuclear Phylogenomics.

PubMed

Huang, Chien-Hsun; Zhang, Caifei; Liu, Mian; Hu, Yi; Gao, Tiangang; Qi, Ji; Ma, Hong

2016-11-01

Biodiversity results from multiple evolutionary mechanisms, including genetic variation and natural selection. Whole-genome duplications (WGDs), or polyploidizations, provide opportunities for large-scale genetic modifications. Many evolutionarily successful lineages, including angiosperms and vertebrates, are ancient polyploids, suggesting that WGDs are a driving force in evolution. However, this hypothesis is challenged by the observed lower speciation and higher extinction rates of recently formed polyploids than diploids. Asteraceae includes about 10% of angiosperm species, is thus undoubtedly one of the most successful lineages and paleopolyploidization was suggested early in this family using a small number of datasets. Here, we used genes from 64 new transcriptome datasets and others to reconstruct a robust Asteraceae phylogeny, covering 73 species from 18 tribes in six subfamilies. We estimated their divergence times and further identified multiple potential ancient WGDs within several tribes and shared by the Heliantheae alliance, core Asteraceae (Asteroideae-Mutisioideae), and also with the sister family Calyceraceae. For two of the WGD events, there were subsequent great increases in biodiversity; the older one proceeded the divergence of at least 10 subfamilies within 10 My, with great variation in morphology and physiology, whereas the other was followed by extremely high species richness in the Heliantheae alliance clade. Our results provide different evidence for several WGDs in Asteraceae and reveal distinct association among WGD events, dramatic changes in environment and species radiations, providing a possible scenario for polyploids to overcome the disadvantages of WGDs and to evolve into lineages with high biodiversity. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

A Comparative Transcriptomic Analysis Reveals Conserved Features of Stem Cell Pluripotency in Planarians and Mammals

PubMed Central

Labbé, Roselyne M.; Irimia, Manuel; Currie, Ko W.; Lin, Alexander; Zhu, Shu Jun; Brown, David D.R.; Ross, Eric J.; Voisin, Veronique; Bader, Gary D.; Blencowe, Benjamin J.; Pearson, Bret J.

2014-01-01

Many long-lived species of animals require the function of adult stem cells throughout their lives. However, the transcriptomes of stem cells in invertebrates and vertebrates have not been compared, and consequently, ancestral regulatory circuits that control stem cell populations remain poorly defined. In this study, we have used data from high-throughput RNA sequencing to compare the transcriptomes of pluripotent adult stem cells from planarians with the transcriptomes of human and mouse pluripotent embryonic stem cells. From a stringently defined set of 4,432 orthologs shared between planarians, mice and humans, we identified 123 conserved genes that are ≥5-fold differentially expressed in stem cells from all three species. Guided by this gene set, we used RNAi screening in adult planarians to discover novel stem cell regulators, which we found to affect the stem cell-associated functions of tissue homeostasis, regeneration, and stem cell maintenance. Examples of genes that disrupted these processes included the orthologs of TBL3, PSD12, TTC27, and RACK1. From these analyses, we concluded that by comparing stem cell transcriptomes from diverse species, it is possible to uncover conserved factors that function in stem cell biology. These results provide insights into which genes comprised the ancestral circuitry underlying the control of stem cell self-renewal and pluripotency. PMID:22696458

Comparative transcriptomics between Synechococcus PCC 7942 and Synechocystis PCC 6803 provide insights into mechanisms of adaptation to stress.

DOE PAGES

Konstantinos, Billis; Billini, Maria; Tripp, Harry J.; ...

2014-09-23

Background: Synechococcus sp. PCC 7942 and Synechocystis sp. PCC 6803 are model cyanobacteria from which the metabolism and adaptive responses of other cyanobacteria are inferred. Here we report the gene expression response of these two strains to a variety of nutrient and environmental stresses of varying duration, using transcriptomics. Our data comprise both stranded and 5' enriched libraries in order to elucidate many aspects of the transcriptome. Results: Both organisms were exposed to stress conditions due to nutrient deficiency (inorganic carbon) or change of environmental conditions (salinity, temperature, pH, light) sampled at 1 and 24 hours after the application ofmore » stress. The transcriptome profile of each strain revealed similarities and differences in gene expression for photosynthetic and respiratory electron transport chains and carbon fixation. Transcriptome profiles also helped us improve the structural annotation of the genome and identify possible missed genes (including anti-sense) and determine transcriptional units (operons). Finally, we predicted association of proteins of unknown function biochemical pathways by associating them to well-characterized ones based on their transcript levels correlation. Conclusions: Overall, this study results an informative annotation of those species and the comparative analysis of the response of the two organisms revealed similarities but also significant changes in the way they respond to external stress and the duration of the response« less

Aging-like Changes in the Transcriptome of Irradiated Microglia

PubMed Central

Li, Matthew D.; Burns, Terry C.; Kumar, Sunny; Morgan, Alexander A.; Sloan, Steven A.; Palmer, Theo D.

2014-01-01

Whole brain irradiation remains important in the management of brain tumors. Although necessary for improving survival outcomes, cranial irradiation also results in cognitive decline in long-term survivors. A chronic inflammatory state characterized by microglial activation has been implicated in radiation-induced brain injury. We here provide the first comprehensive transcriptional profile of irradiated microglia. Fluorescence-activated cell sorting (FACS) was used to isolate CD11b+ microglia from the hippocampi of C57BL/6 and Balb/c mice 1 month after 10Gy cranial irradiation. Affymetrix gene expression profiles were evaluated using linear modeling, rank product analyses. One month after irradiation, a conserved irradiation signature across strains was identified, comprising 448 and 85 differentially up- and down-regulated genes, respectively. Gene set enrichment analysis (GSEA) demonstrated enrichment for inflammation, including M1 macrophage-associated genes, but also an unexpected enrichment for extracellular matrix and blood coagulation-related gene sets, in contrast previously described microglial states. Weighted gene co-expression network analysis (WGCNA) confirmed these findings and further revealed alterations in mitochondrial function. The RNA-seq transcriptome of microglia 24h post-radiation proved similar to the 1-month transcriptome, but additionally featured alterations in apoptotic and lysosomal gene expression. Re-analysis of published aging mouse microglia transcriptome data demonstrated striking similarity to the 1 month irradiated microglia transcriptome, suggesting that shared mechanisms may underlie aging and chronic irradiation-induced cognitive decline. PMID:25690519

Alternative Splicing Profile and Sex-Preferential Gene Expression in the Female and Male Pacific Abalone Haliotis discus hannai.

PubMed

Kim, Mi Ae; Rhee, Jae-Sung; Kim, Tae Ha; Lee, Jung Sick; Choi, Ah-Young; Choi, Beom-Soon; Choi, Ik-Young; Sohn, Young Chang

2017-03-09

In order to characterize the female or male transcriptome of the Pacific abalone and further increase genomic resources, we sequenced the mRNA of full-length complementary DNA (cDNA) libraries derived from pooled tissues of female and male Haliotis discus hannai by employing the Iso-Seq protocol of the PacBio RSII platform. We successfully assembled whole full-length cDNA sequences and constructed a transcriptome database that included isoform information. After clustering, a total of 15,110 and 12,145 genes that coded for proteins were identified in female and male abalones, respectively. A total of 13,057 putative orthologs were retained from each transcriptome in abalones. Overall Gene Ontology terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways analyzed in each database showed a similar composition between sexes. In addition, a total of 519 and 391 isoforms were genome-widely identified with at least two isoforms from female and male transcriptome databases. We found that the number of isoforms and their alternatively spliced patterns are variable and sex-dependent. This information represents the first significant contribution to sex-preferential genomic resources of the Pacific abalone. The availability of whole female and male transcriptome database and their isoform information will be useful to improve our understanding of molecular responses and also for the analysis of population dynamics in the Pacific abalone.

Alternative Splicing Profile and Sex-Preferential Gene Expression in the Female and Male Pacific Abalone Haliotis discus hannai

PubMed Central

Kim, Mi Ae; Rhee, Jae-Sung; Kim, Tae Ha; Lee, Jung Sick; Choi, Ah-Young; Choi, Beom-Soon; Choi, Ik-Young; Sohn, Young Chang

2017-01-01

In order to characterize the female or male transcriptome of the Pacific abalone and further increase genomic resources, we sequenced the mRNA of full-length complementary DNA (cDNA) libraries derived from pooled tissues of female and male Haliotis discus hannai by employing the Iso-Seq protocol of the PacBio RSII platform. We successfully assembled whole full-length cDNA sequences and constructed a transcriptome database that included isoform information. After clustering, a total of 15,110 and 12,145 genes that coded for proteins were identified in female and male abalones, respectively. A total of 13,057 putative orthologs were retained from each transcriptome in abalones. Overall Gene Ontology terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways analyzed in each database showed a similar composition between sexes. In addition, a total of 519 and 391 isoforms were genome-widely identified with at least two isoforms from female and male transcriptome databases. We found that the number of isoforms and their alternatively spliced patterns are variable and sex-dependent. This information represents the first significant contribution to sex-preferential genomic resources of the Pacific abalone. The availability of whole female and male transcriptome database and their isoform information will be useful to improve our understanding of molecular responses and also for the analysis of population dynamics in the Pacific abalone. PMID:28282934

Transcriptome of intraperitoneal organs of starry flounder Platichthys stellatus challenged by Edwardsiella ictaluri JCM1680

NASA Astrophysics Data System (ADS)

Tong, Yanli; Sun, Xiuqin; Wang, Bo; Wang, Ling; Li, Yan; Tian, Jinhu; Zheng, Fengrong; Zheng, Minggang

2015-01-01

Platichthys stellatus is an economically important marine bony fish species that is cultured in China on a large scale. However, very little is known about its immune-related genes. In this study, the transcriptome of the immune organs of P. stellatus that were intraperitoneally challenged with the pathogen E dwardsiella ictaluri JCM1680 is analyzed. Total RNA from four tissues (spleen, kidney, liver, and intestine) was mixed equally and then sequenced on an Illumina HiSeq 2000 platform. Overall, 28 465 813 quality reads were generated and assembled into 43 061 unigenes. Similarity searches against public protein sequence databases were used to annotate 28 291 unigenes (65.7% of the total), 368 of which were associated with immunoregulation, including 188 related to immunity response. Additionally, the transcript levels of immunity response unigenes annotated as related to tumor necrosis factor (TNF), TNF receptor, chemokine, major histocompatibility complex, and interleukin-6 were investigated in the different tissues of normal and infected P. stellatus by real-time quantitative PCR. The results confirmed that the unigenes identified in the transcriptome database were indeed expressed and up-regulated in infected P. stellatus. To our knowledge, this is the first report of the sequencing and analysis of the transcriptome of P. stellatus. These findings provide insights into the transcriptomics and immunogenetics of bony fish.

Novel transcriptome resources for three scleractinian coral species from the Indo-Pacific

PubMed Central

Kenkel, Carly D.; Bay, Line K

2017-01-01

Abstract Transcriptomic resources for coral species can provide insight into coral evolutionary history and stress-response physiology. Goniopora columna, Galaxea astreata, and Galaxea acrhelia are scleractinian corals of the Indo-Pacific, representing a diversity of morphologies and life-history traits. G. columna and G. astreata are common and cosmopolitan, while G. acrhelia is largely restricted to the coral triangle and Great Barrier Reef. Reference transcriptomes for these species were assembled from replicate colony fragments exposed to elevated (31°C) and ambient (27°C) temperatures. Trinity was used to create de novo assemblies for each species from 92–102 million raw Illumina Hiseq 2 × 150 bp reads. Host-specific assemblies contained 65 460–72 405 contigs, representing 26 693–37 894 isogroups (∼genes) with an average N50 of 2254. Gene name and/or gene ontology annotations were possible for 58% of isogroups on average. Transcriptomes contained 93.1–94.3% of EuKaryotic Orthologous Groups comprising the core eukaryotic gene set, and 89.98–91.92% of the single-copy metazoan core gene set orthologs were complete, indicating fairly comprehensive assemblies. This work expands the complement of transcriptomic resources available for scleractinian coral species, including the first reference for a representative of Goniopora spp. as well as species with novel morphology. PMID:28938722

Novel transcriptome resources for three scleractinian coral species from the Indo-Pacific.

PubMed

Kenkel, Carly D; Bay, Line K

2017-09-01

Transcriptomic resources for coral species can provide insight into coral evolutionary history and stress-response physiology. Goniopora columna, Galaxea astreata, and Galaxea acrhelia are scleractinian corals of the Indo-Pacific, representing a diversity of morphologies and life-history traits. G. columna and G. astreata are common and cosmopolitan, while G. acrhelia is largely restricted to the coral triangle and Great Barrier Reef. Reference transcriptomes for these species were assembled from replicate colony fragments exposed to elevated (31°C) and ambient (27°C) temperatures. Trinity was used to create de novo assemblies for each species from 92-102 million raw Illumina Hiseq 2 × 150 bp reads. Host-specific assemblies contained 65 460-72 405 contigs, representing 26 693-37 894 isogroups (∼genes) with an average N50 of 2254. Gene name and/or gene ontology annotations were possible for 58% of isogroups on average. Transcriptomes contained 93.1-94.3% of EuKaryotic Orthologous Groups comprising the core eukaryotic gene set, and 89.98-91.92% of the single-copy metazoan core gene set orthologs were complete, indicating fairly comprehensive assemblies. This work expands the complement of transcriptomic resources available for scleractinian coral species, including the first reference for a representative of Goniopora spp. as well as species with novel morphology. © The Authors 2017. Published by Oxford University Press.

Comparative transcriptomics between Synechococcus PCC 7942 and Synechocystis PCC 6803 provide insights into mechanisms of adaptation to stress.

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

Konstantinos, Billis; Billini, Maria; Tripp, Harry J.

Background: Synechococcus sp. PCC 7942 and Synechocystis sp. PCC 6803 are model cyanobacteria from which the metabolism and adaptive responses of other cyanobacteria are inferred. Here we report the gene expression response of these two strains to a variety of nutrient and environmental stresses of varying duration, using transcriptomics. Our data comprise both stranded and 5' enriched libraries in order to elucidate many aspects of the transcriptome. Results: Both organisms were exposed to stress conditions due to nutrient deficiency (inorganic carbon) or change of environmental conditions (salinity, temperature, pH, light) sampled at 1 and 24 hours after the application ofmore » stress. The transcriptome profile of each strain revealed similarities and differences in gene expression for photosynthetic and respiratory electron transport chains and carbon fixation. Transcriptome profiles also helped us improve the structural annotation of the genome and identify possible missed genes (including anti-sense) and determine transcriptional units (operons). Finally, we predicted association of proteins of unknown function biochemical pathways by associating them to well-characterized ones based on their transcript levels correlation. Conclusions: Overall, this study results an informative annotation of those species and the comparative analysis of the response of the two organisms revealed similarities but also significant changes in the way they respond to external stress and the duration of the response« less

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

PubMed

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

2015-04-22

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

Transcriptome profiling of the floral buds and discovery of genes related to sex-differentiation in the dioecious cucurbit Coccinia grandis (L.) Voigt.

PubMed

Mohanty, Jatindra Nath; Nayak, Sanghamitra; Jha, Sumita; Joshi, Raj Kumar

2017-08-30

Dioecious species offer an inclusive structure to study the molecular basis of sexual dimorphism in angiosperms. Despite having a small genome and heteromorphic sex chromosomes, Coccinia grandis is a highly neglected dioecious species with little information available on its physical state, genetic orientation and key sex-defining elements. In the present study, we performed RNA-Seq and DGE analysis of male (MB) and female (FB) buds in C. grandis to gain insights into the molecular basis of sex determination in this plant. De novo assembly of 75 million clean reads resulted in 72,479 unigenes for male library and 63,308 unigenes for female library with a mean length of 736bp. 61,458 (85.57%) unigenes displayed significant similarity with protein sequences from publicly available databases. Comparative transcriptome analyses revealed 1410 unigenes as differentially expressed (DEGs) between MB and FB samples. A consistent correlation between the expression levels of DEGs was observed for the RNA-Seq pattern and qRT-PCR validation. Functional annotation showed high enrichment of DEGs involved in phytohormone biosynthesis, hormone signaling and transduction, transcriptional regulation and methyltransferase activity. High induction of hormone responsive genes such as ARF6, ACC synthase1, SNRK2 and BRI1-associated receptor kinase 1 (BAK1) suggest that multiple phytohormones and their signaling crosstalk play crucial role in sex determination in this species. Beside, the transcription factors such as zinc fingers, homeodomain leucine zippers and MYBs were identified as major determinants of male specific expression. Moreover, the detection of multiple DEGs as the miRNA target site implies that a small RNA mediated gene silencing cascade may also be regulating gender differentiation in C. grandis. Overall, the present transcriptome resources provide us a large number of DEGs involved in sex expression and could form the groundwork for unravelling the molecular mechanism of sex determination in C. grandis. Copyright © 2017 Elsevier B.V. All rights reserved.

A New Combinatorial Optimization Approach for Integrated Feature Selection Using Different Datasets: A Prostate Cancer Transcriptomic Study

PubMed Central

Puthiyedth, Nisha; Riveros, Carlos; Berretta, Regina; Moscato, Pablo

2015-01-01

Background The joint study of multiple datasets has become a common technique for increasing statistical power in detecting biomarkers obtained from smaller studies. The approach generally followed is based on the fact that as the total number of samples increases, we expect to have greater power to detect associations of interest. This methodology has been applied to genome-wide association and transcriptomic studies due to the availability of datasets in the public domain. While this approach is well established in biostatistics, the introduction of new combinatorial optimization models to address this issue has not been explored in depth. In this study, we introduce a new model for the integration of multiple datasets and we show its application in transcriptomics. Methods We propose a new combinatorial optimization problem that addresses the core issue of biomarker detection in integrated datasets. Optimal solutions for this model deliver a feature selection from a panel of prospective biomarkers. The model we propose is a generalised version of the (α,β)-k-Feature Set problem. We illustrate the performance of this new methodology via a challenging meta-analysis task involving six prostate cancer microarray datasets. The results are then compared to the popular RankProd meta-analysis tool and to what can be obtained by analysing the individual datasets by statistical and combinatorial methods alone. Results Application of the integrated method resulted in a more informative signature than the rank-based meta-analysis or individual dataset results, and overcomes problems arising from real world datasets. The set of genes identified is highly significant in the context of prostate cancer. The method used does not rely on homogenisation or transformation of values to a common scale, and at the same time is able to capture markers associated with subgroups of the disease. PMID:26106884

Use of homologous and heterologous gene expression profiling tools to characterize transcription dynamics during apple fruit maturation and ripening.

PubMed

Costa, Fabrizio; Alba, Rob; Schouten, Henk; Soglio, Valeria; Gianfranceschi, Luca; Serra, Sara; Musacchi, Stefano; Sansavini, Silviero; Costa, Guglielmo; Fei, Zhangjun; Giovannoni, James

2010-10-25

Fruit development, maturation and ripening consists of a complex series of biochemical and physiological changes that in climacteric fruits, including apple and tomato, are coordinated by the gaseous hormone ethylene. These changes lead to final fruit quality and understanding of the functional machinery underlying these processes is of both biological and practical importance. To date many reports have been made on the analysis of gene expression in apple. In this study we focused our investigation on the role of ethylene during apple maturation, specifically comparing transcriptomics of normal ripening with changes resulting from application of the hormone receptor competitor 1-methylcyclopropene. To gain insight into the molecular process regulating ripening in apple, and to compare to tomato (model species for ripening studies), we utilized both homologous and heterologous (tomato) microarray to profile transcriptome dynamics of genes involved in fruit development and ripening, emphasizing those which are ethylene regulated.The use of both types of microarrays facilitated transcriptome comparison between apple and tomato (for the later using data previously published and available at the TED: tomato expression database) and highlighted genes conserved during ripening of both species, which in turn represent a foundation for further comparative genomic studies. The cross-species analysis had the secondary aim of examining the efficiency of heterologous (specifically tomato) microarray hybridization for candidate gene identification as related to the ripening process. The resulting transcriptomics data revealed coordinated gene expression during fruit ripening of a subset of ripening-related and ethylene responsive genes, further facilitating the analysis of ethylene response during fruit maturation and ripening. Our combined strategy based on microarray hybridization enabled transcriptome characterization during normal climacteric apple ripening, as well as definition of ethylene-dependent transcriptome changes. Comparison with tomato fruit maturation and ethylene responsive transcriptome activity facilitated identification of putative conserved orthologous ripening-related genes, which serve as an initial set of candidates for assessing conservation of gene activity across genomes of fruit bearing plant species.

Halvade-RNA: Parallel variant calling from transcriptomic data using MapReduce.

PubMed

Decap, Dries; Reumers, Joke; Herzeel, Charlotte; Costanza, Pascal; Fostier, Jan

2017-01-01

Given the current cost-effectiveness of next-generation sequencing, the amount of DNA-seq and RNA-seq data generated is ever increasing. One of the primary objectives of NGS experiments is calling genetic variants. While highly accurate, most variant calling pipelines are not optimized to run efficiently on large data sets. However, as variant calling in genomic data has become common practice, several methods have been proposed to reduce runtime for DNA-seq analysis through the use of parallel computing. Determining the effectively expressed variants from transcriptomics (RNA-seq) data has only recently become possible, and as such does not yet benefit from efficiently parallelized workflows. We introduce Halvade-RNA, a parallel, multi-node RNA-seq variant calling pipeline based on the GATK Best Practices recommendations. Halvade-RNA makes use of the MapReduce programming model to create and manage parallel data streams on which multiple instances of existing tools such as STAR and GATK operate concurrently. Whereas the single-threaded processing of a typical RNA-seq sample requires ∼28h, Halvade-RNA reduces this runtime to ∼2h using a small cluster with two 20-core machines. Even on a single, multi-core workstation, Halvade-RNA can significantly reduce runtime compared to using multi-threading, thus providing for a more cost-effective processing of RNA-seq data. Halvade-RNA is written in Java and uses the Hadoop MapReduce 2.0 API. It supports a wide range of distributions of Hadoop, including Cloudera and Amazon EMR.

Molecular adaptation in the world's deepest-living animal: Insights from transcriptome sequencing of the hadal amphipod Hirondellea gigas.

PubMed

Lan, Yi; Sun, Jin; Tian, Renmao; Bartlett, Douglas H; Li, Runsheng; Wong, Yue Him; Zhang, Weipeng; Qiu, Jian-Wen; Xu, Ting; He, Li-Sheng; Tabata, Harry G; Qian, Pei-Yuan

2017-07-01

The Challenger Deep in the Mariana Trench is the deepest point in the oceans of our planet. Understanding how animals adapt to this harsh environment characterized by high hydrostatic pressure, food limitation, dark and cold is of great scientific interest. Of the animals dwelling in the Challenger Deep, amphipods have been captured using baited traps. In this study, we sequenced the transcriptome of the amphipod Hirondellea gigas collected at a depth of 10,929 m from the East Pond of the Challenger Deep. Assembly of these sequences resulted in 133,041 contigs and 22,046 translated proteins. Functional annotation of these contigs was made using the go and kegg databases. Comparison of these translated proteins with those of four shallow-water amphipods revealed 10,731 gene families, of which 5659 were single-copy orthologs. Base substitution analysis on these single-copy orthologs showed that 62 genes are positively selected in H. gigas, including genes related to β-alanine biosynthesis, energy metabolism and genetic information processing. For multiple-copy orthologous genes, gene family expansion analysis revealed that cold-inducible proteins (i.e., transcription factors II A and transcription elongation factor 1) as well as zinc finger domains are expanded in H. gigas. Overall, our results indicate that genetic adaptation to the hadal environment by H. gigas may be mediated by both gene family expansion and amino acid substitutions of specific proteins. © 2017 John Wiley & Sons Ltd.

Cooption of heat shock regulatory system for anhydrobiosis in the sleeping chironomid Polypedilum vanderplanki

PubMed Central

Shagimardanova, Elena; Kozlova, Olga; Cherkasov, Alexander; Sutormin, Roman; Stepanova, Vita V.; Stupnikov, Alexey; Logacheva, Maria; Penin, Aleksey; Sogame, Yoichiro; Cornette, Richard; Tokumoto, Shoko; Miyata, Yugo; Gelfand, Mikhail S.; Gusev, Oleg

2018-01-01

Polypedilum vanderplanki is a striking and unique example of an insect that can survive almost complete desiccation. Its genome and a set of dehydration–rehydration transcriptomes, together with the genome of Polypedilum nubifer (a congeneric desiccation-sensitive midge), were recently released. Here, using published and newly generated datasets reflecting detailed transcriptome changes during anhydrobiosis, as well as a developmental series, we show that the TCTAGAA DNA motif, which closely resembles the binding motif of the Drosophila melanogaster heat shock transcription activator (Hsf), is significantly enriched in the promoter regions of desiccation-induced genes in P. vanderplanki, such as genes encoding late embryogenesis abundant (LEA) proteins, thioredoxins, or trehalose metabolism-related genes, but not in P. nubifer. Unlike P. nubifer, P. vanderplanki has double TCTAGAA sites upstream of the Hsf gene itself, which is probably responsible for the stronger activation of Hsf in P. vanderplanki during desiccation compared with P. nubifer. To confirm the role of Hsf in desiccation-induced gene activation, we used the Pv11 cell line, derived from P. vanderplanki embryo. After preincubation with trehalose, Pv11 cells can enter anhydrobiosis and survive desiccation. We showed that Hsf knockdown suppresses trehalose-induced activation of multiple predicted Hsf targets (including P. vanderplanki-specific LEA protein genes) and reduces the desiccation survival rate of Pv11 cells fivefold. Thus, cooption of the heat shock regulatory system has been an important evolutionary mechanism for adaptation to desiccation in P. vanderplanki. PMID:29463761

Comparative Proteomic and Transcriptomic Analysis of Follistatin-Induced Skeletal Muscle Hypertrophy.

PubMed

Barbé, Caroline; Bray, Fabrice; Gueugneau, Marine; Devassine, Stéphanie; Lause, Pascale; Tokarski, Caroline; Rolando, Christian; Thissen, Jean-Paul

2017-10-06

Skeletal muscle, the most abundant body tissue, plays vital roles in locomotion and metabolism. Myostatin is a negative regulator of skeletal muscle mass. In addition to increasing muscle mass, Myostatin inhibition impacts muscle contractility and energy metabolism. To decipher the mechanisms of action of the Myostatin inhibitors, we used proteomic and transcriptomic approaches to investigate the changes induced in skeletal muscles of transgenic mice overexpressing Follistatin, a physiological Myostatin inhibitor. Our proteomic workflow included a fractionation step to identify weakly expressed proteins and a comparison of fast versus slow muscles. Functional annotation of altered proteins supports the phenotypic changes induced by Myostatin inhibition, including modifications in energy metabolism, fiber type, insulin and calcium signaling, as well as membrane repair and regeneration. Less than 10% of the differentially expressed proteins were found to be also regulated at the mRNA level but the Biological Process annotation, and the KEGG pathways analysis of transcriptomic results shows a great concordance with the proteomic data. Thus this study describes the most extensive omics analysis of muscle overexpressing Follistatin, providing molecular-level insights to explain the observed muscle phenotypic changes.

Disturbed Placental Imprinting in Preeclampsia Leads to Altered Expression of DLX5, a Human-Specific Early Trophoblast Marker

PubMed Central

Zadora, Julianna; Singh, Manvendra; Herse, Florian; Przybyl, Lukasz; Haase, Nadine; Golic, Michaela; Yung, Hong Wa; Huppertz, Berthold; Cartwright, Judith E.; Whitley, Guy; Johnsen, Guro M.; Levi, Giovanni; Isbruch, Annette; Schulz, Herbert; Luft, Friedrich C.; Müller, Dominik N.; Staff, Anne Cathrine

2017-01-01

Background: Preeclampsia is a complex and common human-specific pregnancy syndrome associated with placental pathology. The human specificity provides both intellectual and methodological challenges, lacking a robust model system. Given the role of imprinted genes in human placentation and the vulnerability of imprinted genes to loss of imprinting changes, there has been extensive speculation, but no robust evidence, that imprinted genes are involved in preeclampsia. Our study aims to investigate whether disturbed imprinting contributes to preeclampsia. Methods: We first aimed to confirm that preeclampsia is a disease of the placenta by generating and analyzing genome-wide molecular data on well-characterized patient material. We performed high-throughput transcriptome analyses of multiple placenta samples from healthy controls and patients with preeclampsia. Next, we identified differentially expressed genes in preeclamptic placentas and intersected them with the list of human imprinted genes. We used bioinformatics/statistical analyses to confirm association between imprinting and preeclampsia and to predict biological processes affected in preeclampsia. Validation included epigenetic and cellular assays. In terms of human specificity, we established an in vitro invasion-differentiation trophoblast model. Our comparative phylogenetic analysis involved single-cell transcriptome data of human, macaque, and mouse preimplantation embryogenesis. Results: We found disturbed placental imprinting in preeclampsia and revealed potential candidates, including GATA3 and DLX5, with poorly explored imprinted status and no prior association with preeclampsia. As a result of loss of imprinting, DLX5 was upregulated in 69% of preeclamptic placentas. Levels of DLX5 correlated with classic preeclampsia markers. DLX5 is expressed in human but not in murine trophoblast. The DLX5high phenotype resulted in reduced proliferation, increased metabolism, and endoplasmic reticulum stress-response activation in trophoblasts in vitro. The transcriptional profile of such cells mimics the transcriptome of preeclamptic placentas. Pan-mammalian comparative analysis identified DLX5 as part of the human-specific regulatory network of trophoblast differentiation. Conclusions: Our analysis provides evidence of a true association among disturbed imprinting, gene expression, and preeclampsia. As a result of disturbed imprinting, the upregulated DLX5 affects trophoblast proliferation. Our in vitro model might fill a vital niche in preeclampsia research. Human-specific regulatory circuitry of DLX5 might help explain certain aspects of preeclampsia. PMID:28904069

Disturbed Placental Imprinting in Preeclampsia Leads to Altered Expression of DLX5, a Human-Specific Early Trophoblast Marker.

PubMed

Zadora, Julianna; Singh, Manvendra; Herse, Florian; Przybyl, Lukasz; Haase, Nadine; Golic, Michaela; Yung, Hong Wa; Huppertz, Berthold; Cartwright, Judith E; Whitley, Guy; Johnsen, Guro M; Levi, Giovanni; Isbruch, Annette; Schulz, Herbert; Luft, Friedrich C; Müller, Dominik N; Staff, Anne Cathrine; Hurst, Laurence D; Dechend, Ralf; Izsvák, Zsuzsanna

2017-11-07

Preeclampsia is a complex and common human-specific pregnancy syndrome associated with placental pathology. The human specificity provides both intellectual and methodological challenges, lacking a robust model system. Given the role of imprinted genes in human placentation and the vulnerability of imprinted genes to loss of imprinting changes, there has been extensive speculation, but no robust evidence, that imprinted genes are involved in preeclampsia. Our study aims to investigate whether disturbed imprinting contributes to preeclampsia. We first aimed to confirm that preeclampsia is a disease of the placenta by generating and analyzing genome-wide molecular data on well-characterized patient material. We performed high-throughput transcriptome analyses of multiple placenta samples from healthy controls and patients with preeclampsia. Next, we identified differentially expressed genes in preeclamptic placentas and intersected them with the list of human imprinted genes. We used bioinformatics/statistical analyses to confirm association between imprinting and preeclampsia and to predict biological processes affected in preeclampsia. Validation included epigenetic and cellular assays. In terms of human specificity, we established an in vitro invasion-differentiation trophoblast model. Our comparative phylogenetic analysis involved single-cell transcriptome data of human, macaque, and mouse preimplantation embryogenesis. We found disturbed placental imprinting in preeclampsia and revealed potential candidates, including GATA3 and DLX5 , with poorly explored imprinted status and no prior association with preeclampsia. As a result of loss of imprinting, DLX5 was upregulated in 69% of preeclamptic placentas. Levels of DLX5 correlated with classic preeclampsia markers. DLX5 is expressed in human but not in murine trophoblast. The DLX5 high phenotype resulted in reduced proliferation, increased metabolism, and endoplasmic reticulum stress-response activation in trophoblasts in vitro. The transcriptional profile of such cells mimics the transcriptome of preeclamptic placentas. Pan-mammalian comparative analysis identified DLX5 as part of the human-specific regulatory network of trophoblast differentiation. Our analysis provides evidence of a true association among disturbed imprinting, gene expression, and preeclampsia. As a result of disturbed imprinting, the upregulated DLX5 affects trophoblast proliferation. Our in vitro model might fill a vital niche in preeclampsia research. Human-specific regulatory circuitry of DLX5 might help explain certain aspects of preeclampsia. © 2017 The Authors.

Exploring the genetic architecture and improving genomic prediction accuracy for mastitis and milk production traits in dairy cattle by mapping variants to hepatic transcriptomic regions responsive to intra-mammary infection.

PubMed

Fang, Lingzhao; Sahana, Goutam; Ma, Peipei; Su, Guosheng; Yu, Ying; Zhang, Shengli; Lund, Mogens Sandø; Sørensen, Peter

2017-05-12

A better understanding of the genetic architecture of complex traits can contribute to improve genomic prediction. We hypothesized that genomic variants associated with mastitis and milk production traits in dairy cattle are enriched in hepatic transcriptomic regions that are responsive to intra-mammary infection (IMI). Genomic markers [e.g. single nucleotide polymorphisms (SNPs)] from those regions, if included, may improve the predictive ability of a genomic model. We applied a genomic feature best linear unbiased prediction model (GFBLUP) to implement the above strategy by considering the hepatic transcriptomic regions responsive to IMI as genomic features. GFBLUP, an extension of GBLUP, includes a separate genomic effect of SNPs within a genomic feature, and allows differential weighting of the individual marker relationships in the prediction equation. Since GFBLUP is computationally intensive, we investigated whether a SNP set test could be a computationally fast way to preselect predictive genomic features. The SNP set test assesses the association between a genomic feature and a trait based on single-SNP genome-wide association studies. We applied these two approaches to mastitis and milk production traits (milk, fat and protein yield) in Holstein (HOL, n = 5056) and Jersey (JER, n = 1231) cattle. We observed that a majority of genomic features were enriched in genomic variants that were associated with mastitis and milk production traits. Compared to GBLUP, the accuracy of genomic prediction with GFBLUP was marginally improved (3.2 to 3.9%) in within-breed prediction. The highest increase (164.4%) in prediction accuracy was observed in across-breed prediction. The significance of genomic features based on the SNP set test were correlated with changes in prediction accuracy of GFBLUP (P < 0.05). GFBLUP provides a framework for integrating multiple layers of biological knowledge to provide novel insights into the biological basis of complex traits, and to improve the accuracy of genomic prediction. The SNP set test might be used as a first-step to improve GFBLUP models. Approaches like GFBLUP and SNP set test will become increasingly useful, as the functional annotations of genomes keep accumulating for a range of species and traits.

VESPA: software to facilitate genomic annotation of prokaryotic organisms through integration of proteomic and transcriptomic data.

PubMed

Peterson, Elena S; McCue, Lee Ann; Schrimpe-Rutledge, Alexandra C; Jensen, Jeffrey L; Walker, Hyunjoo; Kobold, Markus A; Webb, Samantha R; Payne, Samuel H; Ansong, Charles; Adkins, Joshua N; Cannon, William R; Webb-Robertson, Bobbie-Jo M

2012-04-05

The procedural aspects of genome sequencing and assembly have become relatively inexpensive, yet the full, accurate structural annotation of these genomes remains a challenge. Next-generation sequencing transcriptomics (RNA-Seq), global microarrays, and tandem mass spectrometry (MS/MS)-based proteomics have demonstrated immense value to genome curators as individual sources of information, however, integrating these data types to validate and improve structural annotation remains a major challenge. Current visual and statistical analytic tools are focused on a single data type, or existing software tools are retrofitted to analyze new data forms. We present Visual Exploration and Statistics to Promote Annotation (VESPA) is a new interactive visual analysis software tool focused on assisting scientists with the annotation of prokaryotic genomes though the integration of proteomics and transcriptomics data with current genome location coordinates. VESPA is a desktop Java™ application that integrates high-throughput proteomics data (peptide-centric) and transcriptomics (probe or RNA-Seq) data into a genomic context, all of which can be visualized at three levels of genomic resolution. Data is interrogated via searches linked to the genome visualizations to find regions with high likelihood of mis-annotation. Search results are linked to exports for further validation outside of VESPA or potential coding-regions can be analyzed concurrently with the software through interaction with BLAST. VESPA is demonstrated on two use cases (Yersinia pestis Pestoides F and Synechococcus sp. PCC 7002) to demonstrate the rapid manner in which mis-annotations can be found and explored in VESPA using either proteomics data alone, or in combination with transcriptomic data. VESPA is an interactive visual analytics tool that integrates high-throughput data into a genomic context to facilitate the discovery of structural mis-annotations in prokaryotic genomes. Data is evaluated via visual analysis across multiple levels of genomic resolution, linked searches and interaction with existing bioinformatics tools. We highlight the novel functionality of VESPA and core programming requirements for visualization of these large heterogeneous datasets for a client-side application. The software is freely available at https://www.biopilot.org/docs/Software/Vespa.php.

Transcriptome-enabled marker discovery and mapping of plastochron-related genes in Petunia spp.

PubMed

Guo, Yufang; Wiegert-Rininger, Krystle E; Vallejo, Veronica A; Barry, Cornelius S; Warner, Ryan M

2015-09-24

Petunia (Petunia × hybrida), derived from a hybrid between P. axillaris and P. integrifolia, is one of the most economically important bedding plant crops and Petunia spp. serve as model systems for investigating the mechanisms underlying diverse mating systems and pollination syndromes. In addition, we have previously described genetic variation and quantitative trait loci (QTL) related to petunia development rate and morphology, which represent important breeding targets for the floriculture industry to improve crop production and performance. Despite the importance of petunia as a crop, the floriculture industry has been slow to adopt marker assisted selection to facilitate breeding strategies and there remains a limited availability of sequences and molecular markers from the genus compared to other economically important members of the Solanaceae family such as tomato, potato and pepper. Here we report the de novo assembly, annotation and characterization of transcriptomes from P. axillaris, P. exserta and P. integrifolia. Each transcriptome assembly was derived from five tissue libraries (callus, 3-week old seedlings, shoot apices, flowers of mixed developmental stages, and trichomes). A total of 74,573, 54,913, and 104,739 assembled transcripts were recovered from P. axillaris, P. exserta and P. integrifolia, respectively and following removal of multiple isoforms, 32,994 P. axillaris, 30,225 P. exserta, and 33,540 P. integrifolia high quality representative transcripts were extracted for annotation and expression analysis. The transcriptome data was mined for single nucleotide polymorphisms (SNP) and simple sequence repeat (SSR) markers, yielding 89,007 high quality SNPs and 2949 SSRs, respectively. 15,701 SNPs were computationally converted into user-friendly cleaved amplified polymorphic sequence (CAPS) markers and a subset of SNP and CAPS markers were experimentally verified. CAPS markers developed from plastochron-related homologous transcripts from P. axillaris were mapped in an interspecific Petunia population and evaluated for co-localization with QTL for development rate. The high quality of the three Petunia spp. transcriptomes coupled with the utility of the SNP data will serve as a resource for further exploration of genetic diversity within the genus and will facilitate efforts to develop genetic and physical maps to aid the identification of QTL associated with traits of interest.

VESPA: software to facilitate genomic annotation of prokaryotic organisms through integration of proteomic and transcriptomic data

PubMed Central

2012-01-01

Background The procedural aspects of genome sequencing and assembly have become relatively inexpensive, yet the full, accurate structural annotation of these genomes remains a challenge. Next-generation sequencing transcriptomics (RNA-Seq), global microarrays, and tandem mass spectrometry (MS/MS)-based proteomics have demonstrated immense value to genome curators as individual sources of information, however, integrating these data types to validate and improve structural annotation remains a major challenge. Current visual and statistical analytic tools are focused on a single data type, or existing software tools are retrofitted to analyze new data forms. We present Visual Exploration and Statistics to Promote Annotation (VESPA) is a new interactive visual analysis software tool focused on assisting scientists with the annotation of prokaryotic genomes though the integration of proteomics and transcriptomics data with current genome location coordinates. Results VESPA is a desktop Java™ application that integrates high-throughput proteomics data (peptide-centric) and transcriptomics (probe or RNA-Seq) data into a genomic context, all of which can be visualized at three levels of genomic resolution. Data is interrogated via searches linked to the genome visualizations to find regions with high likelihood of mis-annotation. Search results are linked to exports for further validation outside of VESPA or potential coding-regions can be analyzed concurrently with the software through interaction with BLAST. VESPA is demonstrated on two use cases (Yersinia pestis Pestoides F and Synechococcus sp. PCC 7002) to demonstrate the rapid manner in which mis-annotations can be found and explored in VESPA using either proteomics data alone, or in combination with transcriptomic data. Conclusions VESPA is an interactive visual analytics tool that integrates high-throughput data into a genomic context to facilitate the discovery of structural mis-annotations in prokaryotic genomes. Data is evaluated via visual analysis across multiple levels of genomic resolution, linked searches and interaction with existing bioinformatics tools. We highlight the novel functionality of VESPA and core programming requirements for visualization of these large heterogeneous datasets for a client-side application. The software is freely available at https://www.biopilot.org/docs/Software/Vespa.php. PMID:22480257

Antennal Transcriptome Analysis of Odorant Reception Genes in the Red Turpentine Beetle (RTB), Dendroctonus valens.

PubMed

Gu, Xiao-Cui; Zhang, Ya-Nan; Kang, Ke; Dong, Shuang-Lin; Zhang, Long-Wa

2015-01-01

The red turpentine beetle (RTB), Dendroctonus valens LeConte (Coleoptera: Curculionidae, Scolytinae), is a destructive invasive pest of conifers which has become the second most important forest pest nationwide in China. Dendroctonus valens is known to use host odors and aggregation pheromones, as well as non-host volatiles, in host location and mass-attack modulation, and thus antennal olfaction is of the utmost importance for the beetles' survival and fitness. However, information on the genes underlying olfaction has been lacking in D. valens. Here, we report the antennal transcriptome of D. valens from next-generation sequencing, with the goal of identifying the olfaction gene repertoire that is involved in D. valens odor-processing. We obtained 51 million reads that were assembled into 61,889 genes, including 39,831 contigs and 22,058 unigenes. In total, we identified 68 novel putative odorant reception genes, including 21 transcripts encoding for putative odorant binding proteins (OBP), six chemosensory proteins (CSP), four sensory neuron membrane proteins (SNMP), 22 odorant receptors (OR), four gustatory receptors (GR), three ionotropic receptors (IR), and eight ionotropic glutamate receptors. We also identified 155 odorant/xenobiotic degradation enzymes from the antennal transcriptome, putatively identified to be involved in olfaction processes including cytochrome P450s, glutathione-S-transferases, and aldehyde dehydrogenase. Predicted protein sequences were compared with counterparts in Tribolium castaneum, Megacyllene caryae, Ips typographus, Dendroctonus ponderosae, and Agrilus planipennis. The antennal transcriptome described here represents the first study of the repertoire of odor processing genes in D. valens. The genes reported here provide a significant addition to the pool of identified olfactory genes in Coleoptera, which might represent novel targets for insect management. The results from our study also will assist with evolutionary analyses of coleopteran olfaction.

Antennal Transcriptome Analysis of Odorant Reception Genes in the Red Turpentine Beetle (RTB), Dendroctonus valens

PubMed Central

Dong, Shuang-Lin; Zhang, Long-Wa

2015-01-01

Background The red turpentine beetle (RTB), Dendroctonus valens LeConte (Coleoptera: Curculionidae, Scolytinae), is a destructive invasive pest of conifers which has become the second most important forest pest nationwide in China. Dendroctonus valens is known to use host odors and aggregation pheromones, as well as non-host volatiles, in host location and mass-attack modulation, and thus antennal olfaction is of the utmost importance for the beetles’ survival and fitness. However, information on the genes underlying olfaction has been lacking in D. valens. Here, we report the antennal transcriptome of D. valens from next-generation sequencing, with the goal of identifying the olfaction gene repertoire that is involved in D. valens odor-processing. Results We obtained 51 million reads that were assembled into 61,889 genes, including 39,831 contigs and 22,058 unigenes. In total, we identified 68 novel putative odorant reception genes, including 21 transcripts encoding for putative odorant binding proteins (OBP), six chemosensory proteins (CSP), four sensory neuron membrane proteins (SNMP), 22 odorant receptors (OR), four gustatory receptors (GR), three ionotropic receptors (IR), and eight ionotropic glutamate receptors. We also identified 155 odorant/xenobiotic degradation enzymes from the antennal transcriptome, putatively identified to be involved in olfaction processes including cytochrome P450s, glutathione-S-transferases, and aldehyde dehydrogenase. Predicted protein sequences were compared with counterparts in Tribolium castaneum, Megacyllene caryae, Ips typographus, Dendroctonus ponderosae, and Agrilus planipennis. Conclusion The antennal transcriptome described here represents the first study of the repertoire of odor processing genes in D. valens. The genes reported here provide a significant addition to the pool of identified olfactory genes in Coleoptera, which might represent novel targets for insect management. The results from our study also will assist with evolutionary analyses of coleopteran olfaction. PMID:25938508

Effects of temperature on transcriptome and cuticular hydrocarbon expression in ecologically differentiated populations of desert Drosophila.

PubMed

Etges, William J; de Oliveira, Cássia C; Rajpurohit, Subhash; Gibbs, Allen G

2017-01-01

We assessed the effects of temperature differences on gene expression using whole-transcriptome microarrays and cuticular hydrocarbon variation in populations of cactophilic Drosophila mojavensis . Four populations from Baja California and mainland Mexico and Arizona were each reared on two different host cacti, reared to sexual maturity on laboratory media, and adults were exposed for 12 hr to 15, 25, or 35°C. Temperature differences influenced the expression of 3,294 genes, while population differences and host plants affected >2,400 each in adult flies. Enriched, functionally related groups of genes whose expression changed at high temperatures included heat response genes, as well as genes affecting chromatin structure. Gene expression differences between mainland and peninsular populations included genes involved in metabolism of secondary compounds, mitochondrial activity, and tRNA synthases. Flies reared on the ancestral host plant, pitaya agria cactus, showed upregulation of genes involved in metabolism, while flies reared on organ pipe cactus had higher expression of DNA repair and chromatin remodeling genes. Population × environment (G × E) interactions had widespread effects on the transcriptome where population × temperature interactions affected the expression of >5,000 orthologs, and there were >4,000 orthologs that showed temperature × host plant interactions. Adults exposed to 35°C had lower amounts of most cuticular hydrocarbons than those exposed to 15 or 25°C, including abundant unsaturated alkadienes. For insects adapted to different host plants and climatic regimes, our results suggest that temperature shifts associated with climate change have large and significant effects on transcriptomes of genetically differentiated natural populations.

High-throughput sequencing of black pepper root transcriptome.

PubMed

Gordo, Sheila M C; Pinheiro, Daniel G; Moreira, Edith C O; Rodrigues, Simone M; Poltronieri, Marli C; de Lemos, Oriel F; da Silva, Israel Tojal; Ramos, Rommel T J; Silva, Artur; Schneider, Horacio; Silva, Wilson A; Sampaio, Iracilda; Darnet, Sylvain

2012-09-17

Black pepper (Piper nigrum L.) is one of the most popular spices in the world. It is used in cooking and the preservation of food and even has medicinal properties. Losses in production from disease are a major limitation in the culture of this crop. The major diseases are root rot and foot rot, which are results of root infection by Fusarium solani and Phytophtora capsici, respectively. Understanding the molecular interaction between the pathogens and the host's root region is important for obtaining resistant cultivars by biotechnological breeding. Genetic and molecular data for this species, though, are limited. In this paper, RNA-Seq technology has been employed, for the first time, to describe the root transcriptome of black pepper. The root transcriptome of black pepper was sequenced by the NGS SOLiD platform and assembled using the multiple-k method. Blast2Go and orthoMCL methods were used to annotate 10338 unigenes. The 4472 predicted proteins showed about 52% homology with the Arabidopsis proteome. Two root proteomes identified 615 proteins, which seem to define the plant's root pattern. Simple-sequence repeats were identified that may be useful in studies of genetic diversity and may have applications in biotechnology and ecology. This dataset of 10338 unigenes is crucially important for the biotechnological breeding of black pepper and the ecogenomics of the Magnoliids, a major group of basal angiosperms.

Comparative Transcriptomics of Strawberries (Fragaria spp.) Provides Insights into Evolutionary Patterns.

PubMed

Qiao, Qin; Xue, Li; Wang, Qia; Sun, Hang; Zhong, Yang; Huang, Jinling; Lei, Jiajun; Zhang, Ticao

2016-01-01

Multiple closely related species with genomic sequences provide an ideal system for studies on comparative and evolutionary genomics, as well as the mechanism of speciation. The whole genome sequences of six strawberry species ( Fragaria spp.) have been released, which provide one of the richest genomic resources of any plant genus. In this study, we first generated seven transcriptome sequences of Fragaria species de novo , with a total of 48,557-82,537 unigenes per species. Combined with 13 other species genomes in Rosales, we reconstructed a phylogenetic tree at the genomic level. The phylogenic tree shows that Fragaria closed grouped with Rubus and the Fragaria clade is divided into three subclades. East Asian species appeared in every subclade, suggesting that the genus originated in this area at ∼7.99 Mya. Four species found in mountains of Southwest China originated at ∼3.98 Mya, suggesting that rapid speciation occurred to adapt to changing environments following the uplift of the Qinghai-Tibet Plateau. Moreover, we identified 510 very significantly positively selected genes in the cultivated species F . × ananassa genome. This set of genes was enriched in functions related to specific agronomic traits, such as carbon metabolism and plant hormone signal transduction processes, which are directly related to fruit quality and flavor. These findings illustrate comprehensive evolutionary patterns in Fragaria and the genetic basis of fruit domestication of cultivated strawberry at the genomic/transcriptomic level.

Masculinization of Gene Expression Is Associated with Exaggeration of Male Sexual Dimorphism

PubMed Central

Pointer, Marie A.; Harrison, Peter W.; Wright, Alison E.; Mank, Judith E.

2013-01-01

Gene expression differences between the sexes account for the majority of sexually dimorphic phenotypes, and the study of sex-biased gene expression is important for understanding the genetic basis of complex sexual dimorphisms. However, it has been difficult to test the nature of this relationship due to the fact that sexual dimorphism has traditionally been conceptualized as a dichotomy between males and females, rather than an axis with individuals distributed at intermediate points. The wild turkey (Meleagris gallopavo) exhibits just this sort of continuum, with dominant and subordinate males forming a gradient in male secondary sexual characteristics. This makes it possible for the first time to test the correlation between sex-biased gene expression and sexually dimorphic phenotypes, a relationship crucial to molecular studies of sexual selection and sexual conflict. Here, we show that subordinate male transcriptomes show striking multiple concordances with their relative phenotypic sexual dimorphism. Subordinate males were clearly male rather than intersex, and when compared to dominant males, their transcriptomes were simultaneously demasculinized for male-biased genes and feminized for female-biased genes across the majority of the transcriptome. These results provide the first evidence linking sexually dimorphic transcription and sexually dimorphic phenotypes. More importantly, they indicate that evolutionary changes in sexual dimorphism can be achieved by varying the magnitude of sex-bias in expression across a large proportion of the coding content of a genome. PMID:23966876

Masculinization of gene expression is associated with exaggeration of male sexual dimorphism.

PubMed

Pointer, Marie A; Harrison, Peter W; Wright, Alison E; Mank, Judith E

2013-01-01

Gene expression differences between the sexes account for the majority of sexually dimorphic phenotypes, and the study of sex-biased gene expression is important for understanding the genetic basis of complex sexual dimorphisms. However, it has been difficult to test the nature of this relationship due to the fact that sexual dimorphism has traditionally been conceptualized as a dichotomy between males and females, rather than an axis with individuals distributed at intermediate points. The wild turkey (Meleagris gallopavo) exhibits just this sort of continuum, with dominant and subordinate males forming a gradient in male secondary sexual characteristics. This makes it possible for the first time to test the correlation between sex-biased gene expression and sexually dimorphic phenotypes, a relationship crucial to molecular studies of sexual selection and sexual conflict. Here, we show that subordinate male transcriptomes show striking multiple concordances with their relative phenotypic sexual dimorphism. Subordinate males were clearly male rather than intersex, and when compared to dominant males, their transcriptomes were simultaneously demasculinized for male-biased genes and feminized for female-biased genes across the majority of the transcriptome. These results provide the first evidence linking sexually dimorphic transcription and sexually dimorphic phenotypes. More importantly, they indicate that evolutionary changes in sexual dimorphism can be achieved by varying the magnitude of sex-bias in expression across a large proportion of the coding content of a genome.

High-throughput sequencing of black pepper root transcriptome

PubMed Central

2012-01-01

Background Black pepper (Piper nigrum L.) is one of the most popular spices in the world. It is used in cooking and the preservation of food and even has medicinal properties. Losses in production from disease are a major limitation in the culture of this crop. The major diseases are root rot and foot rot, which are results of root infection by Fusarium solani and Phytophtora capsici, respectively. Understanding the molecular interaction between the pathogens and the host’s root region is important for obtaining resistant cultivars by biotechnological breeding. Genetic and molecular data for this species, though, are limited. In this paper, RNA-Seq technology has been employed, for the first time, to describe the root transcriptome of black pepper. Results The root transcriptome of black pepper was sequenced by the NGS SOLiD platform and assembled using the multiple-k method. Blast2Go and orthoMCL methods were used to annotate 10338 unigenes. The 4472 predicted proteins showed about 52% homology with the Arabidopsis proteome. Two root proteomes identified 615 proteins, which seem to define the plant’s root pattern. Simple-sequence repeats were identified that may be useful in studies of genetic diversity and may have applications in biotechnology and ecology. Conclusions This dataset of 10338 unigenes is crucially important for the biotechnological breeding of black pepper and the ecogenomics of the Magnoliids, a major group of basal angiosperms. PMID:22984782

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

Ruggles, Kelly V.; Tang, Zuojian; Wang, Xuya

Improvements in mass spectrometry (MS)-based peptide sequencing provide a new opportunity to determine whether polymorphisms, mutations and splice variants identified in cancer cells are translated. Herein we therefore describe a proteogenomic data integration tool (QUILTS) and illustrate its application to whole genome, transcriptome and global MS peptide sequence datasets generated from a pair of luminal and basal-like breast cancer patient derived xenografts (PDX). The sensitivity of proteogenomic analysis for singe nucleotide variant (SNV) expression and novel splice junction (NSJ) detection was probed using multiple MS/MS process replicates. Despite over thirty sample replicates, only about 10% of all SNV (somatic andmore » germline) were detected by both DNA and RNA sequencing were observed as peptides. An even smaller proportion of peptides corresponding to NSJ observed by RNA sequencing were detected (<0.1%). Peptides mapping to DNA-detected SNV without a detectable mRNA transcript were also observed demonstrating the transcriptome coverage was also incomplete (~80%). In contrast to germ-line variants, somatic variants were less likely to be detected at the peptide level in the basal-like tumor than the luminal tumor raising the possibility of differential translation or protein degradation effects. In conclusion, the QUILTS program integrates DNA, RNA and peptide sequencing to assess the degree to which somatic mutations are translated and therefore biologically active. By identifying gaps in sequence coverage QUILTS benchmarks current technology and assesses progress towards whole cancer proteome and transcriptome analysis.« less

Algorithms for network-based identification of differential regulators from transcriptome data: a systematic evaluation

PubMed Central

Hui, YU; Ramkrishna, MITRA; Jing, YANG; YuanYuan, LI; ZhongMing, ZHAO

2016-01-01

Identification of differential regulators is critical to understand the dynamics of cellular systems and molecular mechanisms of diseases. Several computational algorithms have recently been developed for this purpose by using transcriptome and network data. However, it remains largely unclear which algorithm performs better under a specific condition. Such knowledge is important for both appropriate application and future enhancement of these algorithms. Here, we systematically evaluated seven main algorithms (TED, TDD, TFactS, RIF1, RIF2, dCSA_t2t, and dCSA_r2t), using both simulated and real datasets. In our simulation evaluation, we artificially inactivated either a single regulator or multiple regulators and examined how well each algorithm detected known gold standard regulators. We found that all these algorithms could effectively discern signals arising from regulatory network differences, indicating the validity of our simulation schema. Among the seven tested algorithms, TED and TFactS were placed first and second when both discrimination accuracy and robustness against data variation were considered. When applied to two independent lung cancer datasets, both TED and TFactS replicated a substantial fraction of their respective differential regulators. Since TED and TFactS rely on two distinct features of transcriptome data, namely differential co-expression and differential expression, both may be applied as mutual references during practical application. PMID:25326829

Phylogenomics, Diversification Dynamics, and Comparative Transcriptomics across the Spider Tree of Life.

PubMed

Fernández, Rosa; Kallal, Robert J; Dimitrov, Dimitar; Ballesteros, Jesús A; Arnedo, Miquel A; Giribet, Gonzalo; Hormiga, Gustavo

2018-05-07

Dating back to almost 400 mya, spiders are among the most diverse terrestrial predators [1]. However, despite considerable effort [1-9], their phylogenetic relationships and diversification dynamics remain poorly understood. Here, we use a synergistic approach to study spider evolution through phylogenomics, comparative transcriptomics, and lineage diversification analyses. Our analyses, based on ca. 2,500 genes from 159 spider species, reject a single origin of the orb web (the "ancient orb-web hypothesis") and suggest that orb webs evolved multiple times since the late Triassic-Jurassic. We find no significant association between the loss of foraging webs and increases in diversification rates, suggesting that other factors (e.g., habitat heterogeneity or biotic interactions) potentially played a key role in spider diversification. Finally, we report notable genomic differences in the main spider lineages: while araneoids (ecribellate orb-weavers and their allies) reveal an enrichment in genes related to behavior and sensory reception, the retrolateral tibial apophysis (RTA) clade-the most diverse araneomorph spider lineage-shows enrichment in genes related to immune responses and polyphenic determination. This study, one of the largest invertebrate phylogenomic analyses to date, highlights the usefulness of transcriptomic data not only to build a robust backbone for the Spider Tree of Life, but also to address the genetic basis of diversification in the spider evolutionary chronicle. Copyright © 2018 Elsevier Ltd. All rights reserved.

Rapid stress-induced transcriptomic changes in the brain depend on beta-adrenergic signaling.

PubMed

Roszkowski, Martin; Manuella, Francesca; von Ziegler, Lukas; Durán-Pacheco, Gonzalo; Moreau, Jean-Luc; Mansuy, Isabelle M; Bohacek, Johannes

2016-08-01

Acute exposure to stressful experiences can rapidly increase anxiety and cause neuropsychiatric disorders. The effects of stress result in part from the release of neurotransmitters and hormones, which regulate gene expression in different brain regions. The fast neuroendocrine response to stress is largely mediated by norepinephrine (NE) and corticotropin releasing hormone (CRH), followed by a slower and more sustained release of corticosterone. While corticosterone is an important regulator of gene expression, it is not clear which stress-signals contribute to the rapid regulation of gene expression observed immediately after stress exposure. Here, we demonstrate in mice that 45 min after an acute swim stress challenge, large changes in gene expression occur across the transcriptome in the hippocampus, a region sensitive to the effects of stress. We identify multiple candidate genes that are rapidly and transiently altered in both males and females. Using a pharmacological approach, we show that most of these rapidly induced genes are regulated by NE through β-adrenergic receptor signaling. We find that CRH and corticosterone can also contribute to rapid changes in gene expression, although these effects appear to be restricted to fewer genes. These results newly reveal a widespread impact of NE on the transcriptome and identify novel genes associated with stress and adrenergic signaling. Copyright © 2016 Elsevier Ltd. All rights reserved.

Elodea nuttallii exposure to mercury exposure under enhanced ultraviolet radiation: Effects on bioaccumulation, transcriptome, pigment content and oxidative stress.

PubMed

Regier, Nicole; Beauvais-Flück, Rebecca; Slaveykova, Vera I; Cosio, Claudia

2016-11-01

The hypothesis that increased UV radiation result in co-tolerance to Hg toxicity in aquatic plants was studied at the physiological and transcriptomic level in Elodea nuttallii. At the transcriptomic level, combined exposure to UV+Hg enhanced the stress response in comparison with single treatments, affecting the expression level of transcripts involved in energy metabolism, lipid metabolism, nutrition, and redox homeostasis. Single and combined UV and Hg treatments dysregulated different genes but with similar functions, suggesting a fine regulation of the plant to stresses triggered by Hg, UV and their combination but lack of co-tolerance. At the physiological level, UV+Hg treatment reduced chlorophyll content and depleted antioxidative compounds such as anthocyanin and GSH/GSSG in E. nuttallii. Nonetheless, combined exposure to UV+Hg resulted in about 30% reduction of Hg accumulation into shoots vs exposure to Hg alone, which was congruent with the level of expression of several transporter genes, as well as the UV effect on Hg bioavailability in water. The findings of the present work underlined the importance of performing experimentation under environmentally realistic conditions and to consider the interplay between contaminants and environmental variables such as light that might have confounding effects to better understand and anticipate the effects of multiple stressors in aquatic environment. Copyright © 2016 Elsevier B.V. All rights reserved.

Breast Cancer Methylomes Establish an Epigenomic Foundation for Metastasis

PubMed Central

Fang, Fang; Turcan, Sevin; Rimner, Andreas; Kaufman, Andrew; Giri, Dilip; Morris, Luc G. T.; Shen, Ronglai; Seshan, Venkatraman; Mo, Qianxing; Heguy, Adriana; Baylin, Stephen B.; Ahuja, Nita; Viale, Agnes; Massague, Joan; Norton, Larry; Vahdat, Linda T.; Moynahan, Mary Ellen; Chan, Timothy A.

2011-01-01

Cancer-specific alterations in DNA methylation are hallmarks of human malignancies; however, the nature of the breast cancer epigenome and its effects on metastatic behavior remain obscure. To address this issue, we used genome-wide analysis to characterize the methylomes of breast cancers with diverse metastatic behavior. Groups of breast tumors were characterized by the presence or absence of coordinate hypermethylation at a large number of genes, demonstrating a breast CpG island methylator phenotype (B-CIMP). The B-CIMP provided a distinct epigenomic profile and was a strong determinant of metastatic potential. Specifically, the presence of the B-CIMP in tumors was associated with low metastatic risk and survival, and the absence of the B-CIMP was associated with high metastatic risk and death. B-CIMP loci were highly enriched for genes that make up the metastasis transcriptome. Methylation at B-CIMP genes accounted for much of the transcriptomal diversity between breast cancers of varying prognosis, indicating a fundamental epigenomic contribution to metastasis. Comparison of the loci affected by the B-CIMP with those affected by the hypermethylator phenotype in glioma and colon cancer revealed that the CIMP signature was shared by multiple human malignancies. Our data provide a unifying epigenomic framework linking breast cancers with varying outcome and transcriptomic changes underlying metastasis. These findings significantly enhance our understanding of breast cancer oncogenesis and aid the development of new prognostic biomarkers for this common malignancy. PMID:21430268

Signatures of inflammation and impending multiple organ dysfunction in the hyperacute phase of trauma: A prospective cohort study

PubMed Central

Longhi, M. Paula; Hoti, Mimoza; Patel, Minal B.; O’Dwyer, Michael; Nourshargh, Sussan; Barnes, Michael R.; Brohi, Karim

2017-01-01

Background Severe trauma induces a widespread response of the immune system. This “genomic storm” can lead to poor outcomes, including Multiple Organ Dysfunction Syndrome (MODS). MODS carries a high mortality and morbidity rate and adversely affects long-term health outcomes. Contemporary management of MODS is entirely supportive, and no specific therapeutics have been shown to be effective in reducing incidence or severity. The pathogenesis of MODS remains unclear, and several models are proposed, such as excessive inflammation, a second-hit insult, or an imbalance between pro- and anti-inflammatory pathways. We postulated that the hyperacute window after trauma may hold the key to understanding how the genomic storm is initiated and may lead to a new understanding of the pathogenesis of MODS. Methods and findings We performed whole blood transcriptome and flow cytometry analyses on a total of 70 critically injured patients (Injury Severity Score [ISS] ≥ 25) at The Royal London Hospital in the hyperacute time period within 2 hours of injury. We compared transcriptome findings in 36 critically injured patients with those of 6 patients with minor injuries (ISS ≤ 4). We then performed flow cytometry analyses in 34 critically injured patients and compared findings with those of 9 healthy volunteers. Immediately after injury, only 1,239 gene transcripts (4%) were differentially expressed in critically injured patients. By 24 hours after injury, 6,294 transcripts (21%) were differentially expressed compared to the hyperacute window. Only 202 (16%) genes differentially expressed in the hyperacute window were still expressed in the same direction at 24 hours postinjury. Pathway analysis showed principally up-regulation of pattern recognition and innate inflammatory pathways, with down-regulation of adaptive responses. Immune deconvolution, flow cytometry, and modular analysis suggested a central role for neutrophils and Natural Killer (NK) cells, with underexpression of T- and B cell responses. In the transcriptome cohort, 20 critically injured patients later developed MODS. Compared with the 16 patients who did not develop MODS (NoMODS), maximal differential expression was seen within the hyperacute window. In MODS versus NoMODS, 363 genes were differentially expressed on admission, compared to only 33 at 24 hours postinjury. MODS transcripts differentially expressed in the hyperacute window showed enrichment among diseases and biological functions associated with cell survival and organismal death rather than inflammatory pathways. There was differential up-regulation of NK cell signalling pathways and markers in patients who would later develop MODS, with down-regulation of neutrophil deconvolution markers. This study is limited by its sample size, precluding more detailed analyses of drivers of the hyperacute response and different MODS phenotypes, and requires validation in other critically injured cohorts. Conclusions In this study, we showed how the hyperacute postinjury time window contained a focused, specific signature of the response to critical injury that led to widespread genomic activation. A transcriptomic signature for later development of MODS was present in this hyperacute window; it showed a strong signal for cell death and survival pathways and implicated NK cells and neutrophil populations in this differential response. PMID:28715416

Transcriptomic resources for the medicinal legume Mucuna pruriens: de novo transcriptome assembly, annotation, identification and validation of EST-SSR markers.

PubMed

Sathyanarayana, N; Pittala, Ranjith Kumar; Tripathi, Pankaj Kumar; Chopra, Ratan; Singh, Heikham Russiachand; Belamkar, Vikas; Bhardwaj, Pardeep Kumar; Doyle, Jeff J; Egan, Ashley N

2017-05-25

The medicinal legume Mucuna pruriens (L.) DC. has attracted attention worldwide as a source of the anti-Parkinson's drug L-Dopa. It is also a popular green manure cover crop that offers many agronomic benefits including high protein content, nitrogen fixation and soil nutrients. The plant currently lacks genomic resources and there is limited knowledge on gene expression, metabolic pathways, and genetics of secondary metabolite production. Here, we present transcriptomic resources for M. pruriens, including a de novo transcriptome assembly and annotation, as well as differential transcript expression analyses between root, leaf, and pod tissues. We also develop microsatellite markers and analyze genetic diversity and population structure within a set of Indian germplasm accessions. One-hundred ninety-one million two hundred thirty-three thousand two hundred forty-two bp cleaned reads were assembled into 67,561 transcripts with mean length of 626 bp and N50 of 987 bp. Assembled sequences were annotated using BLASTX against public databases with over 80% of transcripts annotated. We identified 7,493 simple sequence repeat (SSR) motifs, including 787 polymorphic repeats between the parents of a mapping population. 134 SSRs from expressed sequenced tags (ESTs) were screened against 23 M. pruriens accessions from India, with 52 EST-SSRs retained after quality control. Population structure analysis using a Bayesian framework implemented in fastSTRUCTURE showed nearly similar groupings as with distance-based (neighbor-joining) and principal component analyses, with most of the accessions clustering per geographical origins. Pair-wise comparison of transcript expression in leaves, roots and pods identified 4,387 differentially expressed transcripts with the highest number occurring between roots and leaves. Differentially expressed transcripts were enriched with transcription factors and transcripts annotated as belonging to secondary metabolite pathways. The M. pruriens transcriptomic resources generated in this study provide foundational resources for gene discovery and development of molecular markers. Polymorphic SSRs identified can be used for genetic diversity, marker-trait analyses, and development of functional markers for crop improvement. The results of differential expression studies can be used to investigate genes involved in L-Dopa synthesis and other key metabolic pathways in M. pruriens.

The exosome component Rrp6 is required for RNA polymerase II termination at specific targets of the Nrd1-Nab3 pathway.

PubMed

Fox, Melanie J; Gao, Hongyu; Smith-Kinnaman, Whitney R; Liu, Yunlong; Mosley, Amber L

2015-01-01

The exosome and its nuclear specific subunit Rrp6 form a 3'-5' exonuclease complex that regulates diverse aspects of RNA biology including 3' end processing and degradation of a variety of noncoding RNAs (ncRNAs) and unstable transcripts. Known targets of the nuclear exosome include short (<1000 bp) RNAPII transcripts such as small noncoding RNAs (snRNAs), cryptic unstable transcripts (CUTs), and some stable unannotated transcripts (SUTs) that are terminated by an Nrd1, Nab3, and Sen1 (NNS) dependent mechanism. NNS-dependent termination is coupled to RNA 3' end processing and/or degradation by the Rrp6/exosome in yeast. Recent work suggests Nrd1 is necessary for transcriptome surveillance, regulating promoter directionality and suppressing antisense transcription independently of, or prior to, Rrp6 activity. It remains unclear whether Rrp6 is directly involved in termination; however, Rrp6 has been implicated in the 3' end processing and degradation of ncRNA transcripts including CUTs. To determine the role of Rrp6 in NNS termination globally, we performed RNA sequencing (RNA-Seq) on total RNA and perform ChIP-exo analysis of RNA Polymerase II (RNAPII) localization. Deletion of RRP6 promotes hyper-elongation of multiple NNS-dependent transcripts resulting from both improperly processed 3' RNA ends and faulty transcript termination at specific target genes. The defects in RNAPII termination cause transcriptome-wide changes in mRNA expression through transcription interference and/or antisense repression, similar to previously reported effects of depleting Nrd1 from the nucleus. Elongated transcripts were identified within all classes of known NNS targets with the largest changes in transcription termination occurring at CUTs. Interestingly, the extended transcripts that we have detected in our studies show remarkable similarity to Nrd1-unterminated transcripts at many locations, suggesting that Rrp6 acts with the NNS complex globally to promote transcription termination in addition to 3' end RNA processing and/or degradation at specific targets.

The Physalis peruviana leaf transcriptome: assembly, annotation and gene model prediction

PubMed Central

2012-01-01

Background Physalis peruviana commonly known as Cape gooseberry is a member of the Solanaceae family that has an increasing popularity due to its nutritional and medicinal values. A broad range of genomic tools is available for other Solanaceae, including tomato and potato. However, limited genomic resources are currently available for Cape gooseberry. Results We report the generation of a total of 652,614 P. peruviana Expressed Sequence Tags (ESTs), using 454 GS FLX Titanium technology. ESTs, with an average length of 371 bp, were obtained from a normalized leaf cDNA library prepared using a Colombian commercial variety. De novo assembling was performed to generate a collection of 24,014 isotigs and 110,921 singletons, with an average length of 1,638 bp and 354 bp, respectively. Functional annotation was performed using NCBI’s BLAST tools and Blast2GO, which identified putative functions for 21,191 assembled sequences, including gene families involved in all the major biological processes and molecular functions as well as defense response and amino acid metabolism pathways. Gene model predictions in P. peruviana were obtained by using the genomes of Solanum lycopersicum (tomato) and Solanum tuberosum (potato). We predict 9,436 P. peruviana sequences with multiple-exon models and conserved intron positions with respect to the potato and tomato genomes. Additionally, to study species diversity we developed 5,971 SSR markers from assembled ESTs. Conclusions We present the first comprehensive analysis of the Physalis peruviana leaf transcriptome, which will provide valuable resources for development of genetic tools in the species. Assembled transcripts with gene models could serve as potential candidates for marker discovery with a variety of applications including: functional diversity, conservation and improvement to increase productivity and fruit quality. P. peruviana was estimated to be phylogenetically branched out before the divergence of five other Solanaceae family members, S. lycopersicum, S. tuberosum, Capsicum spp, S. melongena and Petunia spp. PMID:22533342

The Physalis peruviana leaf transcriptome: assembly, annotation and gene model prediction.

PubMed

Garzón-Martínez, Gina A; Zhu, Z Iris; Landsman, David; Barrero, Luz S; Mariño-Ramírez, Leonardo

2012-04-25

Physalis peruviana commonly known as Cape gooseberry is a member of the Solanaceae family that has an increasing popularity due to its nutritional and medicinal values. A broad range of genomic tools is available for other Solanaceae, including tomato and potato. However, limited genomic resources are currently available for Cape gooseberry. We report the generation of a total of 652,614 P. peruviana Expressed Sequence Tags (ESTs), using 454 GS FLX Titanium technology. ESTs, with an average length of 371 bp, were obtained from a normalized leaf cDNA library prepared using a Colombian commercial variety. De novo assembling was performed to generate a collection of 24,014 isotigs and 110,921 singletons, with an average length of 1,638 bp and 354 bp, respectively. Functional annotation was performed using NCBI's BLAST tools and Blast2GO, which identified putative functions for 21,191 assembled sequences, including gene families involved in all the major biological processes and molecular functions as well as defense response and amino acid metabolism pathways. Gene model predictions in P. peruviana were obtained by using the genomes of Solanum lycopersicum (tomato) and Solanum tuberosum (potato). We predict 9,436 P. peruviana sequences with multiple-exon models and conserved intron positions with respect to the potato and tomato genomes. Additionally, to study species diversity we developed 5,971 SSR markers from assembled ESTs. We present the first comprehensive analysis of the Physalis peruviana leaf transcriptome, which will provide valuable resources for development of genetic tools in the species. Assembled transcripts with gene models could serve as potential candidates for marker discovery with a variety of applications including: functional diversity, conservation and improvement to increase productivity and fruit quality. P. peruviana was estimated to be phylogenetically branched out before the divergence of five other Solanaceae family members, S. lycopersicum, S. tuberosum, Capsicum spp, S. melongena and Petunia spp.

Ovary transcriptome profiling via artificial intelligence reveals a transcriptomic fingerprint predicting egg quality in striped bass, Morone saxatilis.

PubMed

Chapman, Robert W; Reading, Benjamin J; Sullivan, Craig V

2014-01-01

Inherited gene transcripts deposited in oocytes direct early embryonic development in all vertebrates, but transcript profiles indicative of embryo developmental competence have not previously been identified. We employed artificial intelligence to model profiles of maternal ovary gene expression and their relationship to egg quality, evaluated as production of viable mid-blastula stage embryos, in the striped bass (Morone saxatilis), a farmed species with serious egg quality problems. In models developed using artificial neural networks (ANNs) and supervised machine learning, collective changes in the expression of a limited suite of genes (233) representing <2% of the queried ovary transcriptome explained >90% of the eventual variance in embryo survival. Egg quality related to minor changes in gene expression (<0.2-fold), with most individual transcripts making a small contribution (<1%) to the overall prediction of egg quality. These findings indicate that the predictive power of the transcriptome as regards egg quality resides not in levels of individual genes, but rather in the collective, coordinated expression of a suite of transcripts constituting a transcriptomic "fingerprint". Correlation analyses of the corresponding candidate genes indicated that dysfunction of the ubiquitin-26S proteasome, COP9 signalosome, and subsequent control of the cell cycle engenders embryonic developmental incompetence. The affected gene networks are centrally involved in regulation of early development in all vertebrates, including humans. By assessing collective levels of the relevant ovarian transcripts via ANNs we were able, for the first time in any vertebrate, to accurately predict the subsequent embryo developmental potential of eggs from individual females. Our results show that the transcriptomic fingerprint evidencing developmental dysfunction is highly predictive of, and therefore likely to regulate, egg quality, a biologically complex trait crucial to reproductive fitness.

RNA-seq analysis of Rubus idaeus cv. Nova: transcriptome sequencing and de novo assembly for subsequent functional genomics approaches.

PubMed

Hyun, Tae Kyung; Lee, Sarah; Kumar, Dhinesh; Rim, Yeonggil; Kumar, Ritesh; Lee, Sang Yeol; Lee, Choong Hwan; Kim, Jae-Yean

2014-10-01

Using Illumina sequencing technology, we have generated the large-scale transcriptome sequencing data containing abundant information on genes involved in the metabolic pathways in R. idaeus cv. Nova fruits. Rubus idaeus (Red raspberry) is one of the important economical crops that possess numerous nutrients, micronutrients and phytochemicals with essential health benefits to human. The molecular mechanism underlying the ripening process and phytochemical biosynthesis in red raspberry is attributed to the changes in gene expression, but very limited transcriptomic and genomic information in public databases is available. To address this issue, we generated more than 51 million sequencing reads from R. idaeus cv. Nova fruit using Illumina RNA-Seq technology. After de novo assembly, we obtained 42,604 unigenes with an average length of 812 bp. At the protein level, Nova fruit transcriptome showed 77 and 68 % sequence similarities with Rubus coreanus and Fragaria versa, respectively, indicating the evolutionary relationship between them. In addition, 69 % of assembled unigenes were annotated using public databases including NCBI non-redundant, Cluster of Orthologous Groups and Gene ontology database, suggesting that our transcriptome dataset provides a valuable resource for investigating metabolic processes in red raspberry. To analyze the relationship between several novel transcripts and the amounts of metabolites such as γ-aminobutyric acid and anthocyanins, real-time PCR and target metabolite analysis were performed on two different ripening stages of Nova. This is the first attempt using Illumina sequencing platform for RNA sequencing and de novo assembly of Nova fruit without reference genome. Our data provide the most comprehensive transcriptome resource available for Rubus fruits, and will be useful for understanding the ripening process and for breeding R. idaeus cultivars with improved fruit quality.

Transcriptome analysis and gene expression profiling of abortive and developing ovules during fruit development in hazelnut.

PubMed

Cheng, Yunqing; Liu, Jianfeng; Zhang, Huidi; Wang, Ju; Zhao, Yixin; Geng, Wanting

2015-01-01

A high ratio of blank fruit in hazelnut (Corylus heterophylla Fisch) is a very common phenomenon that causes serious yield losses in northeast China. The development of blank fruit in the Corylus genus is known to be associated with embryo abortion. However, little is known about the molecular mechanisms responsible for embryo abortion during the nut development stage. Genomic information for C. heterophylla Fisch is not available; therefore, data related to transcriptome and gene expression profiling of developing and abortive ovules are needed. In this study, de novo transcriptome sequencing and RNA-seq analysis were conducted using short-read sequencing technology (Illumina HiSeq 2000). The results of the transcriptome assembly analysis revealed genetic information that was associated with the fruit development stage. Two digital gene expression libraries were constructed, one for a full (normally developing) ovule and one for an empty (abortive) ovule. Transcriptome sequencing and assembly results revealed 55,353 unigenes, including 18,751 clusters and 36,602 singletons. These results were annotated using the public databases NR, NT, Swiss-Prot, KEGG, COG, and GO. Using digital gene expression profiling, gene expression differences in developing and abortive ovules were identified. A total of 1,637 and 715 unigenes were significantly upregulated and downregulated, respectively, in abortive ovules, compared with developing ovules. Quantitative real-time polymerase chain reaction analysis was used in order to verify the differential expression of some genes. The transcriptome and digital gene expression profiling data of normally developing and abortive ovules in hazelnut provide exhaustive information that will improve our understanding of the molecular mechanisms of abortive ovule formation in hazelnut.

Prior to extension, Transcriptomes of fibroblast-like Synoviocytes from extended and Polyarticular juvenile idiopathic arthritis are indistinguishable.

PubMed

Brescia, AnneMarie C; Simonds, Megan M; McCahan, Suzanne M; Sullivan, Kathleen E; Rose, Carlos D

2018-01-08

Our intent was to identify differences between the transcriptome of fibroblast-like synoviocytes (FLS) in oligoarticular juvenile idiopathic arthritis (JIA) before extension when compared to persistent subtype of JIA, when the two are clinically indistinguishable. Additionally, we sought to determine if differences between the transcriptomes of FLS from extended-to-be and polyarticular course JIA could be detected. Our hypothesis was that intrinsic differences in the transcriptome of the FLS from extended-to-be JIA would distinguish them from persistent oligoarticular JIA, before the course is clinically apparent. Global gene expression was defined in cultured FLS from 6 controls, 12 JIA with persistent course, 7 JIA prior to extension (extended-to-be), 4 JIA with extended course and 6 polyarticular onset, using Affymetrix Human GeneChips 133plus2.0. Bioconductor Linear Models for Microarray Analysis revealed 22 probesets with differential expression between persistent and extended-to-be FLS at 15% FDR, however only 2 probesets distinguished extended-to-be from extended and none distinguished extended-to-be and polyarticular at 15% FDR. Differences in extended and polyarticular gene expression profiles were not detected. Confirmation of select genes was done on the RNA level by RT-qPCR and on the protein level in synovial fluid by ELISA. The transcriptome of FLS from extended-to-be juvenile idiopathic arthritis is distinct from persistent course before a clinical distinction can be made. Additionally, the transcriptome of extended-to-be and polyarticular course, including those who have already extended, are indistinguishable. These gene expression data suggest that FLS already reflect a polyarticular behavior early in disease course, suggesting that extended-to-be may be "latent polyarticular" at onset. These differences can be used to develop early biomarkers of disease course, allowing for better-informed treatment decisions.

Next-Generation Transcriptome Profiling of the Salmon Louse Caligus rogercresseyi Exposed to Deltamethrin (AlphaMax™): Discovery of Relevant Genes and Sex-Related Differences.

PubMed

Chávez-Mardones, Jacqueline; Gallardo-Escárate, Cristian

2015-12-01

Sea lice are one of the main parasites affecting the salmon aquaculture industry, causing significant economic losses worldwide. Increased resistance to traditional chemical treatments has created the need to find alternative control methods. Therefore, the objective of this study was to identify the transcriptome response of the salmon louse Caligus rogercresseyi to the delousing drug deltamethrin (AlphaMax™). Through bioassays with different concentrations of deltamethrin, adult salmon lice transcriptomes were sequenced from cDNA libraries in the MiSeq Illumina platform. A total of 78 million reads for females and males were assembled in 30,212 and 38,536 contigs, respectively. De novo assembly yielded 86,878 high-quality contigs and, based on published data, it was possible to annotate and identify relevant genes involved in several biological processes. RNA-seq analysis in conjunction with heatmap hierarchical clustering evidenced that pyrethroids modify the ectoparasitic transcriptome in adults, affecting molecular processes associated with the nervous system, cuticle formation, oxidative stress, reproduction, and metabolism, among others. Furthermore, sex-related transcriptome differences were evidenced. Specifically, 534 and 1033 exclusive transcripts were identified for males and females, respectively, and 154 were shared between sexes. For males, estradiol 17-beta-dehydrogenase, sphingolipid delta4-desaturase DES1, ketosamine-3-kinase, and arylsulfatase A, among others, were discovered, while for females, vitellogenin 1, glycoprotein G, transaldolase, and nitric oxide synthase were among those identified. The shared transcripts included annotations for tropomyosin, γ-crystallin A, glutamate receptor-metabotropic, glutathione S-transferase, and carboxipeptidase B. The present study reveals that deltamethrin generates a complex transcriptome response in C. rogercresseyi, thus providing valuable genomic information for developing new delousing drugs.

Mining genes involved in insecticide resistance of Liposcelis bostrychophila Badonnel by transcriptome and expression profile analysis.

PubMed

Dou, Wei; Shen, Guang-Mao; Niu, Jin-Zhi; Ding, Tian-Bo; Wei, Dan-Dan; Wang, Jin-Jun

2013-01-01

Recent studies indicate that infestations of psocids pose a new risk for global food security. Among the psocids species, Liposcelis bostrychophila Badonnel has gained recognition in importance because of its parthenogenic reproduction, rapid adaptation, and increased worldwide distribution. To date, the molecular data available for L. bostrychophila is largely limited to genes identified through homology. Also, no transcriptome data relevant to psocids infection is available. In this study, we generated de novo assembly of L. bostrychophila transcriptome performed through the short read sequencing technology (Illumina). In a single run, we obtained more than 51 million sequencing reads that were assembled into 60,012 unigenes (mean size = 711 bp) by Trinity. The transcriptome sequences from different developmental stages of L. bostrychophila including egg, nymph and adult were annotated with non-redundant (Nr) protein database, gene ontology (GO), cluster of orthologous groups of proteins (COG), and KEGG orthology (KO). The analysis revealed three major enzyme families involved in insecticide metabolism as differentially expressed in the L. bostrychophila transcriptome. A total of 49 P450-, 31 GST- and 21 CES-specific genes representing the three enzyme families were identified. Besides, 16 transcripts were identified to contain target site sequences of resistance genes. Furthermore, we profiled gene expression patterns upon insecticide (malathion and deltamethrin) exposure using the tag-based digital gene expression (DGE) method. The L. bostrychophila transcriptome and DGE data provide gene expression data that would further our understanding of molecular mechanisms in psocids. In particular, the findings of this investigation will facilitate identification of genes involved in insecticide resistance and designing of new compounds for control of psocids.

Mining Genes Involved in Insecticide Resistance of Liposcelis bostrychophila Badonnel by Transcriptome and Expression Profile Analysis

PubMed Central

Dou, Wei; Shen, Guang-Mao; Niu, Jin-Zhi; Ding, Tian-Bo; Wei, Dan-Dan; Wang, Jin-Jun

2013-01-01

Background Recent studies indicate that infestations of psocids pose a new risk for global food security. Among the psocids species, Liposcelis bostrychophila Badonnel has gained recognition in importance because of its parthenogenic reproduction, rapid adaptation, and increased worldwide distribution. To date, the molecular data available for L. bostrychophila is largely limited to genes identified through homology. Also, no transcriptome data relevant to psocids infection is available. Methodology and Principal Findings In this study, we generated de novo assembly of L. bostrychophila transcriptome performed through the short read sequencing technology (Illumina). In a single run, we obtained more than 51 million sequencing reads that were assembled into 60,012 unigenes (mean size = 711 bp) by Trinity. The transcriptome sequences from different developmental stages of L. bostrychophila including egg, nymph and adult were annotated with non-redundant (Nr) protein database, gene ontology (GO), cluster of orthologous groups of proteins (COG), and KEGG orthology (KO). The analysis revealed three major enzyme families involved in insecticide metabolism as differentially expressed in the L. bostrychophila transcriptome. A total of 49 P450-, 31 GST- and 21 CES-specific genes representing the three enzyme families were identified. Besides, 16 transcripts were identified to contain target site sequences of resistance genes. Furthermore, we profiled gene expression patterns upon insecticide (malathion and deltamethrin) exposure using the tag-based digital gene expression (DGE) method. Conclusion The L. bostrychophila transcriptome and DGE data provide gene expression data that would further our understanding of molecular mechanisms in psocids. In particular, the findings of this investigation will facilitate identification of genes involved in insecticide resistance and designing of new compounds for control of psocids. PMID:24278202

The Long Noncoding RNA Transcriptome of Dictyostelium discoideum Development.

PubMed

Rosengarten, Rafael D; Santhanam, Balaji; Kokosar, Janez; Shaulsky, Gad

2017-02-09

Dictyostelium discoideum live in the soil as single cells, engulfing bacteria and growing vegetatively. Upon starvation, tens of thousands of amoebae enter a developmental program that includes aggregation, multicellular differentiation, and sporulation. Major shifts across the protein-coding transcriptome accompany these developmental changes. However, no study has presented a global survey of long noncoding RNAs (ncRNAs) in D. discoideum To characterize the antisense and long intergenic noncoding RNA (lncRNA) transcriptome, we analyzed previously published developmental time course samples using an RNA-sequencing (RNA-seq) library preparation method that selectively depletes ribosomal RNAs (rRNAs). We detected the accumulation of transcripts for 9833 protein-coding messenger RNAs (mRNAs), 621 lncRNAs, and 162 putative antisense RNAs (asRNAs). The noncoding RNAs were interspersed throughout the genome, and were distinct in expression level, length, and nucleotide composition. The noncoding transcriptome displayed a temporal profile similar to the coding transcriptome, with stages of gradual change interspersed with larger leaps. The transcription profiles of some noncoding RNAs were strongly correlated with known differentially expressed coding RNAs, hinting at a functional role for these molecules during development. Examining the mitochondrial transcriptome, we modeled two novel antisense transcripts. We applied yet another ribosomal depletion method to a subset of the samples to better retain transfer RNA (tRNA) transcripts. We observed polymorphisms in tRNA anticodons that suggested a post-transcriptional means by which D. discoideum compensates for codons missing in the genomic complement of tRNAs. We concluded that the prevalence and characteristics of long ncRNAs indicate that these molecules are relevant to the progression of molecular and cellular phenotypes during development. Copyright © 2017 Rosengarten et al.

Selenium supplementation prevents metabolic and transcriptomic responses to cadmium in mouse lung.

PubMed

Hu, Xin; Chandler, Joshua D; Fernandes, Jolyn; Orr, Michael L; Hao, Li; Uppal, Karan; Neujahr, David C; Jones, Dean P; Go, Young-Mi

2018-04-12

The protective effect of selenium (Se) on cadmium (Cd) toxicity is well documented, but underlying mechanisms are unclear. Male mice fed standard diet were given Cd (CdCl 2 , 18 μmol/L) in drinking water with or without Se (Na 2 SeO 4, 20 μmol/L) for 16 weeks. Lungs were analyzed for Cd concentration, transcriptomics and metabolomics. Data were analyzed with biostatistics, bioinformatics, pathway enrichment analysis, and combined transcriptome-metabolome-wide association study. Mice treated with Cd had higher lung Cd content (1.7 ± 0.4 pmol/mg protein) than control mice (0.8 ± 0.3 pmol/mg protein) or mice treated with Cd and Se (0.4 ± 0.1 pmol/mg protein). Gene set enrichment analysis of transcriptomics data showed that Se prevented Cd effects on inflammatory and myogenesis genes and diminished Cd effects on several other pathways. Similarly, Se prevented Cd-disrupted metabolic pathways in amino acid metabolism and urea cycle. Integrated transcriptome and metabolome network analysis showed that Cd treatment had a network structure with fewer gene-metabolite clusters compared to control. Centrality measurements showed that Se counteracted changes in a group of Cd-responsive genes including Zdhhc11, (protein-cysteine S-palmitoyltransferase), Ighg1 (immunoglobulin heavy constant gamma-1) and associated changes in metabolite concentrations. Co-administration of Se with Cd prevented Cd increase in lung and prevented Cd-associated pathway and network responses of the transcriptome and metabolome. Se protection against Cd toxicity in lung involves complex systems responses. Environmental Cd stimulates proinflammatory and profibrotic signaling. The present results indicate that dietary or supplemental Se could be useful to mitigate Cd toxicity. Published by Elsevier B.V.

Ovary Transcriptome Profiling via Artificial Intelligence Reveals a Transcriptomic Fingerprint Predicting Egg Quality in Striped Bass, Morone saxatilis

PubMed Central

2014-01-01

Inherited gene transcripts deposited in oocytes direct early embryonic development in all vertebrates, but transcript profiles indicative of embryo developmental competence have not previously been identified. We employed artificial intelligence to model profiles of maternal ovary gene expression and their relationship to egg quality, evaluated as production of viable mid-blastula stage embryos, in the striped bass (Morone saxatilis), a farmed species with serious egg quality problems. In models developed using artificial neural networks (ANNs) and supervised machine learning, collective changes in the expression of a limited suite of genes (233) representing <2% of the queried ovary transcriptome explained >90% of the eventual variance in embryo survival. Egg quality related to minor changes in gene expression (<0.2-fold), with most individual transcripts making a small contribution (<1%) to the overall prediction of egg quality. These findings indicate that the predictive power of the transcriptome as regards egg quality resides not in levels of individual genes, but rather in the collective, coordinated expression of a suite of transcripts constituting a transcriptomic “fingerprint”. Correlation analyses of the corresponding candidate genes indicated that dysfunction of the ubiquitin-26S proteasome, COP9 signalosome, and subsequent control of the cell cycle engenders embryonic developmental incompetence. The affected gene networks are centrally involved in regulation of early development in all vertebrates, including humans. By assessing collective levels of the relevant ovarian transcripts via ANNs we were able, for the first time in any vertebrate, to accurately predict the subsequent embryo developmental potential of eggs from individual females. Our results show that the transcriptomic fingerprint evidencing developmental dysfunction is highly predictive of, and therefore likely to regulate, egg quality, a biologically complex trait crucial to reproductive fitness. PMID:24820964

Origin and functional diversification of an amphibian defense peptide arsenal.

PubMed

Roelants, Kim; Fry, Bryan G; Ye, Lumeng; Stijlemans, Benoit; Brys, Lea; Kok, Philippe; Clynen, Elke; Schoofs, Liliane; Cornelis, Pierre; Bossuyt, Franky

2013-01-01

The skin secretion of many amphibians contains an arsenal of bioactive molecules, including hormone-like peptides (HLPs) acting as defense toxins against predators, and antimicrobial peptides (AMPs) providing protection against infectious microorganisms. Several amphibian taxa seem to have independently acquired the genes to produce skin-secreted peptide arsenals, but it remains unknown how these originated from a non-defensive ancestral gene and evolved diverse defense functions against predators and pathogens. We conducted transcriptome, genome, peptidome and phylogenetic analyses to chart the full gene repertoire underlying the defense peptide arsenal of the frog Silurana tropicalis and reconstruct its evolutionary history. Our study uncovers a cluster of 13 transcriptionally active genes, together encoding up to 19 peptides, including diverse HLP homologues and AMPs. This gene cluster arose from a duplicated gastrointestinal hormone gene that attained a HLP-like defense function after major remodeling of its promoter region. Instead, new defense functions, including antimicrobial activity, arose by mutation of the precursor proteins, resulting in the proteolytic processing of secondary peptides alongside the original ones. Although gene duplication did not trigger functional innovation, it may have subsequently facilitated the convergent loss of the original function in multiple gene lineages (subfunctionalization), completing their transformation from HLP gene to AMP gene. The processing of multiple peptides from a single precursor entails a mechanism through which peptide-encoding genes may establish new functions without the need for gene duplication to avoid adaptive conflicts with older ones.

Adaptation Genomics of a Small-Colony Variant in a Pseudomonas chlororaphis 30-84 Biofilm

PubMed Central

Dorosky, Robert J.; Han, Cliff S.; Lo, Chien-chi; Dichosa, Armand E. K.; Chain, Patrick S.; Yu, Jun Myoung; Pierson, Leland S.

2014-01-01

The rhizosphere-colonizing bacterium Pseudomonas chlororaphis 30-84 is an effective biological control agent against take-all disease of wheat. In this study, we characterize a small-colony variant (SCV) isolated from a P. chlororaphis 30-84 biofilm. The SCV exhibited pleiotropic phenotypes, including small cell size, slow growth and motility, low levels of phenazine production, and increased biofilm formation and resistance to antimicrobials. To better understand the genetic alterations underlying these phenotypes, RNA and whole-genome sequencing analyses were conducted comparing an SCV to the wild-type strain. Of the genome's 5,971 genes, transcriptomic profiling indicated that 1,098 (18.4%) have undergone substantial reprograming of gene expression in the SCV. Whole-genome sequence analysis revealed multiple alterations in the SCV, including mutations in yfiR (cyclic-di-GMP production), fusA (elongation factor), and cyoE (heme synthesis) and a 70-kb deletion. Genetic analysis revealed that the yfiR locus plays a major role in controlling SCV phenotypes, including colony size, growth, motility, and biofilm formation. Moreover, a point mutation in the fusA gene contributed to kanamycin resistance. Interestingly, the SCV can partially switch back to wild-type morphologies under specific conditions. Our data also support the idea that phenotypic switching in P. chlororaphis is not due to simple genetic reversions but may involve multiple secondary mutations. The emergence of these highly adherent and antibiotic-resistant SCVs within the biofilm might play key roles in P. chlororaphis natural persistence. PMID:25416762

Origin and Functional Diversification of an Amphibian Defense Peptide Arsenal

PubMed Central

Roelants, Kim; Fry, Bryan G.; Ye, Lumeng; Stijlemans, Benoit; Brys, Lea; Kok, Philippe; Clynen, Elke; Schoofs, Liliane; Cornelis, Pierre; Bossuyt, Franky

2013-01-01

The skin secretion of many amphibians contains an arsenal of bioactive molecules, including hormone-like peptides (HLPs) acting as defense toxins against predators, and antimicrobial peptides (AMPs) providing protection against infectious microorganisms. Several amphibian taxa seem to have independently acquired the genes to produce skin-secreted peptide arsenals, but it remains unknown how these originated from a non-defensive ancestral gene and evolved diverse defense functions against predators and pathogens. We conducted transcriptome, genome, peptidome and phylogenetic analyses to chart the full gene repertoire underlying the defense peptide arsenal of the frog Silurana tropicalis and reconstruct its evolutionary history. Our study uncovers a cluster of 13 transcriptionally active genes, together encoding up to 19 peptides, including diverse HLP homologues and AMPs. This gene cluster arose from a duplicated gastrointestinal hormone gene that attained a HLP-like defense function after major remodeling of its promoter region. Instead, new defense functions, including antimicrobial activity, arose by mutation of the precursor proteins, resulting in the proteolytic processing of secondary peptides alongside the original ones. Although gene duplication did not trigger functional innovation, it may have subsequently facilitated the convergent loss of the original function in multiple gene lineages (subfunctionalization), completing their transformation from HLP gene to AMP gene. The processing of multiple peptides from a single precursor entails a mechanism through which peptide-encoding genes may establish new functions without the need for gene duplication to avoid adaptive conflicts with older ones. PMID:23935531

De novo analysis of the Nilaparvata lugens (Stål) antenna transcriptome and expression patterns of olfactory genes.

PubMed

Zhou, Shuang-Shuang; Sun, Ze; Ma, Weihua; Chen, Wei; Wang, Man-Qun

2014-03-01

We sequenced the antenna transcriptome of the brown planthopper (BPH), Nilaparvata lugens (Stål), a global rice pest, and performed transcriptome analysis on BPH antenna. We obtained about 40million 90bp reads that were assembled into 75,874 unigenes with a mean size of 456bp. Among the antenna transcripts, 32,856 (43%) showed significant similarity (E-value <1e(-5)) to known proteins in the NCBI database. Gene ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were used to classify functions of BPH antenna genes. We identified 10 odorant-binding proteins (OBPs), including 7 previously unidentified, and 11 chemosensory proteins (CSPs), including two new members. The expression profiles of 4 OBPs and 2 CSPs were determined by q-PCR for antenna, abdomen, leg and wing of insects of different age, gender, and mating status including two BPH adult wing-morphology types. NlugCSP10 and 4 OBPs appeared to be antenna-specific because they were highly and differentially expressed in male and female antennae. NlugCSP11 was expressed ubiquitously, with particularly high expression in wings. The transcript levels of several olfactory genes depended on adult wing form, age, gender, and mating status, although no clear expression patterns were determined. Copyright © 2013 Elsevier Inc. All rights reserved.

The genomic and transcriptomic architecture of 2,000 breast tumours reveals novel subgroups.

PubMed

Curtis, Christina; Shah, Sohrab P; Chin, Suet-Feung; Turashvili, Gulisa; Rueda, Oscar M; Dunning, Mark J; Speed, Doug; Lynch, Andy G; Samarajiwa, Shamith; Yuan, Yinyin; Gräf, Stefan; Ha, Gavin; Haffari, Gholamreza; Bashashati, Ali; Russell, Roslin; McKinney, Steven; Langerød, Anita; Green, Andrew; Provenzano, Elena; Wishart, Gordon; Pinder, Sarah; Watson, Peter; Markowetz, Florian; Murphy, Leigh; Ellis, Ian; Purushotham, Arnie; Børresen-Dale, Anne-Lise; Brenton, James D; Tavaré, Simon; Caldas, Carlos; Aparicio, Samuel

2012-04-18

The elucidation of breast cancer subgroups and their molecular drivers requires integrated views of the genome and transcriptome from representative numbers of patients. We present an integrated analysis of copy number and gene expression in a discovery and validation set of 997 and 995 primary breast tumours, respectively, with long-term clinical follow-up. Inherited variants (copy number variants and single nucleotide polymorphisms) and acquired somatic copy number aberrations (CNAs) were associated with expression in ~40% of genes, with the landscape dominated by cis- and trans-acting CNAs. By delineating expression outlier genes driven in cis by CNAs, we identified putative cancer genes, including deletions in PPP2R2A, MTAP and MAP2K4. Unsupervised analysis of paired DNA–RNA profiles revealed novel subgroups with distinct clinical outcomes, which reproduced in the validation cohort. These include a high-risk, oestrogen-receptor-positive 11q13/14 cis-acting subgroup and a favourable prognosis subgroup devoid of CNAs. Trans-acting aberration hotspots were found to modulate subgroup-specific gene networks, including a TCR deletion-mediated adaptive immune response in the ‘CNA-devoid’ subgroup and a basal-specific chromosome 5 deletion-associated mitotic network. Our results provide a novel molecular stratification of the breast cancer population, derived from the impact of somatic CNAs on the transcriptome.

Population- and individual-specific regulatory variation in Sardinia.

PubMed

Pala, Mauro; Zappala, Zachary; Marongiu, Mara; Li, Xin; Davis, Joe R; Cusano, Roberto; Crobu, Francesca; Kukurba, Kimberly R; Gloudemans, Michael J; Reinier, Frederic; Berutti, Riccardo; Piras, Maria G; Mulas, Antonella; Zoledziewska, Magdalena; Marongiu, Michele; Sorokin, Elena P; Hess, Gaelen T; Smith, Kevin S; Busonero, Fabio; Maschio, Andrea; Steri, Maristella; Sidore, Carlo; Sanna, Serena; Fiorillo, Edoardo; Bassik, Michael C; Sawcer, Stephen J; Battle, Alexis; Novembre, John; Jones, Chris; Angius, Andrea; Abecasis, Gonçalo R; Schlessinger, David; Cucca, Francesco; Montgomery, Stephen B

2017-05-01

Genetic studies of complex traits have mainly identified associations with noncoding variants. To further determine the contribution of regulatory variation, we combined whole-genome and transcriptome data for 624 individuals from Sardinia to identify common and rare variants that influence gene expression and splicing. We identified 21,183 expression quantitative trait loci (eQTLs) and 6,768 splicing quantitative trait loci (sQTLs), including 619 new QTLs. We identified high-frequency QTLs and found evidence of selection near genes involved in malarial resistance and increased multiple sclerosis risk, reflecting the epidemiological history of Sardinia. Using family relationships, we identified 809 segregating expression outliers (median z score of 2.97), averaging 13.3 genes per individual. Outlier genes were enriched for proximal rare variants, providing a new approach to study large-effect regulatory variants and their relevance to traits. Our results provide insight into the effects of regulatory variants and their relationship to population history and individual genetic risk.

Molecular and chemical dialogues in bacteria-protozoa interactions.

PubMed

Song, Chunxu; Mazzola, Mark; Cheng, Xu; Oetjen, Janina; Alexandrov, Theodore; Dorrestein, Pieter; Watrous, Jeramie; van der Voort, Menno; Raaijmakers, Jos M

2015-08-06

Protozoan predation of bacteria can significantly affect soil microbial community composition and ecosystem functioning. Bacteria possess diverse defense strategies to resist or evade protozoan predation. For soil-dwelling Pseudomonas species, several secondary metabolites were proposed to provide protection against different protozoan genera. By combining whole-genome transcriptome analyses with (live) imaging mass spectrometry (IMS), we observed multiple changes in the molecular and chemical dialogues between Pseudomonas fluorescens and the protist Naegleria americana. Lipopeptide (LP) biosynthesis was induced in Pseudomonas upon protozoan grazing and LP accumulation transitioned from homogeneous distributions across bacterial colonies to site-specific accumulation at the bacteria-protist interface. Also putrescine biosynthesis was upregulated in P. fluorescens upon predation. We demonstrated that putrescine induces protozoan trophozoite encystment and adversely affects cyst viability. This multifaceted study provides new insights in common and strain-specific responses in bacteria-protozoa interactions, including responses that contribute to bacterial survival in highly competitive soil and rhizosphere environments.

Discrete domains of gene expression in germinal layers distinguish the development of gyrencephaly

PubMed Central

de Juan Romero, Camino; Bruder, Carl; Tomasello, Ugo; Sanz-Anquela, José Miguel; Borrell, Víctor

2015-01-01

Gyrencephalic species develop folds in the cerebral cortex in a stereotypic manner, but the genetic mechanisms underlying this patterning process are unknown. We present a large-scale transcriptomic analysis of individual germinal layers in the developing cortex of the gyrencephalic ferret, comparing between regions prospective of fold and fissure. We find unique transcriptional signatures in each germinal compartment, where thousands of genes are differentially expressed between regions, including ∼80% of genes mutated in human cortical malformations. These regional differences emerge from the existence of discrete domains of gene expression, which occur at multiple locations across the developing cortex of ferret and human, but not the lissencephalic mouse. Complex expression patterns emerge late during development and map the eventual location of folds or fissures. Protomaps of gene expression within germinal layers may contribute to define cortical folds or functional areas, but our findings demonstrate that they distinguish the development of gyrencephalic cortices. PMID:25916825

Deep functional analysis of synII, a 770-kilobase synthetic yeast chromosome.

PubMed

Shen, Yue; Wang, Yun; Chen, Tai; Gao, Feng; Gong, Jianhui; Abramczyk, Dariusz; Walker, Roy; Zhao, Hongcui; Chen, Shihong; Liu, Wei; Luo, Yisha; Müller, Carolin A; Paul-Dubois-Taine, Adrien; Alver, Bonnie; Stracquadanio, Giovanni; Mitchell, Leslie A; Luo, Zhouqing; Fan, Yanqun; Zhou, Baojin; Wen, Bo; Tan, Fengji; Wang, Yujia; Zi, Jin; Xie, Zexiong; Li, Bingzhi; Yang, Kun; Richardson, Sarah M; Jiang, Hui; French, Christopher E; Nieduszynski, Conrad A; Koszul, Romain; Marston, Adele L; Yuan, Yingjin; Wang, Jian; Bader, Joel S; Dai, Junbiao; Boeke, Jef D; Xu, Xun; Cai, Yizhi; Yang, Huanming

2017-03-10

Here, we report the successful design, construction, and characterization of a 770-kilobase synthetic yeast chromosome II (synII). Our study incorporates characterization at multiple levels-including phenomics, transcriptomics, proteomics, chromosome segregation, and replication analysis-to provide a thorough and comprehensive analysis of a synthetic chromosome. Our Trans-Omics analyses reveal a modest but potentially relevant pervasive up-regulation of translational machinery observed in synII, mainly caused by the deletion of 13 transfer RNAs. By both complementation assays and SCRaMbLE (synthetic chromosome rearrangement and modification by loxP -mediated evolution), we targeted and debugged the origin of a growth defect at 37°C in glycerol medium, which is related to misregulation of the high-osmolarity glycerol response. Despite the subtle differences, the synII strain shows highly consistent biological processes comparable to the native strain. Copyright © 2017, American Association for the Advancement of Science.

In silico Pathway Activation Network Decomposition Analysis (iPANDA) as a method for biomarker development.

PubMed

Ozerov, Ivan V; Lezhnina, Ksenia V; Izumchenko, Evgeny; Artemov, Artem V; Medintsev, Sergey; Vanhaelen, Quentin; Aliper, Alexander; Vijg, Jan; Osipov, Andreyan N; Labat, Ivan; West, Michael D; Buzdin, Anton; Cantor, Charles R; Nikolsky, Yuri; Borisov, Nikolay; Irincheeva, Irina; Khokhlovich, Edward; Sidransky, David; Camargo, Miguel Luiz; Zhavoronkov, Alex

2016-11-16

Signalling pathway activation analysis is a powerful approach for extracting biologically relevant features from large-scale transcriptomic and proteomic data. However, modern pathway-based methods often fail to provide stable pathway signatures of a specific phenotype or reliable disease biomarkers. In the present study, we introduce the in silico Pathway Activation Network Decomposition Analysis (iPANDA) as a scalable robust method for biomarker identification using gene expression data. The iPANDA method combines precalculated gene coexpression data with gene importance factors based on the degree of differential gene expression and pathway topology decomposition for obtaining pathway activation scores. Using Microarray Analysis Quality Control (MAQC) data sets and pretreatment data on Taxol-based neoadjuvant breast cancer therapy from multiple sources, we demonstrate that iPANDA provides significant noise reduction in transcriptomic data and identifies highly robust sets of biologically relevant pathway signatures. We successfully apply iPANDA for stratifying breast cancer patients according to their sensitivity to neoadjuvant therapy.

Ensemble analyses improve signatures of tumour hypoxia and reveal inter-platform differences

PubMed Central

2014-01-01

Background The reproducibility of transcriptomic biomarkers across datasets remains poor, limiting clinical application. We and others have suggested that this is in-part caused by differential error-structure between datasets, and their incomplete removal by pre-processing algorithms. Methods To test this hypothesis, we systematically assessed the effects of pre-processing on biomarker classification using 24 different pre-processing methods and 15 distinct signatures of tumour hypoxia in 10 datasets (2,143 patients). Results We confirm strong pre-processing effects for all datasets and signatures, and find that these differ between microarray versions. Importantly, exploiting different pre-processing techniques in an ensemble technique improved classification for a majority of signatures. Conclusions Assessing biomarkers using an ensemble of pre-processing techniques shows clear value across multiple diseases, datasets and biomarkers. Importantly, ensemble classification improves biomarkers with initially good results but does not result in spuriously improved performance for poor biomarkers. While further research is required, this approach has the potential to become a standard for transcriptomic biomarkers. PMID:24902696

Streptococcus pneumoniae Supragenome Hybridization Arrays for Profiling of Genetic Content and Gene Expression.

PubMed

Kadam, Anagha; Janto, Benjamin; Eutsey, Rory; Earl, Joshua P; Powell, Evan; Dahlgren, Margaret E; Hu, Fen Z; Ehrlich, Garth D; Hiller, N Luisa

2015-02-02

There is extensive genomic diversity among Streptococcus pneumoniae isolates. Approximately half of the comprehensive set of genes in the species (the supragenome or pangenome) is present in all the isolates (core set), and the remaining is unevenly distributed among strains (distributed set). The Streptococcus pneumoniae Supragenome Hybridization (SpSGH) array provides coverage for an extensive set of genes and polymorphisms encountered within this species, capturing this genomic diversity. Further, the capture is quantitative. In this manner, the SpSGH array allows for both genomic and transcriptomic analyses of diverse S. pneumoniae isolates on a single platform. In this unit, we present the SpSGH array, and describe in detail its design and implementation for both genomic and transcriptomic analyses. The methodology can be applied to construction and modification of SpSGH array platforms, as well to other bacterial species as long as multiple whole-genome sequences are available that collectively capture the vast majority of the species supragenome. Copyright © 2015 John Wiley & Sons, Inc.

Comprehensive discovery of noncoding RNAs in acute myeloid leukemia cell transcriptomes.

PubMed

Zhang, Jin; Griffith, Malachi; Miller, Christopher A; Griffith, Obi L; Spencer, David H; Walker, Jason R; Magrini, Vincent; McGrath, Sean D; Ly, Amy; Helton, Nichole M; Trissal, Maria; Link, Daniel C; Dang, Ha X; Larson, David E; Kulkarni, Shashikant; Cordes, Matthew G; Fronick, Catrina C; Fulton, Robert S; Klco, Jeffery M; Mardis, Elaine R; Ley, Timothy J; Wilson, Richard K; Maher, Christopher A

2017-11-01

To detect diverse and novel RNA species comprehensively, we compared deep small RNA and RNA sequencing (RNA-seq) methods applied to a primary acute myeloid leukemia (AML) sample. We were able to discover previously unannotated small RNAs using deep sequencing of a library method using broader insert size selection. We analyzed the long noncoding RNA (lncRNA) landscape in AML by comparing deep sequencing from multiple RNA-seq library construction methods for the sample that we studied and then integrating RNA-seq data from 179 AML cases. This identified lncRNAs that are completely novel, differentially expressed, and associated with specific AML subtypes. Our study revealed the complexity of the noncoding RNA transcriptome through a combined strategy of strand-specific small RNA and total RNA-seq. This dataset will serve as an invaluable resource for future RNA-based analyses. Copyright © 2017 ISEH – Society for Hematology and Stem Cells. Published by Elsevier Inc. All rights reserved.

Integrative approaches for large-scale transcriptome-wide association studies

PubMed Central

Gusev, Alexander; Ko, Arthur; Shi, Huwenbo; Bhatia, Gaurav; Chung, Wonil; Penninx, Brenda W J H; Jansen, Rick; de Geus, Eco JC; Boomsma, Dorret I; Wright, Fred A; Sullivan, Patrick F; Nikkola, Elina; Alvarez, Marcus; Civelek, Mete; Lusis, Aldons J.; Lehtimäki, Terho; Raitoharju, Emma; Kähönen, Mika; Seppälä, Ilkka; Raitakari, Olli T.; Kuusisto, Johanna; Laakso, Markku; Price, Alkes L.; Pajukanta, Päivi; Pasaniuc, Bogdan

2016-01-01

Many genetic variants influence complex traits by modulating gene expression, thus altering the abundance levels of one or multiple proteins. Here, we introduce a powerful strategy that integrates gene expression measurements with summary association statistics from large-scale genome-wide association studies (GWAS) to identify genes whose cis-regulated expression is associated to complex traits. We leverage expression imputation to perform a transcriptome wide association scan (TWAS) to identify significant expression-trait associations. We applied our approaches to expression data from blood and adipose tissue measured in ~3,000 individuals overall. We imputed gene expression into GWAS data from over 900,000 phenotype measurements to identify 69 novel genes significantly associated to obesity-related traits (BMI, lipids, and height). Many of the novel genes are associated with relevant phenotypes in the Hybrid Mouse Diversity Panel. Our results showcase the power of integrating genotype, gene expression and phenotype to gain insights into the genetic basis of complex traits. PMID:26854917

In silico Pathway Activation Network Decomposition Analysis (iPANDA) as a method for biomarker development

PubMed Central

Ozerov, Ivan V.; Lezhnina, Ksenia V.; Izumchenko, Evgeny; Artemov, Artem V.; Medintsev, Sergey; Vanhaelen, Quentin; Aliper, Alexander; Vijg, Jan; Osipov, Andreyan N.; Labat, Ivan; West, Michael D.; Buzdin, Anton; Cantor, Charles R.; Nikolsky, Yuri; Borisov, Nikolay; Irincheeva, Irina; Khokhlovich, Edward; Sidransky, David; Camargo, Miguel Luiz; Zhavoronkov, Alex

2016-01-01

Signalling pathway activation analysis is a powerful approach for extracting biologically relevant features from large-scale transcriptomic and proteomic data. However, modern pathway-based methods often fail to provide stable pathway signatures of a specific phenotype or reliable disease biomarkers. In the present study, we introduce the in silico Pathway Activation Network Decomposition Analysis (iPANDA) as a scalable robust method for biomarker identification using gene expression data. The iPANDA method combines precalculated gene coexpression data with gene importance factors based on the degree of differential gene expression and pathway topology decomposition for obtaining pathway activation scores. Using Microarray Analysis Quality Control (MAQC) data sets and pretreatment data on Taxol-based neoadjuvant breast cancer therapy from multiple sources, we demonstrate that iPANDA provides significant noise reduction in transcriptomic data and identifies highly robust sets of biologically relevant pathway signatures. We successfully apply iPANDA for stratifying breast cancer patients according to their sensitivity to neoadjuvant therapy. PMID:27848968

Genomic and transcriptomic analysis of NDM-1 Klebsiella pneumoniae in spaceflight reveal mechanisms underlying environmental adaptability

PubMed Central

Li, Jia; Liu, Fei; Wang, Qi; Ge, Pupu; Woo, Patrick C. Y.; Yan, Jinghua; Zhao, Yanlin; Gao, George F.; Liu, Cui Hua; Liu, Changting

2014-01-01

The emergence and rapid spread of New Delhi Metallo-beta-lactamase-1 (NDM-1)-producing Klebsiella pneumoniae strains has caused a great concern worldwide. To better understand the mechanisms underlying environmental adaptation of those highly drug-resistant K. pneumoniae strains, we took advantage of the China's Shenzhou 10 spacecraft mission to conduct comparative genomic and transcriptomic analysis of a NDM-1 K. pneumoniae strain (ATCC BAA-2146) being cultivated under different conditions. The samples were recovered from semisolid medium placed on the ground (D strain), in simulated space condition (M strain), or in Shenzhou 10 spacecraft (T strain) for analysis. Our data revealed multiple variations underlying pathogen adaptation into different environments in terms of changes in morphology, H2O2 tolerance and biofilm formation ability, genomic stability and regulation of metabolic pathways. Additionally, we found a few non-coding RNAs to be differentially regulated. The results are helpful for better understanding the adaptive mechanisms of drug-resistant bacterial pathogens. PMID:25163721

A Single-Cell Roadmap of Lineage Bifurcation in Human ESC Models of Embryonic Brain Development.

PubMed

Yao, Zizhen; Mich, John K; Ku, Sherman; Menon, Vilas; Krostag, Anne-Rachel; Martinez, Refugio A; Furchtgott, Leon; Mulholland, Heather; Bort, Susan; Fuqua, Margaret A; Gregor, Ben W; Hodge, Rebecca D; Jayabalu, Anu; May, Ryan C; Melton, Samuel; Nelson, Angelique M; Ngo, N Kiet; Shapovalova, Nadiya V; Shehata, Soraya I; Smith, Michael W; Tait, Leah J; Thompson, Carol L; Thomsen, Elliot R; Ye, Chaoyang; Glass, Ian A; Kaykas, Ajamete; Yao, Shuyuan; Phillips, John W; Grimley, Joshua S; Levi, Boaz P; Wang, Yanling; Ramanathan, Sharad

2017-01-05

During human brain development, multiple signaling pathways generate diverse cell types with varied regional identities. Here, we integrate single-cell RNA sequencing and clonal analyses to reveal lineage trees and molecular signals underlying early forebrain and mid/hindbrain cell differentiation from human embryonic stem cells (hESCs). Clustering single-cell transcriptomic data identified 41 distinct populations of progenitor, neuronal, and non-neural cells across our differentiation time course. Comparisons with primary mouse and human gene expression data demonstrated rostral and caudal progenitor and neuronal identities from early brain development. Bayesian analyses inferred a unified cell-type lineage tree that bifurcates between cortical and mid/hindbrain cell types. Two methods of clonal analyses confirmed these findings and further revealed the importance of Wnt/β-catenin signaling in controlling this lineage decision. Together, these findings provide a rich transcriptome-based lineage map for studying human brain development and modeling developmental disorders. Copyright © 2017 Elsevier Inc. All rights reserved.

Engineered reversal of drug resistance in cancer cells--metastases suppressor factors as change agents.

PubMed

Yadav, Vinod Kumar; Kumar, Akinchan; Mann, Anita; Aggarwal, Suruchi; Kumar, Maneesh; Roy, Sumitabho Deb; Pore, Subrata Kumar; Banerjee, Rajkumar; Mahesh Kumar, Jerald; Thakur, Ram Krishna; Chowdhury, Shantanu

2014-01-01

Building molecular correlates of drug resistance in cancer and exploiting them for therapeutic intervention remains a pressing clinical need. To identify factors that impact drug resistance herein we built a model that couples inherent cell-based response toward drugs with transcriptomes of resistant/sensitive cells. To test this model, we focused on a group of genes called metastasis suppressor genes (MSGs) that influence aggressiveness and metastatic potential of cancers. Interestingly, modeling of 84 000 drug response transcriptome combinations predicted multiple MSGs to be associated with resistance of different cell types and drugs. As a case study, on inducing MSG levels in a drug resistant breast cancer line resistance to anticancer drugs caerulomycin, camptothecin and topotecan decreased by more than 50-60%, in both culture conditions and also in tumors generated in mice, in contrast to control un-induced cells. To our knowledge, this is the first demonstration of engineered reversal of drug resistance in cancer cells based on a model that exploits inherent cellular response profiles.

Identification of the RNA recognition element of the RBPMS family of RNA-binding proteins and their transcriptome-wide mRNA targets

PubMed Central

Farazi, Thalia A.; Leonhardt, Carl S.; Mukherjee, Neelanjan; Mihailovic, Aleksandra; Li, Song; Max, Klaas E.A.; Meyer, Cindy; Yamaji, Masashi; Cekan, Pavol; Jacobs, Nicholas C.; Gerstberger, Stefanie; Bognanni, Claudia; Larsson, Erik; Ohler, Uwe; Tuschl, Thomas

2014-01-01

Recent studies implicated the RNA-binding protein with multiple splicing (RBPMS) family of proteins in oocyte, retinal ganglion cell, heart, and gastrointestinal smooth muscle development. These RNA-binding proteins contain a single RNA recognition motif (RRM), and their targets and molecular function have not yet been identified. We defined transcriptome-wide RNA targets using photoactivatable-ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) in HEK293 cells, revealing exonic mature and intronic pre-mRNA binding sites, in agreement with the nuclear and cytoplasmic localization of the proteins. Computational and biochemical approaches defined the RNA recognition element (RRE) as a tandem CAC trinucleotide motif separated by a variable spacer region. Similar to other mRNA-binding proteins, RBPMS family of proteins relocalized to cytoplasmic stress granules under oxidative stress conditions suggestive of a support function for mRNA localization in large and/or multinucleated cells where it is preferentially expressed. PMID:24860013

Natural genetic variation profoundly regulates gene expression in immune cells and dictates susceptibility to CNS autoimmunity

PubMed Central

Bearoff, Frank; del Rio, Roxana; Case, Laure K.; Dragon, Julie A.; Nguyen-Vu, Trang; Lin, Chin-Yo; Blankenhorn, Elizabeth P.; Teuscher, Cory; Krementsov, Dimitry N.

2016-01-01

Regulation of gene expression in immune cells is known to be under genetic control, and likely contributes to susceptibility to autoimmune diseases, such as multiple sclerosis (MS). How this occurs in concert across multiple immune cell types is poorly understood. Using a mouse model that harnesses the genetic diversity of wild-derived mice, more accurately reflecting genetically diverse human populations, we provide an extensive characterization of the genetic regulation of gene expression in five different naïve immune cell types relevant to MS. The immune cell transcriptome is shown to be under profound genetic control, exhibiting diverse patterns: global, cell-specific, and sex-specific. Bioinformatic analysis of the genetically-controlled transcript networks reveals reduced cell type-specificity and inflammatory activity in wild-derived PWD/PhJ mice, compared with the conventional laboratory strain C57BL/6J. Additionally, candidate MS-GWAS genes were significantly enriched among transcripts overrepresented in C57BL/6J cells compared to PWD. These expression level differences correlate with robust differences in susceptibility to experimental autoimmune encephalomyelitis, the principal model of MS, and skewing of the encephalitogenic T cell responses. Taken together, our results provide functional insights into the genetic regulation of the immune transcriptome, and shed light on how this in turn contributes to susceptibility to autoimmune disease. PMID:27653816

Identification of Mild Freezing Shock Response Pathways in Barley Based on Transcriptome Profiling.

PubMed

Wang, Xiaolei; Wu, Dezhi; Yang, Qian; Zeng, Jianbin; Jin, Gulei; Chen, Zhong-Hua; Zhang, Guoping; Dai, Fei

2016-01-01

Low temperature is a major abiotic stress affecting crop growth and productivity. A better understanding of low temperature tolerance mechanisms is imperative for developing the crop cultivars with improved tolerance. We herein performed an Illumina RNA-sequencing experiment using two barley genotypes differing in freezing tolerance (Nure, tolerant and Tremois, sensitive), to determine the transcriptome profiling and genotypic difference under mild freezing shock treatment after a very short acclimation for gene induction. A total of 6474 differentially expressed genes, almost evenly distributed on the seven chromosomes, were identified. The key DEGs could be classified into six signaling pathways, i.e., Ca(2+) signaling, PtdOH signaling, CBFs pathway, ABA pathway, jasmonate pathway, and amylohydrolysis pathway. Expression values of DEGs in multiple signaling pathways were analyzed and a hypothetical model of mild freezing shock tolerance mechanism was proposed. Expression and sequence profile of HvCBFs cluster within Frost resistance-H2, a major quantitative trait locus on 5H being closely related to low temperature tolerance in barley, were further illustrated, considering the crucial role of HvCBFs on freezing tolerance. It may be concluded that multiple signaling pathways are activated in concert when barley is exposed to mild freezing shock. The pathway network we presented may provide a platform for further exploring the functions of genes involved in low temperature tolerance in barley.