Comparative genomics of bdelloid rotifers: Insights from desiccating and nondesiccating species.

Bdelloid rotifers are a class of microscopic invertebrates that have existed for millions of years apparently without sex or meiosis. They inhabit a variety of temporary and permanent freshwater habitats globally, and many species are remarkably tolerant of desiccation. Bdelloids offer an opportunity to better understand the evolution of sex and recombination, but previous work has emphasised desiccation as the cause of several unusual genomic features in this group. Here, we present high-quality whole-genome sequences of 3 bdelloid species: Rotaria macrura and R. magnacalcarata, which are both desiccation intolerant, and Adineta ricciae, which is desiccation tolerant. In combination with the published assembly of A. vaga, which is also desiccation tolerant, we apply a comparative genomics approach to evaluate the potential effects of desiccation tolerance and asexuality on genome evolution in bdelloids. We find that ancestral tetraploidy is conserved among all 4 bdelloid species, but homologous divergence in obligately aquatic Rotaria genomes is unexpectedly low. This finding is contrary to current models regarding the role of desiccation in shaping bdelloid genomes. In addition, we find that homologous regions in A. ricciae are largely collinear and do not form palindromic repeats as observed in the published A. vaga assembly. Consequently, several features interpreted as genomic evidence for long-term ameiotic evolution are not general to all bdelloid species, even within the same genus. Finally, we substantiate previous findings of high levels of horizontally transferred nonmetazoan genes in both desiccating and nondesiccating bdelloid species and show that this unusual feature is not shared by other animal phyla, even those with desiccation-tolerant representatives. These comparisons call into question the proposed role of desiccation in mediating horizontal genetic transfer.

Insights into olfactory ensheathing cell development from a laser-microdissection and transcriptome-profiling approach.

Olfactory ensheathing cells (OECs) are neural crest-derived glia that ensheath bundles of olfactory axons from their peripheral origins in the olfactory epithelium to their central targets in the olfactory bulb. We took an unbiased laser microdissection and differential RNA-seq approach, validated by in situ hybridization, to identify candidate molecular mechanisms underlying mouse OEC development and differences with the neural crest-derived Schwann cells developing on other peripheral nerves. We identified 25 novel markers for developing OECs in the olfactory mucosa and/or the olfactory nerve layer surrounding the olfactory bulb, of which 15 were OEC-specific (that is, not expressed by Schwann cells). One pan-OEC-specific gene, Ptprz1, encodes a receptor-like tyrosine phosphatase that blocks oligodendrocyte differentiation. Mutant analysis suggests Ptprz1 may also act as a brake on OEC differentiation, and that its loss disrupts olfactory axon targeting. Overall, our results provide new insights into OEC development and the diversification of neural crest-derived glia.

InterMineR: an R package for InterMine databases.

SUMMARY: InterMineR is a package designed to provide a flexible interface between the R programming environment and biological databases built using the InterMine platform. The package offers access to the flexible query builder and the library of term enrichment tools of the InterMine framework, as well as interoperability with other Bioconductor packages. This facilitates automation of data retrieval tasks as well as downstream analysis with existing statistical tools in the R environment. AVAILABILITY AND IMPLEMENTATION: InterMineR is free and open source, released under the LGPL licence and available from the Bioconductor project and Github (https://bioconductor.org/packages/release/bioc/html/InterMineR.html, https://github.com/intermine/interMineR). SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

ComplexViewer: visualization of curated macromolecular complexes.

SUMMARY: Proteins frequently function as parts of complexes, assemblages of multiple proteins and other biomolecules, yet network visualizations usually only show proteins as parts of binary interactions. ComplexViewer visualizes interactions with more than two participants and thereby avoids the need to first expand these into multiple binary interactions. Furthermore, if binding regions between molecules are known then these can be displayed in the context of the larger complex. AVAILABILITY AND IMPLEMENTATION: freely available under Apache version 2 license; EMBL-EBI Complex Portal: http://www.ebi.ac.uk/complexportal; Source code: https://github.com/MICommunity/ComplexViewer; Package: https://www.npmjs.com/package/complexviewer; http://biojs.io/d/complexviewer. Language: JavaScript; Web technology: Scalable Vector Graphics; Libraries: D3.js. CONTACT: colin.combe@ed.ac.uk or juri.rappsilber@ed.ac.uk.

Empirical Bayes method for reducing false discovery rates of correlation matrices with block diagonal structure.

BACKGROUND: Correlation matrices are important in inferring relationships and networks between regulatory or signalling elements in biological systems. With currently available technology sample sizes for experiments are typically small, meaning that these correlations can be difficult to estimate. At a genome-wide scale estimation of correlation matrices can also be computationally demanding. RESULTS: We develop an empirical Bayes approach to improve covariance estimates for gene expression, where we assume the covariance matrix takes a block diagonal form. Our method shows lower false discovery rates than existing methods on simulated data. Applied to a real data set from Bacillus subtilis we demonstrate it's ability to detecting known regulatory units and interactions between them. CONCLUSIONS: We demonstrate that, compared to existing methods, our method is able to find significant covariances and also to control false discovery rates, even when the sample size is small (n=10). The method can be used to find potential regulatory networks, and it may also be used as a pre-processing step for methods that calculate, for example, partial correlations, so enabling the inference of the causal and hierarchical structure of the networks.

ThaleMine: A Warehouse for Arabidopsis Data Integration and Discovery.

ThaleMine (https://apps.araport.org/thalemine/) is a comprehensive data warehouse that integrates a wide array of genomic information of the model plant Arabidopsis thaliana. The data collection currently includes the latest structural and functional annotation from the Araport11 update, the Col-0 genome sequence, RNA-seq and array expression, co-expression, protein interactions, homologs, pathways, publications, alleles, germplasm and phenotypes. The data are collected from a wide variety of public resources. Users can browse gene-specific data through Gene Report pages, identify and create gene lists based on experiments or indexed keywords, and run GO enrichment analysis to investigate the biological significance of selected gene sets. Developed by the Arabidopsis Information Portal project (Araport, https://www.araport.org/), ThaleMine uses the InterMine software framework, which builds well-structured data, and provides powerful data query and analysis functionality. The warehoused data can be accessed by users via graphical interfaces, as well as programmatically via web-services. Here we describe recent developments in ThaleMine including new features and extensions, and discuss future improvements. InterMine has been broadly adopted by the model organism research community including nematode, rat, mouse, zebrafish, budding yeast, the modENCODE project, as well as being used for human data. ThaleMine is the first InterMine developed for a plant model. As additional new plant InterMines are developed by the legume and other plant research communities, the potential of cross-organism integrative data analysis will be further enabled.

Araport: the Arabidopsis information portal.

The Arabidopsis Information Portal (https://www.araport.org) is a new online resource for plant biology research. It houses the Arabidopsis thaliana genome sequence and associated annotation. It was conceived as a framework that allows the research community to develop and release 'modules' that integrate, analyze and visualize Arabidopsis data that may reside at remote sites. The current implementation provides an indexed database of core genomic information. These data are made available through feature-rich web applications that provide search, data mining, and genome browser functionality, and also by bulk download and web services. Araport uses software from the InterMine and JBrowse projects to expose curated data from TAIR, GO, BAR, EBI, UniProt, PubMed and EPIC CoGe. The site also hosts 'science apps,' developed as prototypes for community modules that use dynamic web pages to present data obtained on-demand from third-party servers via RESTful web services. Designed for sustainability, the Arabidopsis Information Portal strategy exploits existing scientific computing infrastructure, adopts a practical mixture of data integration technologies and encourages collaborative enhancement of the resource by its user community.

InterMine: extensive web services for modern biology.

InterMine (www.intermine.org) is a biological data warehousing system providing extensive automatically generated and configurable RESTful web services that underpin the web interface and can be re-used in many other applications: to find and filter data; export it in a flexible and structured way; to upload, use, manipulate and analyze lists; to provide services for flexible retrieval of sequence segments, and for other statistical and analysis tools. Here we describe these features and discuss how they can be used separately or in combinations to support integrative and comparative analysis.

Cross-organism analysis using InterMine.

InterMine is a data integration warehouse and analysis software system developed for large and complex biological data sets. Designed for integrative analysis, it can be accessed through a user-friendly web interface. For bioinformaticians, extensive web services as well as programming interfaces for most common scripting languages support access to all features. The web interface includes a useful identifier look-up system, and both simple and sophisticated search options. Interactive results tables enable exploration, and data can be filtered, summarized, and browsed. A set of graphical analysis tools provide a rich environment for data exploration including statistical enrichment of sets of genes or other entities. InterMine databases have been developed for the major model organisms, budding yeast, nematode worm, fruit fly, zebrafish, mouse, and rat together with a newly developed human database. Here, we describe how this has facilitated interoperation and development of cross-organism analysis tools and reports. InterMine as a data exploration and analysis tool is also described. All the InterMine-based systems described in this article are resources freely available to the scientific community.

Activity of a heptad of transcription factors is associated with stem cell programs and clinical outcome in acute myeloid leukemia.

Aberrant transcriptional programs in combination with abnormal proliferative signaling drive leukemic transformation. These programs operate in normal hematopoiesis where they are involved in hematopoietic stem cell (HSC) proliferation and maintenance. Ets Related Gene (ERG) is a component of normal and leukemic stem cell signatures and high ERG expression is a risk factor for poor prognosis in acute myeloid leukemia (AML). However, mechanisms that underlie ERG expression in AML and how its expression relates to leukemic stemness are unknown. We report that ERG expression in AML is associated with activity of the ERG promoters and +85 stem cell enhancer and a heptad of transcription factors that combinatorially regulate genes in HSCs. Gene expression signatures derived from ERG promoter-stem cell enhancer and heptad activity are associated with clinical outcome when ERG expression alone fails. We also show that the heptad signature is associated with AMLs that lack somatic mutations in NPM1 and confers an adverse prognosis when associated with FLT3 mutations. Taken together, these results suggest that transcriptional regulators cooperate to establish or maintain primitive stem cell-like signatures in leukemic cells and that the underlying pattern of somatic mutations contributes to the development of these signatures and modulate their influence on clinical outcome.

Biochemical diversification through foreign gene expression in bdelloid rotifers.

Bdelloid rotifers are microinvertebrates with unique characteristics: they have survived tens of millions of years without sexual reproduction; they withstand extreme desiccation by undergoing anhydrobiosis; and they tolerate very high levels of ionizing radiation. Recent evidence suggests that subtelomeric regions of the bdelloid genome contain sequences originating from other organisms by horizontal gene transfer (HGT), of which some are known to be transcribed. However, the extent to which foreign gene expression plays a role in bdelloid physiology is unknown. We address this in the first large scale analysis of the transcriptome of the bdelloid Adineta ricciae: cDNA libraries from hydrated and desiccated bdelloids were subjected to massively parallel sequencing and assembled transcripts compared against the UniProtKB database by blastx to identify their putative products. Of ~29,000 matched transcripts, ~10% were inferred from blastx matches to be horizontally acquired, mainly from eubacteria but also from fungi, protists, and algae. After allowing for possible sources of error, the rate of HGT is at least 8%-9%, a level significantly higher than other invertebrates. We verified their foreign nature by phylogenetic analysis and by demonstrating linkage of foreign genes with metazoan genes in the bdelloid genome. Approximately 80% of horizontally acquired genes expressed in bdelloids code for enzymes, and these represent 39% of enzymes in identified pathways. Many enzymes encoded by foreign genes enhance biochemistry in bdelloids compared to other metazoans, for example, by potentiating toxin degradation or generation of antioxidants and key metabolites. They also supplement, and occasionally potentially replace, existing metazoan functions. Bdelloid rotifers therefore express horizontally acquired genes on a scale unprecedented in animals, and foreign genes make a profound contribution to their metabolism. This represents a potential mechanism for ancient asexuals to adapt rapidly to changing environments and thereby persist over long evolutionary time periods in the absence of sex.

modMine: flexible access to modENCODE data.

In an effort to comprehensively characterize the functional elements within the genomes of the important model organisms Drosophila melanogaster and Caenorhabditis elegans, the NHGRI model organism Encyclopaedia of DNA Elements (modENCODE) consortium has generated an enormous library of genomic data along with detailed, structured information on all aspects of the experiments. The modMine database (http://intermine.modencode.org) described here has been built by the modENCODE Data Coordination Center to allow the broader research community to (i) search for and download data sets of interest among the thousands generated by modENCODE; (ii) access the data in an integrated form together with non-modENCODE data sets; and (iii) facilitate fine-grained analysis of the above data. The sophisticated search features are possible because of the collection of extensive experimental metadata by the consortium. Interfaces are provided to allow both biologists and bioinformaticians to exploit these rich modENCODE data sets now available via modMine.

Identification of multiple transcription factor genes potentially involved in the development of electrosensory versus mechanosensory lateral line organs.

In electroreceptive jawed vertebrates, embryonic lateral line placodes give rise to electrosensory ampullary organs as well as mechanosensory neuromasts. Previous reports of shared gene expression suggest that conserved mechanisms underlie electroreceptor and mechanosensory hair cell development and that electroreceptors evolved as a transcriptionally related "sister cell type" to hair cells. We previously identified only one transcription factor gene, Neurod4, as ampullary organ-restricted in the developing lateral line system of a chondrostean ray-finned fish, the Mississippi paddlefish (Polyodon spathula). The other 16 transcription factor genes we previously validated in paddlefish were expressed in both ampullary organs and neuromasts. Here, we used our published lateral line organ-enriched gene-set (arising from differential bulk RNA-seq in late-larval paddlefish), together with a candidate gene approach, to identify 25 transcription factor genes expressed in the developing lateral line system of a more experimentally tractable chondrostean, the sterlet (Acipenser ruthenus, a small sturgeon), and/or that of paddlefish. Thirteen are expressed in both ampullary organs and neuromasts, consistent with conservation of molecular mechanisms. Seven are electrosensory-restricted on the head (Irx5, Irx3, Insm1, Sp5, Satb2, Mafa and Rorc), and five are the first-reported mechanosensory-restricted transcription factor genes (Foxg1, Sox8, Isl1, Hmx2 and Rorb). However, as previously reported, Sox8 is expressed in ampullary organs as well as neuromasts in a catshark (Scyliorhinus canicula), suggesting the existence of lineage-specific differences between cartilaginous and ray-finned fishes. Overall, our results support the hypothesis that ampullary organs and neuromasts develop via largely conserved transcriptional mechanisms, and identify multiple transcription factors potentially involved in the formation of electrosensory versus mechanosensory lateral line organs.

Integrating microRNA and mRNA expression profiling in Symbiodinium microadriaticum, a dinoflagellate symbiont of reef-building corals.

BACKGROUND: Animal and plant genomes produce numerous small RNAs (smRNAs) that regulate gene expression post-transcriptionally affecting metabolism, development, and epigenetic inheritance. In order to characterize the repertoire of endogenous smRNAs and potential gene targets in dinoflagellates, we conducted smRNA and mRNA expression profiling over 9 experimental treatments of cultures from Symbiodinium microadriaticum, a photosynthetic symbiont of scleractinian corals. RESULTS: We identified a set of 21 novel smRNAs that share stringent key features with functional microRNAs from other model organisms. smRNAs were predicted independently over all 9 treatments and their putative gene targets were identified. We found 1,720 animal-like target sites in the 3'UTRs of 12,858 mRNAs and 19 plant-like target sites in 51,917 genes. We assembled a transcriptome of 58,649 genes and determined differentially expressed genes (DEGs) between treatments. Heat stress was found to produce a much larger number of DEGs than other treatments that yielded only few DEGs. Analysis of DEGs also revealed that minicircle-encoded photosynthesis proteins seem to be common targets of transcriptional regulation. Furthermore, we identified the core RNAi protein machinery in Symbiodinium. CONCLUSIONS: Integration of smRNA and mRNA expression profiling identified a variety of processes that could be under microRNA control, e.g. protein modification, signaling, gene expression, and response to DNA damage. Given that Symbiodinium seems to have a paucity of transcription factors and differentially expressed genes, identification and characterization of its smRNA repertoire establishes the possibility of a range of gene regulatory mechanisms in dinoflagellates acting post-transcriptionally.

InterMine: a flexible data warehouse system for the integration and analysis of heterogeneous biological data.

SUMMARY: InterMine is an open-source data warehouse system that facilitates the building of databases with complex data integration requirements and a need for a fast customizable query facility. Using InterMine, large biological databases can be created from a range of heterogeneous data sources, and the extensible data model allows for easy integration of new data types. The analysis tools include a flexible query builder, genomic region search and a library of 'widgets' performing various statistical analyses. The results can be exported in many commonly used formats. InterMine is a fully extensible framework where developers can add new tools and functionality. Additionally, there is a comprehensive set of web services, for which client libraries are provided in five commonly used programming languages. AVAILABILITY: Freely available from http://www.intermine.org under the LGPL license. CONTACT: g.micklem@gen.cam.ac.uk SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Transcriptional landscape of oncogene-induced senescence: a machine learning-based meta-analytic approach.

Oncogene-induced senescence (OIS) is highly heterogeneous, varying by oncogenic signals and cellular context. While its dual role, in the initial inhibition potentially later leading to promotion of tumors through the senescence-associated secretory phenotype, is still a matter of debate, it is undeniable that OIS is critical to understanding tumorigenesis. A major obstacle to OIS research is the absence of a universally accepted marker. Here, we present a robust OIS-specific transcriptomic secretory phenotype, termed oncogene-induced senescence secretory phenotype (OIS-SP), which can identify OIS across multiple biological contexts from in vitro datasets to in vivo human samples. We apply a meta-analytic machine learning pipeline to harmonize a deliberately varied selection of Ras-Raf-MEK-induced senescence datasets of differing origins, oncogenic signals and cell types. Finally we make use of bypass data to identify key genes and eliminate genes associated with quiescence, so identifying 40 OIS-SP genes. Within this set, we determined a robust core of five OIS-SP genes (FBLN1, CXCL12, EREG, CST1 and MMP10). Importantly, these 5 OIS-SP genes showed clear, consistent regulation patterns across various human Ras-Raf-MEK-mutated tumor tissues, which suggests that OIS-SP may be a novel cancer driver phenotype with an unexpectedly critical role in tumorigenesis.

Multiple functionally divergent and conserved copies of alpha tubulin in bdelloid rotifers.

BACKGROUND: Bdelloid rotifers are microscopic animals that have apparently survived without sex for millions of years and are able to survive desiccation at all life stages through a process called anhydrobiosis. Both of these characteristics are believed to have played a role in shaping several unusual features of bdelloid genomes discovered in recent years. Studies into the impact of asexuality and anhydrobiosis on bdelloid genomes have focused on understanding gene copy number. Here we investigate copy number and sequence divergence in alpha tubulin. Alpha tubulin is conserved and normally present in low copy numbers in animals, but multiplication of alpha tubulin copies has occurred in animals adapted to extreme environments, such as cold-adapted Antarctic fish. Using cloning and sequencing we compared alpha tubulin copy variation in four species of bdelloid rotifers and four species of monogonont rotifers, which are facultatively sexual and cannot survive desiccation as adults. Results were verified using transcriptome data from one bdelloid species, Adineta ricciae. RESULTS: In common with the typical pattern for animals, monogonont rotifers contain either one or two copies of alpha tubulin, but bdelloid species contain between 11 and 13 different copies, distributed across five classes. Approximately half of the copies form a highly conserved group that vary by only 1.1% amino acid pairwise divergence with each other and with the monogonont copies. The other copies have divergent amino acid sequences that evolved significantly faster between classes than within them, relative to synonymous changes, and vary in predicted biochemical properties. Copies of each class were expressed under the laboratory conditions used to construct the transcriptome. CONCLUSIONS: Our findings are consistent with recent evidence that bdelloids are degenerate tetraploids and that functional divergence of ancestral copies of genes has occurred, but show how further duplication events in the ancestor of bdelloids led to proliferation in both conserved and functionally divergent copies of this gene.

HumanMine: advanced data searching, analysis and cross-species comparison.

HumanMine (www.humanmine.org) is an integrated database of human genomics and proteomics data that provides a powerful interface to support sophisticated exploration and analysis of data compiled from experimental, computational and curated data sources. Built using the InterMine data integration platform, HumanMine includes genes, proteins, pathways, expression levels, Single nucleotide polymorphism (SNP), diseases and more, integrated into a single searchable database. HumanMine promotes integrative analysis, a powerful approach in modern biology that allows many sources of evidence to be analysed together. The data can be accessed through a user-friendly web interface as well as a powerful, scriptable web service Application programming interface (API) to allow programmatic access to data. The web interface includes a useful identifier resolution system, sophisticated query options and interactive results tables that enable powerful exploration of data, including data summaries, filtering, browsing and export. A set of graphical analysis tools provide a rich environment for data exploration including statistical enrichment of sets of genes or other biological entities. HumanMine can be used for integrative multistaged analysis that can lead to new insights and uncover previously unknown relationships. Database URL: https://www.humanmine.org.

The impact of quantitative optimization of hybridization conditions on gene expression analysis.

BACKGROUND: With the growing availability of entire genome sequences, an increasing number of scientists can exploit oligonucleotide microarrays for genome-scale expression studies. While probe-design is a major research area, relatively little work has been reported on the optimization of microarray protocols. RESULTS: As shown in this study, suboptimal conditions can have considerable impact on biologically relevant observations. For example, deviation from the optimal temperature by one degree Celsius lead to a loss of up to 44% of differentially expressed genes identified. While genes from thousands of Gene Ontology categories were affected, transcription factors and other low-copy-number regulators were disproportionately lost. Calibrated protocols are thus required in order to take full advantage of the large dynamic range of microarrays.For an objective optimization of protocols we introduce an approach that maximizes the amount of information obtained per experiment. A comparison of two typical samples is sufficient for this calibration. We can ensure, however, that optimization results are independent of the samples and the specific measures used for calibration. Both simulations and spike-in experiments confirmed an unbiased determination of generally optimal experimental conditions. CONCLUSIONS: Well calibrated hybridization conditions are thus easily achieved and necessary for the efficient detection of differential expression. They are essential for the sensitive pro filing of low-copy-number molecules. This is particularly critical for studies of transcription factor expression, or the inference and study of regulatory networks.

Prospero acts as a binary switch between self-renewal and differentiation in Drosophila neural stem cells.

Stem cells have the remarkable ability to give rise to both self-renewing and differentiating daughter cells. Drosophila neural stem cells segregate cell-fate determinants from the self-renewing cell to the differentiating daughter at each division. Here, we show that one such determinant, the homeodomain transcription factor Prospero, regulates the choice between stem cell self-renewal and differentiation. We have identified the in vivo targets of Prospero throughout the entire genome. We show that Prospero represses genes required for self-renewal, such as stem cell fate genes and cell-cycle genes. Surprisingly, Prospero is also required to activate genes for terminal differentiation. We further show that in the absence of Prospero, differentiating daughters revert to a stem cell-like fate: they express markers of self-renewal, exhibit increased proliferation, and fail to differentiate. These results define a blueprint for the transition from stem cell self-renewal to terminal differentiation.