EuPathDB: the eukaryotic pathogen genomics database resource.

The Eukaryotic Pathogen Genomics Database Resource (EuPathDB, http://eupathdb.org) is a collection of databases covering 170+ eukaryotic pathogens (protists & fungi), along with relevant free-living and non-pathogenic species, and select pathogen hosts. To facilitate the discovery of meaningful biological relationships, the databases couple preconfigured searches with visualization and analysis tools for comprehensive data mining via intuitive graphical interfaces and APIs. All data are analyzed with the same workflows, including creation of gene orthology profiles, so data are easily compared across data sets, data types and organisms. EuPathDB is updated with numerous new analysis tools, features, data sets and data types. New tools include GO, metabolic pathway and word enrichment analyses plus an online workspace for analysis of personal, non-public, large-scale data. Expanded data content is mostly genomic and functional genomic data while new data types include protein microarray, metabolic pathways, compounds, quantitative proteomics, copy number variation, and polysomal transcriptomics. New features include consistent categorization of searches, data sets and genome browser tracks; redesigned gene pages; effective integration of alternative transcripts; and a EuPathDB Galaxy instance for private analyses of a user's data. Forthcoming upgrades include user workspaces for private integration of data with existing EuPathDB data and improved integration and presentation of host-pathogen interactions.

Local admixture of amplified and diversified secreted pathogenesis determinants shapes mosaic Toxoplasma gondii genomes.

Toxoplasma gondii is among the most prevalent parasites worldwide, infecting many wild and domestic animals and causing zoonotic infections in humans. T. gondii differs substantially in its broad distribution from closely related parasites that typically have narrow, specialized host ranges. To elucidate the genetic basis for these differences, we compared the genomes of 62 globally distributed T. gondii isolates to several closely related coccidian parasites. Our findings reveal that tandem amplification and diversification of secretory pathogenesis determinants is the primary feature that distinguishes the closely related genomes of these biologically diverse parasites. We further show that the unusual population structure of T. gondii is characterized by clade-specific inheritance of large conserved haploblocks that are significantly enriched in tandemly clustered secretory pathogenesis determinants. The shared inheritance of these conserved haploblocks, which show a different ancestry than the genome as a whole, may thus influence transmission, host range and pathogenicity.

EuPathDB: the eukaryotic pathogen database.

EuPathDB (http://eupathdb.org) resources include 11 databases supporting eukaryotic pathogen genomic and functional genomic data, isolate data and phylogenomics. EuPathDB resources are built using the same infrastructure and provide a sophisticated search strategy system enabling complex interrogations of underlying data. Recent advances in EuPathDB resources include the design and implementation of a new data loading workflow, a new database supporting Piroplasmida (i.e. Babesia and Theileria), the addition of large amounts of new data and data types and the incorporation of new analysis tools. New data include genome sequences and annotation, strand-specific RNA-seq data, splice junction predictions (based on RNA-seq), phosphoproteomic data, high-throughput phenotyping data, single nucleotide polymorphism data based on high-throughput sequencing (HTS) and expression quantitative trait loci data. New analysis tools enable users to search for DNA motifs and define genes based on their genomic colocation, view results from searches graphically (i.e. genes mapped to chromosomes or isolates displayed on a map) and analyze data from columns in result tables (word cloud and histogram summaries of column content). The manuscript herein describes updates to EuPathDB since the previous report published in NAR in 2010.

FungiDB: an integrated functional genomics database for fungi.

FungiDB (http://FungiDB.org) is a functional genomic resource for pan-fungal genomes that was developed in partnership with the Eukaryotic Pathogen Bioinformatic resource center (http://EuPathDB.org). FungiDB uses the same infrastructure and user interface as EuPathDB, which allows for sophisticated and integrated searches to be performed using an intuitive graphical system. The current release of FungiDB contains genome sequence and annotation from 18 species spanning several fungal classes, including the Ascomycota classes, Eurotiomycetes, Sordariomycetes, Saccharomycetes and the Basidiomycota orders, Pucciniomycetes and Tremellomycetes, and the basal 'Zygomycete' lineage Mucormycotina. Additionally, FungiDB contains cell cycle microarray data, hyphal growth RNA-sequence data and yeast two hybrid interaction data. The underlying genomic sequence and annotation combined with functional data, additional data from the FungiDB standard analysis pipeline and the ability to leverage orthology provides a powerful resource for in silico experimentation.

AmoebaDB and MicrosporidiaDB: functional genomic resources for Amoebozoa and Microsporidia species.

AmoebaDB (http://AmoebaDB.org) and MicrosporidiaDB (http://MicrosporidiaDB.org) are new functional genomic databases serving the amoebozoa and microsporidia research communities, respectively. AmoebaDB contains the genomes of three Entamoeba species (E. dispar, E. invadens and E. histolityca) and microarray expression data for E. histolytica. MicrosporidiaDB contains the genomes of Encephalitozoon cuniculi, E. intestinalis and E. bieneusi. The databases belong to the National Institute of Allergy and Infectious Diseases (NIAID) funded EuPathDB (http://EuPathDB.org) Bioinformatics Resource Center family of integrated databases and assume the same architectural and graphical design as other EuPathDB resources such as PlasmoDB and TriTrypDB. Importantly they utilize the graphical strategy builder that affords a database user the ability to ask complex multi-data-type questions with relative ease and versatility. Genomic scale data can be queried based on BLAST searches, annotation keywords and gene ID searches, GO terms, sequence motifs, protein characteristics, phylogenetic relationships and functional data such as transcript (microarray and EST evidence) and protein expression data. Search strategies can be saved within a user's profile for future retrieval and may also be shared with other researchers using a unique strategy web address.

EuPathDB: a portal to eukaryotic pathogen databases.

EuPathDB (http://EuPathDB.org; formerly ApiDB) is an integrated database covering the eukaryotic pathogens of the genera Cryptosporidium, Giardia, Leishmania, Neospora, Plasmodium, Toxoplasma, Trichomonas and Trypanosoma. While each of these groups is supported by a taxon-specific database built upon the same infrastructure, the EuPathDB portal offers an entry point to all these resources, and the opportunity to leverage orthology for searches across genera. The most recent release of EuPathDB includes updates and changes affecting data content, infrastructure and the user interface, improving data access and enhancing the user experience. EuPathDB currently supports more than 80 searches and the recently-implemented 'search strategy' system enables users to construct complex multi-step searches via a graphical interface. Search results are dynamically displayed as the strategy is constructed or modified, and can be downloaded, saved, revised, or shared with other database users.

TriTrypDB: a functional genomic resource for the Trypanosomatidae.

TriTrypDB (http://tritrypdb.org) is an integrated database providing access to genome-scale datasets for kinetoplastid parasites, and supporting a variety of complex queries driven by research and development needs. TriTrypDB is a collaborative project, utilizing the GUS/WDK computational infrastructure developed by the Eukaryotic Pathogen Bioinformatics Resource Center (EuPathDB.org) to integrate genome annotation and analyses from GeneDB and elsewhere with a wide variety of functional genomics datasets made available by members of the global research community, often pre-publication. Currently, TriTrypDB integrates datasets from Leishmania braziliensis, L. infantum, L. major, L. tarentolae, Trypanosoma brucei and T. cruzi. Users may examine individual genes or chromosomal spans in their genomic context, including syntenic alignments with other kinetoplastid organisms. Data within TriTrypDB can be interrogated utilizing a sophisticated search strategy system that enables a user to construct complex queries combining multiple data types. All search strategies are stored, allowing future access and integrated searches. 'User Comments' may be added to any gene page, enhancing available annotation; such comments become immediately searchable via the text search, and are forwarded to curators for incorporation into the reference annotation when appropriate.

GiardiaDB and TrichDB: integrated genomic resources for the eukaryotic protist pathogens Giardia lamblia and Trichomonas vaginalis.

GiardiaDB (http://GiardiaDB.org) and TrichDB (http://TrichDB.org) house the genome databases for Giardia lamblia and Trichomonas vaginalis, respectively, and represent the latest additions to the EuPathDB (http://EuPathDB.org) family of functional genomic databases. GiardiaDB and TrichDB employ the same framework as other EuPathDB sites (CryptoDB, PlasmoDB and ToxoDB), supporting fully integrated and searchable databases. Genomic-scale data available via these resources may be queried based on BLAST searches, annotation keywords and gene ID searches, GO terms, sequence motifs and other protein characteristics. Functional queries may also be formulated, based on transcript and protein expression data from a variety of platforms. Phylogenetic relationships may also be interrogated. The ability to combine the results from independent queries, and to store queries and query results for future use facilitates complex, genome-wide mining of functional genomic data.

PlasmoDB: a functional genomic database for malaria parasites.

PlasmoDB (http://PlasmoDB.org) is a functional genomic database for Plasmodium spp. that provides a resource for data analysis and visualization in a gene-by-gene or genome-wide scale. PlasmoDB belongs to a family of genomic resources that are housed under the EuPathDB (http://EuPathDB.org) Bioinformatics Resource Center (BRC) umbrella. The latest release, PlasmoDB 5.5, contains numerous new data types from several broad categories--annotated genomes, evidence of transcription, proteomics evidence, protein function evidence, population biology and evolution. Data in PlasmoDB can be queried by selecting the data of interest from a query grid or drop down menus. Various results can then be combined with each other on the query history page. Search results can be downloaded with associated functional data and registered users can store their query history for future retrieval or analysis.

ToxoDB: an integrated Toxoplasma gondii database resource.

ToxoDB (http://ToxoDB.org) is a genome and functional genomic database for the protozoan parasite Toxoplasma gondii. It incorporates the sequence and annotation of the T. gondii ME49 strain, as well as genome sequences for the GT1, VEG and RH (Chr Ia, Chr Ib) strains. Sequence information is integrated with various other genomic-scale data, including community annotation, ESTs, gene expression and proteomics data. ToxoDB has matured significantly since its initial release. Here we outline the numerous updates with respect to the data and increased functionality available on the website.

EPConDB: a web resource for gene expression related to pancreatic development, beta-cell function and diabetes.

EPConDB (http://www.cbil.upenn.edu/EPConDB) is a public web site that supports research in diabetes, pancreatic development and beta-cell function by providing information about genes expressed in cells of the pancreas. EPConDB displays expression profiles for individual genes and information about transcripts, promoter elements and transcription factor binding sites. Gene expression results are obtained from studies examining tissue expression, pancreatic development and growth, differentiation of insulin-producing cells, islet or beta-cell injury, and genetic models of impaired beta-cell function. The expression datasets are derived using different microarray platforms, including the BCBC PancChips and Affymetrix gene expression arrays. Other datasets include semi-quantitative RT-PCR and MPSS expression studies. For selected microarray studies, lists of differentially expressed genes, derived from PaGE analysis, are displayed on the site. EPConDB provides database queries and tools to examine the relationship between a gene, its transcriptional regulation, protein function and expression in pancreatic tissues.

Novel genes identified by manual annotation and microarray expression analysis in the pancreas.

The mouse PancChip, a microarray developed for studying endocrine pancreatic development and diabetes, represents over 13,000 cDNAs. After computationally assigning the cDNAs on the array to known genes, manual curation of the remaining sequences identified 211 novel transcripts. In microarray experiments, we found that 196 of these transcripts were expressed in total pancreas and/or pancreatic islets. Of 50 randomly selected clones from these 196 transcripts, 92% were confirmed as expressed by qRT-PCR. We evaluated the coding potential of the novel transcripts and found that 74% of the clones had low coding potential. Since the transcripts may be partial mRNAs, we examined their translated proteins for transmembrane or signal peptide domains and found that about 40 proteins had one of these predicted domains. Interestingly, when we investigated the novel transcripts for their overlap with noncoding microRNAs, we found that 1 of the novel transcripts overlapped a known microRNA gene.

ApiEST-DB: analyzing clustered EST data of the apicomplexan parasites.

ApiEST-DB (http://www.cbil.upenn.edu/paradbs-servlet/) provides integrated access to publicly available EST data from protozoan parasites in the phylum Apicomplexa. The database currently incorporates a total of nearly 100,000 ESTs from several parasite species of clinical and/or veterinary interest, including Eimeria tenella, Neospora caninum, Plasmodium falciparum, Sarcocystis neurona and Toxoplasma gondii. To facilitate analysis of these data, EST sequences were clustered and assembled to form consensus sequences for each organism, and these assemblies were then subjected to automated annotation via similarity searches against protein and domain databases. The underlying relational database infrastructure, Genomics Unified Schema (GUS), enables complex biologically based queries, facilitating validation of gene models, identification of alternative splicing, detection of single nucleotide polymorphisms, identification of stage-specific genes and recognition of phylogenetically conserved and phylogenetically restricted sequences.

Transcriptional program of the endocrine pancreas in mice and humans.

The Endocrine Pancreas Consortium was formed in late 1999 to derive and sequence cDNA libraries enriched for rare transcripts expressed in the mammalian endocrine pancreas. Over the past 3 years, the Consortium has generated 20 cDNA libraries from mouse and human pancreatic tissues and deposited >150,000 sequences into the public expressed sequence tag databases. A special effort was made to enrich for cDNAs from the endocrine pancreas by constructing libraries from isolated islets. In addition, we constructed a library in which fetal pancreas from Neurogenin 3 null mice, which consists of only exocrine and duct cells, was subtracted from fetal wild-type pancreas to enrich for the transcripts from the endocrine compartment. Sequence analysis showed that these clones cluster into 9,464 assembly groups (approximating unique transcripts) for the mouse and 13,910 for the human sequences. Of these, >4,300 were unique to Consortium libraries. We have assembled a core clone set containing one cDNA for each assembly group for the mouse and have constructed the corresponding microarray, termed "PancChip 4.0," which contains >9,000 nonredundant elements. We show that this PancChip is highly enriched for genes expressed in the endocrine pancreas. The mouse and human clone sets and corresponding arrays will be important resources for diabetes research.

Selective repression of myoD transcription by v-Myc prevents terminal differentiation of quail embryo myoblasts transformed by the MC29 strain of avian myelocytomatosis virus.

We have investigated the mechanism by which expression of the v-myc oncogene interferes with the competence of primary quail myoblasts to undergo terminal differentiation. Previous studies have established that quail myoblasts transformed by myc oncogenes are severely impaired in the accumulation of mRNAs encoding the myogenic transcription factors Myf-5, MyoD and Myogenin. However, the mechanism responsible for such a repression remains largely unknown. Here we present evidence that v-Myc selectively interferes with quail myoD expression at the transcriptional level. Cis-regulatory elements involved in the auto-activation of qmyoD are specifically targeted in this unique example of transrepression by v-Myc, without the apparent participation of Myc-specific E-boxes or InR sequences. Transiently expressed v-Myc efficiently interfered with MyoD-dependent transactivation of the qmyoD regulatory elements, while the myogenin promoter was unaffected. Finally, we show that forced expression of MyoD in v-myc-transformed quail myoblasts restored myogenin expression and promoted extensive terminal differentiation. These data suggest that transcriptional repression of qmyoD is a major and rate-limiting step in the molecular pathway by which v-Myc severely inhibits terminal differentiation in myogenic cells.

Functional genomics of the endocrine pancreas: the pancreas clone set and PancChip, new resources for diabetes research.

Over the past 5 years, microarrays have greatly facilitated large-scale analysis of gene expression levels. Although these arrays were not specifically geared to represent tissues and pathways known to be affected by diabetes, they have been used in both type 1 and type 2 diabetes research. To prepare a tool that is particularly useful in the study of type 1 diabetes, we have assembled a nonredundant set of 3,400 clones representing genes expressed in the mouse pancreas or pathways known to be affected by diabetes. We have demonstrated the usefulness of this clone set by preparing a cDNA glass microarray, the PancChip, and using it to analyze pancreatic gene expression from embryonic day 14.5 through adulthood in mice. The clone set and corresponding array are useful resources for diabetes research.

Myf5 is a direct target of long-range Shh signaling and Gli regulation for muscle specification.

Sonic hedgehog (Shh) is a secreted signaling molecule for tissue patterning and stem cell specification in vertebrate embryos. Shh mediates both long-range and short-range signaling responses in embryonic tissues through the activation and repression of target genes by its Gli transcription factor effectors. Despite the well-established functions of Shh signaling in development and human disease, developmental target genes of Gli regulation are virtually unknown. In this study, we investigate the role of Shh signaling in the control of Myf5, a skeletal muscle regulatory gene for specification of muscle stem cells in vertebrate embryos. In previous genetic studies, we showed that Shh is required for Myf5 expression in the specification of dorsal somite, epaxial muscle progenitors. However, these studies did not distinguish whether Myf5 is a direct target of Gli regulation through long-range Shh signaling, or alternatively, whether Myf5 regulation is a secondary response to Shh signaling. To address this question, we have used transgenic analysis with lacZ reporter genes to characterize an Myf5 transcription enhancer that controls the activation of Myf5 expression in the somite epaxial muscle progenitors in mouse embryos. This Myf5 epaxial somite (ES) enhancer is Shh-dependent, as shown by its complete inactivity in somites of homozygous Shh mutant embryos, and by its reduced activity in heterozygous Shh mutant embryos. Furthermore, Shh and downstream Shh signal transducers specifically induce ES enhancer/luciferase reporters in Shh-responsive 3T3 cells. A Gli-binding site located within the ES enhancer is required for enhancer activation by Shh signaling in transfected 3T3 cells and in epaxial somite progenitors in transgenic embryos. These findings establish that Myf5 is a direct target of long-range Shh signaling through positive regulation by Gli transcription factors, providing evidence that Shh signaling has a direct inductive function in cell lineage specification.