wFleaBase: the Daphnia genome database

BACKGROUND: wFleaBase is a database with the necessary infrastructure to curate, archive and share genetic, molecular and functional genomic data and protocols for an emerging model organism, the microcrustacean Daphnia. Commonly known as the water-flea, Daphnia's ecological merit is unequaled among metazoans, largely because of its sentinel role within freshwater ecosystems and over 200 years of biological investigations. By consequence, the Daphnia Genomics Consortium (DGC) has launched an interdisciplinary research program to create the resources needed to study genes that affect ecological and evolutionary success in natural environments. DISCUSSION: These tools include the genome database wFleaBase, which currently contains functions to search and extract information from expressed sequenced tags, genome survey sequences and full genome sequencing projects. This new database is built primarily from core components of the Generic Model Organism Database project, and related bioinformatics tools. SUMMARY: Over the coming year, preliminary genetic maps and the nearly complete genomic sequence of Daphnia pulex will be integrated into wFleaBase, including gene predictions and ortholog assignments based on sequence similarities with eukaryote genes of known function. wFleaBase aims to serve a large ecological and evolutionary research community. Our challenge is to rapidly expand its content and to ultimately integrate genetic and functional genomic information with population-level responses to environmental challenges. URL:

Centennial clonal stability of asexual Daphnia in Greenland lakes despite climate variability

Climate and environmental condition drive biodiversity at many levels of biological organization, from populations to ecosystems. Combined with paleoecological reconstructions, palaeogenetic information on resident populations provides novel insights into evolutionary trajectories and genetic diversity driven by environmental variability. While temporal observations of changing genetic structure are often made of sexual populations, little is known about how environmental change affects the long‐term fate of asexual lineages. Here, we provide information on obligately asexual, triploid Daphnia populations from three Arctic lakes in West Greenland through the past 200–300 years to test the impact of environmental change on the temporal and spatial population genetic structure. The contrasting ecological state of the lakes, specifically regarding salinity and habitat structure may explain the observed lake‐specific clonal composition over time. Palaeolimnological reconstructions show considerable regional environmental fluctuations since 1,700 (the end of the Little Ice Age), but the population genetic structure in two lakes was almost unchanged with at most two clones per time period. Their local populations were strongly dominated by a single clone that has persisted for 250–300 years. We discuss possible explanations for the apparent population genetic stability: (a) persistent clones are general‐purpose genotypes that thrive under broad environmental conditions, (b) clonal lineages evolved subtle genotypic differences unresolved by microsatellite markers, or (c) epigenetic modifications allow for clonal adaptation to changing environmental conditions. Our results motivate research into the mechanisms of adaptation in these populations, as well as their evolutionary fate in the light of accelerating climate change in the polar regions

Profiling sex-biased gene expression during parthenogenetic reproduction in Daphnia pulex

<p>Abstract</p> <p>Background</p> <p>Sexual reproduction is a core biological function that is conserved throughout eukaryotic evolution, yet breeding systems are extremely variable. Genome-wide comparative studies can be effectively used to identify genes and regulatory patterns that are constrained to preserve core functions from those that may help to account for the diversity of animal reproductive strategies. We use a custom microarray to investigate gene expression in males and two reproductive stages of females in the crustacean <it>Daphnia pulex</it>. Most <it>Daphnia </it>species reproduce by cyclical parthenogenesis, alternating between sexual and clonal reproduction. Both sex determination and the switch in their mode of reproduction is environmentally induced, making <it>Daphnia </it>an interesting comparative system for the study of sex-biased and reproductive genes.</p> <p>Results</p> <p>Patterns of gene expression in females and males reveal that 50% of assayed transcripts show some degree of sex-bias. Female-biased transcription is enriched for translation, metabolic and regulatory genes associated with development. Male-biased expression is enriched for cuticle and protease function. Comparison with well studied arthropods such as <it>Drosophila melanogaster </it>and <it>Anopheles gambiae </it>suggests that female-biased patterns tend to be conserved, whereas male-biased genes are evolving faster in <it>D. pulex</it>. These findings are based on the proportion of female-biased, male-biased, and unbiased genes that share sequence similarity with proteins in other animal genomes.</p> <p>Conclusion</p> <p>Some transcriptional differences between males and females appear to be conserved across Arthropoda, including the rapid evolution of male-biased genes which is observed in insects and now in a crustacean. Yet, novel patterns of male-biased gene expression are also uncovered. This study is an important first step towards a detailed understanding of the genetic basis and evolution of parthenogenesis, environmental sex determination, and adaptation to aquatic environments.</p

Genomic data integration for ecological and evolutionary traits in non-model organisms

Why is it needed to develop system biology initiatives such as ENCODE on non-model organisms

Sampling Daphnia's expressed genes: preservation, expansion and invention of crustacean genes with reference to insect genomes

<p>Abstract</p> <p>Background</p> <p>Functional and comparative studies of insect genomes have shed light on the complement of genes, which in part, account for shared morphologies, developmental programs and life-histories. Contrasting the gene inventories of insects to those of the nematodes provides insight into the genomic changes responsible for their diversification. However, nematodes have weak relationships to insects, as each belongs to separate animal phyla. A better outgroup to distinguish lineage specific novelties would include other members of Arthropoda. For example, crustaceans are close allies to the insects (together forming Pancrustacea) and their fascinating aquatic lifestyle provides an important comparison for understanding the genetic basis of adaptations to life on land versus life in water.</p> <p>Results</p> <p>This study reports on the first characterization of cDNA libraries and sequences for the model crustacean <it>Daphnia pulex</it>. We analyzed 1,546 ESTs of which 1,414 represent approximately 787 nuclear genes, by measuring their sequence similarities with insect and nematode proteomes. The provisional annotation of genes is supported by expression data from microarray studies described in companion papers. Loci expected to be shared between crustaceans and insects because of their mutual biological features are identified, including genes for reproduction, regulation and cellular processes. We identify genes that are likely derived within Pancrustacea or lost within the nematodes. Moreover, lineage specific gene family expansions are identified, which suggest certain biological demands associated with their ecological setting. In particular, up to seven distinct ferritin loci are found in <it>Daphnia </it>compared to three in most insects. Finally, a substantial fraction of the sampled gene transcripts shares no sequence similarity with those from other arthropods. Genes functioning during development and reproduction are comparatively well conserved between crustaceans and insects. By contrast, genes that were responsive to environmental conditions (metal stress) and not sex-biased included the greatest proportion of genes with no matches to insect proteomes.</p> <p>Conclusion</p> <p>This study along with associated microarray experiments are the initial steps in a coordinated effort by the <it>Daphnia </it>Genomics Consortium to build the necessary genomic platform needed to discover genes that account for the phenotypic diversity within the genus and to gain new insights into crustacean biology. This effort will soon include the first crustacean genome sequence.</p

A comprehensive epigenomic analysis of phenotypically distinguishable, genetically identical female and male Daphnia pulex

Background Daphnia species reproduce by cyclic parthenogenesis involving both sexual and asexual reproduction. The sex of the offspring is environmentally determined and mediated via endocrine signalling by the mother. Interestingly, male and female Daphnia can be genetically identical, yet display large differences in behaviour, morphology, lifespan and metabolic activity. Our goal was to integrate multiple omics datasets, including gene expression, splicing, histone modification and DNA methylation data generated from genetically identical female and male Daphnia pulex under controlled laboratory settings with the aim of achieving a better understanding of the underlying epigenetic factors that may contribute to the phenotypic differences observed between the two genders. Results In this study we demonstrate that gene expression level is positively correlated with increased DNA methylation, and histone H3 trimethylation at lysine 4 (H3K4me3) at predicted promoter regions. Conversely, elevated histone H3 trimethylation at lysine 27 (H3K27me3), distributed across the entire transcript length, is negatively correlated with gene expression level. Interestingly, male Daphnia are dominated with epigenetic modifications that globally promote elevated gene expression, while female Daphnia are dominated with epigenetic modifications that reduce gene expression globally. For examples, CpG methylation (positively correlated with gene expression level) is significantly higher in almost all differentially methylated sites in male compared to female Daphnia. Furthermore, H3K4me3 modifications are higher in male compared to female Daphnia in more than 3/4 of the differentially regulated promoters. On the other hand, H3K27me3 is higher in female compared to male Daphnia in more than 5/6 of differentially modified sites. However, both sexes demonstrate roughly equal number of genes that are up-regulated in one gender compared to the other sex. Since, gene expression analyses typically assume that most genes are expressed at equal level among samples and different conditions, and thus cannot detect global changes affecting most genes. Conclusions The epigenetic differences between male and female in Daphnia pulex are vast and dominated by changes that promote elevated gene expression in male Daphnia. Furthermore, the differences observed in both gene expression changes and epigenetic modifications between the genders relate to pathways that are physiologically relevant to the observed phenotypic differences.Peer reviewe

Predator-induced defences in Daphnia pulex: Selection and evaluation of internal reference genes for gene expression studies with real-time PCR

<p>Abstract</p> <p>Background</p> <p>The planktonic microcrustacean <it>Daphnia pulex </it>is among the best-studied animals in ecological, toxicological and evolutionary research. One aspect that has sustained interest in the study system is the ability of <it>D. pulex </it>to develop inducible defence structures when exposed to predators, such as the phantom midge larvae <it>Chaoborus</it>. The available draft genome sequence for <it>D. pulex </it>is accelerating research to identify genes that confer plastic phenotypes that are regularly cued by environmental stimuli. Yet for quantifying gene expression levels, no experimentally validated set of internal control genes exists for the accurate normalization of qRT-PCR data.</p> <p>Results</p> <p>In this study, we tested six candidate reference genes for normalizing transcription levels of <it>D. pulex </it>genes; alpha tubulin (aTub), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), TATA box binding protein (Tbp) syntaxin 16 (Stx16), X-box binding protein 1 (Xbp1) and CAPON, a protein associated with the neuronal nitric oxide synthase, were selected on the basis of an earlier study and from microarray studies. One additional gene, a matrix metalloproteinase (MMP), was tested to validate its transcriptional response to <it>Chaoborus</it>, which was earlier observed in a microarray study. The transcription profiles of these seven genes were assessed by qRT-PCR from RNA of juvenile <it>D. pulex </it>that showed induced defences in comparison to untreated control animals. We tested the individual suitability of genes for expression normalization using the programs geNorm, NormFinder and BestKeeper. Intriguingly, Xbp1, Tbp, CAPON and Stx16 were selected as ideal reference genes. Analyses on the relative expression level using the software REST showed that both classical housekeeping candidate genes (aTub and GAPDH) were significantly downregulated, whereas the MMP gene was shown to be significantly upregulated, as predicted. aTub is a particularly ill suited reference gene because five copies are found in the <it>D. pulex </it>genome sequence. When applying aTub for expression normalization Xbp1 and Tbp are falsely reported as significantly upregulated.</p> <p>Conclusions</p> <p>Our results suggest that the genes Xbp1, Tbp, CAPON and Stx16 are suitable reference genes for accurate normalization in qRT-PCR studies using <it>Chaoborus</it>-induced <it>D. pulex </it>specimens. Furthermore, our study underscores the importance of verifying the expression stability of putative reference genes for normalization of expression levels.</p

Why so many unknown genes?:Partitioning orphans from a representative transcriptome of the lone star tick Amblyomma americanum

BACKGROUND: Genomic resources within the phylum Arthropoda are largely limited to the true insects but are beginning to include unexplored subphyla, such as the Crustacea and Chelicerata. Investigations of these understudied taxa uncover high frequencies of orphan genes, which lack detectable sequence homology to genes in pre-existing databases. The ticks (Acari: Chelicerata) are one such understudied taxon for which genomic resources are urgently needed. Ticks are obligate blood-feeders that vector major diseases of humans, domesticated animals, and wildlife. In analyzing a transcriptome of the lone star tick Amblyomma americanum, one of the most abundant disease vectors in the United States, we find a high representation of unannotated sequences. We apply a general framework for quantifying the origin and true representation of unannotated sequences in a dataset and for evaluating the biological significance of orphan genes. RESULTS: Expressed sequence tags (ESTs) were derived from different life stages and populations of A. americanum and combined with ESTs available from GenBank to produce 14,310 ESTs, over twice the number previously available. The vast majority (71%) has no sequence homology to proteins archived in UniProtKB. We show that poor sequence or assembly quality is not a major contributor to this high representation by orphan genes. Moreover, most unannotated sequences are functional: a microarray experiment demonstrates that 59% of functional ESTs are unannotated. Lastly, we attempt to further annotate our EST dataset using genomic datasets from other members of the Acari, including Ixodes scapularis, four other tick species and the mite Tetranychus urticae. We find low homology with these species, consistent with significant divergence within this subclass. CONCLUSIONS: We conclude that the abundance of orphan genes in A. americanum likely results from 1) taxonomic isolation stemming from divergence within the tick lineage and limited genomic resources for ticks and 2) lineage-specific genes needing functional genomic studies to evaluate their association with the unique biology of ticks. The EST sequences described here will contribute substantially to the development of tick genomics. Moreover, the framework provided for the evaluation of orphan genes can guide analyses of future transcriptome sequencing projects