Dataset Information

Shotgun ecotoxicoproteomics of Daphnia pulex: biochemical effects of the anticancer drug tamoxifen

ABSTRACT: Among pollutants released into the environment by human activities, residues of pharmaceutical agents have been identified worldwide, and are an increasing matter of concern because of their potential impact on ecosystems. Because micro-crustaceans, such as daphnids, are sensitive to chemicals and are key organisms in the food chain, they are particularly convenient for studying the effects of these drugs in the aquatic environment. The aim of this study was to analyse differences of protein expression resulting from acute (2 days) and middle-term (7 days) exposure of Daphnia pulex daphnids to the anticancer drug tamoxifen, with the hope to unveil links between the effects of xenobiotics at the organism level and changes at the molecular level. Using a liquid chromatography - mass spectrometry shotgun approach, about 4000 proteins could be identified with a minimum of one unique peptide, providing the largest proteomics dataset of D. pulex published up to now. Considering both time points and tested concentrations, 189 proteins showed a significant fold change, most of the regulated proteins being observed at the highest concentration tested and for the longer exposure time. About one third of significant proteins were positively regulated, whereas two thirds showed decreased levels after tamoxifen treatment. The identity of regulated proteins suggested a decrease in translation, an increase in protein degradation and changes in carbohydrate and lipid metabolism as the major effects of the drug. Besides these impacted processes, which reflect a general stress response of the organism, some other regulated proteins play a role in Daphnia reproduction. These latter results are in accordance with our previous observations of the impact of tamoxifen on D. pulex reproduction, and illustrate the potential of ecotoxicoproteomics to unravel links between xenobiotic effects at the biochemical and at the organismal level.

INSTRUMENT(S): LTQ Orbitrap

ORGANISM(S): Daphnia Pulex (water Flea)

TISSUE(S): Whole Body

SUBMITTER: Patrice Waridel

LAB HEAD: Manfredo Quadroni

PROVIDER: PXD001257 | Pride | 2014-10-31

REPOSITORIES: Pride

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Shotgun ecotoxicoproteomics of Daphnia pulex: biochemical effects of the anticancer drug tamoxifen.

Borgatta Myriam M Hernandez Céline C Decosterd Laurent Arthur LA Chèvre Nathalie N Waridel Patrice P

Journal of proteome research 20141113 1

Among pollutants released into the environment by human activities, residues of pharmaceuticals are an increasing matter of concern because of their potential impact on ecosystems. The aim of this study was to analyze differences of protein expression resulting from acute (2 days) and middle-term (7 days) exposure of aquatic microcrustacean Daphnia pulex to the anticancer drug tamoxifen. Using a liquid chromatography-mass spectrometry shotgun approach, about 4000 proteins could be identified, pr ...[more]

PMID: 25350372

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Project description:The US Department of Defense (DOD) is developing insensitive munitions (IMs) that are resistant to unintended detonation to protect warfighters. To enable evaluation of the material life-cycle for the IM, 1-methyl-3-nitro-1-nitroguanidine (MeNQ), ecotoxicological impacts assessment was required. A previous investigation of MeNQ exposures in Daphnia pulex revealed concentration-responsive decreases in reproduction relative to controls (0 mg/L) across a 174, 346, 709, 1385, and 2286 mg/L exposure range. The present study used those exposures to conduct global transcriptomic expression analyses to establish hypothetical mechanisms underlying inhibited reproduction. Changes in transcript expression tended to increase with increasing MeNQ concentration where hierarchical clustering analysis identified separation among the “low” (174 and 346 mg/L) and “high” (709, 1385, and 2286 mg/L) exposures. The MeNQ exposures significantly decreased transcriptional expression for a vitellogenin precursor and juvenile hormone-inducible protein, each having plausible mechanistic connections to impaired reproductive physiology and reduced offspring production in Daphnia. Further, MeNQ affected chorion peroxidase and high choriolytic enzyme transcriptional expression which are known to influence egg production / maturation processes in arthropods. Additionally, MeNQ exposure caused dose-responsive decreases in transcriptional expression for a diverse set of cuticle proteins which requires closer examination regarding Daphnia embryo development and brood release. Finally, canonical-pathway enrichment analysis identified significant effects on a broad array of energy metabolic processes in D. pulex which require consideration regarding potential shifts in homeostatic equilibrium that can have indirect costs on reproduction. Overall, plausible mechanisms of inhibited D. pulex reproduction have been identified and should be tested in targeted bioassays. see previous row.

Project description: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.

Project description:"Gene response profiles for Daphnia pulex exposed to the environmental stressor cadmium reveals novel crustacean metallothioneins." Joseph.R.Shaw, John.K.Colbourne, Jennifer.C.Davey, Stephen.P.Glaholt, Thomas.H.Hampton, Celia.Y.Chen, Carol.L.Folt, Joshua.W.Hamilton (BMC Genomics 2007). Genomic research tools such as microarrays are proving to be important resources to study the complex regulation of genes that respond to environmental perturbations. A first generation cDNA microarray was developed for the environmental indicator species Daphnia pulex, to identify genes whose regulation is modulated following exposure to the metal stressor cadmium. Our experiments revealed interesting changes in gene transcription that suggest their biological roles and their potentially toxicological features in responding to this important environmental contaminant. Our microarray identified genes reported in the literature to be regulated in response to cadmium exposure, suggested functional attributes for genes that share no sequence similarity to proteins in the public databases, and pointed to genes that are likely members of expanded gene families in the Daphnia genome. Genes identified on the microarray also were associated with cadmium induced phenotypes and population-level outcomes that we experimentally determined. A subset of genes regulated in response to cadmium exposure was independently validated using quantitative-realtime (Q-RT)-PCR. These microarray studies led to the discovery of three genes coding for the metal detoxication protein metallothionein (MT). The gene structures and predicted translated sequences of D. pulex MTs clearly place them in this gene family. Yet, they share little homology with previously characterized MTs. The genomic information obtained from this study represents an important first step in characterizing microarray patterns that may be diagnostic to specific environmental contaminants and give insights into their toxicological mechanisms, while also providing a practical tool for evolutionary, ecological, and toxicological functional gene discovery studies. Advances in Daphnia genomics will enable the further development of this species as a model organism for the environmental sciences. Keywords: cDNA microarrays, stress response, expressed sequence tags (EST), Daphnia Genomics Consortium, Arthropod, Gene Expression Profiling, Gene Expression Regulation

Dataset Information

Shotgun ecotoxicoproteomics of Daphnia pulex: biochemical effects of the anticancer drug tamoxifen

Publications

Shotgun ecotoxicoproteomics of Daphnia pulex: biochemical effects of the anticancer drug tamoxifen.

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