Changes in lipid profiles induced by bisphenol A (BPA) in zebrafish eleutheroembryos during the yolk sac absorption stage

Bisphenol A (BPA; 4,4’-(propane-2,2-diyl)diphenol) has been shown to act as an obesogen and to disrupt lipid metabolism in zebrafish eleutheroembryos (ZE). To characterize the consequences of this disruption, we performed a detailed lipidomic study using ZE exposed to different BPA concentrations (0, 4, 6 and 8 mg/L of BPA) from day 2 to up to day 6 post fertilization (dpf). Total lipids at 4, 5 and 6 dpf were extracted by Folch method and analyzed by high-performance thin layer chromatography (HPTLC) as wide-range preliminary screening. Selected conditions (0 and 6 mg/L of BPA) were used to obtain a high-quality lipid profile using ultra high-performance liquid chromatography/time-of-flight mass spectrometry (UHPLC-TOFMS). BPA exposed ZE exhibited increased amounts of triglycerides (TG), diglycerides (DG), phosphatidylcholines (PC) and phosphatidylinositols (PI), regarding the control group. Analysis of time- and BPA exposure-related patterns of specific lipid species showed a clear influence of unsaturation degree (mostly in DG and PC) and/or fatty acid chain length (mostly in TG and PC derivatives) on their response to the presence of BPA. A decreased yolk-sac and energy consumption in exposed individuals appeared as the main reason for the observed BPA-driven effects. Integration of these results with previous morphological, biochemical, transcriptomic, metabolomic and behavioral data suggests a disruption of different signalling pathways by BPA that starts at very low BPAconcentrations, whose effects propagate across different organization levels, and that cannot be only explained by the relatively weak estrogenic effect of BPA. © 2019 Elsevier LtdThis work was supported by grants from the Spanish Ministry of Economy and Competitiveness (CTQ2014-56777-R) and the Spanish Ministry of Science, Innovation and University (RTI2018-096175-B-I00). RM was supported by a FPU predoctoral fellow from the Spanish Ministry of Education, Culture and Sport (ref. FPU15/03332). IF was supported by a FPI predoctoral fellow from the Spanish Ministry of Economy and Competitiveness (FPI-MICINN BES-2015-075023). We thank Juan Carlos Navarro and Josefina Casas for his valuable advice and expertise in the optimization of the thin-layer chromatography technique and liquid chromatography/mass spectrometry, respectively.Peer reviewe

[Raw Data] Lipidomic effects of bisphenol A in zebrafish eleutheroembryos during the yolk sac absorption stage

Previous studies on the effects of bisphenol A (BPA; 4,4'-(propane-2,2-diyl)diphenol) in zebrafish identified lipid metabolism as particularly affected by BPA exposure. We perform here a lipidomic analysis in zebrafish eleutheroembryos exposed up to 8 mg/L of BPA from day 2 to day 6 post fertilization. Total lipids were analyzed by high-performance thin layer chromatography as wide-range preliminary screening. Selected conditions were used to obtain a high-quality lipid profile using ultra high-performance liquid chromatography/time-of-flight mass spectrometry. BPA specifically impaired the physiological consumption of triglycerides (TG), diglycerides (DG), phosphatidylcholines (PC) and phosphatidylinositols (PI), most of them yolk sac-related lipids. These effects seemed more related to a decrease in energy consumption in BPA exposed individuals than to an impairment of lipid transport from the yolk sac. Integration of these results with previous morphological, biochemical, transcriptomic, metabolomic and behavioral data suggests that changes on lipid metabolism first appeared at very low BPA concentrations, and that they propagated across different organization levels. The analysis also suggests that the disruption of different signalling pathways by BPA results in a combination of adverse outcomes (estrogenic and lipid disruption, eye development), making it advisable a tighter limitation on BPA exposure limits for both wildlife and human populations.This work was supported by the European Research Council under the European Union's Seventh Framework Programme (FP/ 2007-2013)/ERC Grant Agreement n. 320737. Some part of this study was also supported by grants from the Spanish Ministry of Economy and Competitiveness (CTQ2014-56777-R) and the Spanish Ministry of Science, Innovation and University (RTI2018-096175-B-I00). RM was supported by a FPU predoctoral fellow from the Spanish Ministry of Education and Science (ref. FPU15/03332). IF was supported by a FPI predoctoral fellow from the Spanish Ministry of Economy and Competitiveness (FPI-MICINN BES-2015-075023). We thank Juan Carlos Navarro and Josefina Casas for his valuable advice and expertise in the optimization of the thin-layer chromatography technique and liquid chromatography/ mass spectrometry, respectively.Peer reviewe

DNA methylation in clonal Duckweed lineages ( Lemna minor L.) reflects current and historical environmental exposures

Environmentally induced DNA methylation variants may mediate gene expression responses to environmental changes. If such induced variants are transgenerationally stable, there is potential for expression responses to persist over multiple generations. Our current knowledge in plants, however, is almost exclusively based on studies conducted in sexually reproducing species where the majority of DNA methylation changes are subject to resetting in germlines, limiting the potential for transgenerational epigenetics stress memory. Asexual reproduction circumvents germlines, and may therefore be more conducive to long-term inheritance of epigenetic marks. Taking advantage of the rapid clonal reproduction of the common duckweed Lemna minor, we hypothesize that long-term, transgenerational stress memory from exposure to high temperature can be detected in DNA methylation profiles. Using a reduced representation bisulphite sequencing approach (epiGBS), we show that temperature stress induces DNA hypermethylation at many CG and CHG cytosine contexts but not CHH. Additionally, differential methylation in CHG context that was observed was still detected in a subset of cytosines, even after 3–12 generations of culturing in a common environment. This demonstrates a memory effect of stress reflected in the methylome and that persists over multiple clonal generations. Structural annotation revealed that this memory effect in CHG methylation was enriched in transposable elements. The observed epigenetic stress memory is probably caused by stable transgenerational persistence of temperature-induced DNA methylation variants across clonal generations. To the extent that such epigenetic memory has functional consequences for gene expression and phenotypes, this result suggests potential for long-term modulation of stress responses in asexual plants

Data Analysis : Morphometric signatures of endocrine disrupting chemicals exposures in zebrafish eleutheroembryos [Dataset]

Understanding the mode of action of the different pollutants in human and wildlife health is a key step in environmental risk assessment. The aim of this study was to determine signatures that could link morphological phenotypes to the toxicity mechanisms of four Endocrine Disrupting Chemicals (EDCs): bisphenol A (BPA), perfluorooctanesulfonate potassium salt (PFOS), tributyltin chloride (TBT), and 17-ß-estradiol (E2). Zebrafish (Danio rerio) eleutheroembryos were exposed from 2 to 5 dpf to a wide range of BPA, PFOS, TBT and E2 concentrations. At the end of the exposures several morphometric features were assessed. Common and non-specific effects on larvae pigmentation or swim bladder area were observed after exposures to all compounds. BPA specifically induced yolk sac malabsorption syndrome and altered craniofacial parameters, whereas PFOS had specific effects on the notochord formation presenting higher rates of scoliosis and kyphosis. The main effect of E2 was an increase in the body length of the exposed eleutheroembryos. In the case of TBT, main alterations on the morphological traits were related to developmental delays. When integrating all morphometrical parameters, BPA showed the highest rates of malformations in terms of equilethality, followed by PFOS and, distantly, by TBT and E2. In the case of BPA and PFOS, we were able to relate our results with effects on the transcriptome and metabolome, previously reported. We propose that methodized morphometric analyses in zebrafish embryo model can be used as an inexpensive and easy screening tool to predict modes of action of a wide-range number of contaminants.This work was supported by a grant from the Spanish Ministry of Economy and Competitiveness (ref. CTQ2014-56777-R). LNM was supported by a H2020-Marie Skłodowska-Curie Action MSCA-IF-RI-2017 awarded by the European Commission (ref.797725 EpiSTOX). RM was supported by an FPU predoctoral fellow from the Spanish Ministry of Education and Science (ref. FPU15/03332). We thank Morgane Van Antro and Chiara Luccarelli for helping with eleutheroembryos measurements.Peer reviewe

Morphometric signatures of exposure to endocrine disrupting chemicals in zebrafish eleutheroembryos

Understanding the mode of action of the different pollutants in human and wildlife health is a key step in environmental risk assessment. The aim of this study was to determine signatures that could link morphological phenotypes to the toxicity mechanisms of four Endocrine Disrupting Chemicals (EDCs): bisphenol A (BPA), perfluorooctanesulfonate potassium salt (PFOS), tributyltin chloride (TBT), and 17-ß-estradiol (E2). Zebrafish (Danio rerio) eleutheroembryos were exposed from 2 to 5 dpf to a wide range of BPA, PFOS, TBT and E2 concentrations. At the end of the exposures several morphometric features were assessed. Common and non-specific effects on larvae pigmentation or swim bladder area were observed after exposures to all compounds. BPA specifically induced yolk sac malabsorption syndrome and altered craniofacial parameters, whereas PFOS had specific effects on the notochord formation presenting higher rates of scoliosis and kyphosis. The main effect of E2 was an increase in the body length of the exposed eleutheroembryos. In the case of TBT, main alterations on the morphological traits were related to developmental delays. When integrating all morphometrical parameters, BPA showed the highest rates of malformations in terms of equilethality, followed by PFOS and, distantly, by TBT and E2. In the case of BPA and PFOS, we were able to relate our results with effects on the transcriptome and metabolome, previously reported. We propose that methodized morphometric analyses in zebrafish embryo model can be used as an inexpensive and easy screening tool to predict modes of action of a wide-range number of contaminants.This work was supported by a grant from the Spanish Ministry of Economy and Competitiveness (ref. CTQ2014-56777-R). LNM was supported by a H2020-Marie Skłodowska-Curie Action MSCA-IF-RI-2017 awarded by the European Commission (ref. 797725-EpiSTOX). RM was supported by an FPU predoctoral fellow from the Spanish Ministry of Education and Science (ref. FPU15/03332). We thank Chiara Luccarelli for helping with eleutheroembryos measurements.Peer reviewe

Environmental and genealogical effects on DNA methylation in a widespread apomictic dandelion lineage

International audienceAbstract DNA methylation in plant genomes occurs in different sequences and genomic contexts that have very different properties. DNA methylation that occurs in CG (mCG) sequence context shows transgenerational stability and high epimutation rate, and can thus provide genealogical information at short time scales. However, due to meta‐stability and because mCG variants may arise due to other factors than epimutation, such as environmental stress exposure, it is not clear how well mCG captures genealogical information at micro‐evolutionary time scales. Here, we analysed DNA methylation variation between accessions from a geographically widespread, apomictic common dandelion ( Taraxacum officinale ) lineage when grown experimentally under different light conditions. Using a reduced‐representation bisulphite sequencing approach, we show that the light treatment induced differentially methylated cytosines (DMCs) in all sequence contexts, with a bias towards transposable elements. Accession differences were associated mainly with DMCs in CG context. Hierarchical clustering of samples based on total mCG profiles revealed a perfect clustering of samples by accession identity, irrespective of light conditions. Using microsatellite information as a benchmark of genetic divergence within the clonal lineage, we show that genetic divergence between accessions correlates strongly with overall mCG profiles. However, our results suggest that environmental effects that do occur in CG context may produce a heritable signal that partly dilutes the genealogical signal. Our study shows that methylation information in plants can be used to reconstruct micro‐evolutionary genealogy, providing a useful tool in systems that lack genetic variation such as clonal and vegetatively propagated plants

epiGBS2: Improvements and evaluation of highly demultiplexed, epiGBS-based RRBS

This dataset was create to test the performance capabilities of the epiGBS 2.0 pipeline. We created a epiGBS dataset containing 44 A.thaliana individuals from 6 different accessions