Zebrafish embryos as a screen for DNA methylation modifications after compound exposure

Bouwmeester, Manon C.; Ruiter, Sander; Lommelaars, Tobias; Sippel, Josefine; Hodemaekers, Hennie M.; Brandhof, Evert-Jan van den; Pennings, Jeroen L. A.; Kamstra, Jorke H.; Jelı́nek, Jaroslav; Issa, Jean–Pierre J.; Legler, Juliette; Ven, Leo T.M. van der

doi:10.1016/j.taap.2015.12.012

Cited by 59 publications

(38 citation statements)

References 63 publications

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“…We exposed embryos to 5AC at concentrations that were below the effect concentration in zebrafish embryo toxicity tests based on our own results and others [29, 41, 42]. Nevertheless, developmental exposure of zebrafish embryos to 5AC resulted in transgenerational effects on larval body length as well as direct effects on gastrointestinal development in F0 larvae at 15 dpf.…”

Section: Discussionmentioning

confidence: 99%

Differential DNA methylation at conserved non-genic elements and evidence for transgenerational inheritance following developmental exposure to mono(2-ethylhexyl) phthalate and 5-azacytidine in zebrafish

Kamstra

Sales

Aleström

et al. 2017

Epigenetics & Chromatin

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BackgroundExposure to environmental stressors during development may lead to latent and transgenerational adverse health effects. To understand the role of DNA methylation in these effects, we used zebrafish as a vertebrate model to investigate heritable changes in DNA methylation following chemical-induced stress during early development. We exposed zebrafish embryos to non-embryotoxic concentrations of the biologically active phthalate metabolite mono(2-ethylhexyl) phthalate (MEHP, 30 µM) and the DNA methyltransferase 1 inhibitor 5-azacytidine (5AC, 10 µM). Direct, latent and transgenerational effects on DNA methylation were assessed using global, genome-wide and locus-specific DNA methylation analyses.ResultsFollowing direct exposure in zebrafish embryos from 0 to 6 days post-fertilization, genome-wide analysis revealed a multitude of differentially methylated regions, strongly enriched at conserved non-genic elements for both compounds. Pathways involved in adipogenesis were enriched with the putative obesogenic compound MEHP. Exposure to 5AC resulted in enrichment of pathways involved in embryonic development and transgenerational effects on larval body length. Locus-specific methylation analysis of 10 differentially methylated sites revealed six of these loci differentially methylated in sperm sampled from adult zebrafish exposed during development to 5AC, and in first and second generation larvae. With MEHP, consistent changes were found at 2 specific loci in first and second generation larvae.ConclusionsOur results suggest a functional role for DNA methylation on cis-regulatory conserved elements following developmental exposure to compounds. Effects on these regions are potentially transferred to subsequent generations.Electronic supplementary materialThe online version of this article (doi:10.1186/s13072-017-0126-4) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 99%

Differential DNA methylation at conserved non-genic elements and evidence for transgenerational inheritance following developmental exposure to mono(2-ethylhexyl) phthalate and 5-azacytidine in zebrafish

Kamstra

Sales

Aleström

et al. 2017

Epigenetics & Chromatin

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“…In general, this approach falls into the category of “MSRE-Seq” methods. There have been other methodological implementations of methylation-sensitive assays like this, most recently “DREAM” in 2012 (Jelinek et al, 2012; van Esterik et al, 2015; Bouwmeester et al, 2016), but the challenge of assessing the probability of methylation events from NGS sequence data across whole-genomes has limited earlier work from being successfully adopted in the field beyond a few laboratories.…”

Section: Methodsmentioning

confidence: 99%

Epigenetic DNA Methylation Profiling with MSRE: A Quantitative NGS Approach Using a Parkinson's Disease Test Case

Marsh

Cottrell

Goldman³

2016

Front. Genet.

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Epigenetics is a rapidly developing field focused on deciphering chemical fingerprints that accumulate on human genomes over time. As the nascent idea of precision medicine expands to encompass epigenetic signatures of diagnostic and prognostic relevance, there is a need for methodologies that provide high-throughput DNA methylation profiling measurements. Here we report a novel quantification methodology for computationally reconstructing site-specific CpG methylation status from next generation sequencing (NGS) data using methyl-sensitive restriction endonucleases (MSRE). An integrated pipeline efficiently incorporates raw NGS metrics into a statistical discrimination platform to identify functional linkages between shifts in epigenetic DNA methylation and disease phenotypes in samples being analyzed. In this pilot proof-of-concept study we quantify and compare DNA methylation in blood serum of individuals with Parkinson's Disease relative to matched healthy blood profiles. Even with a small study of only six samples, a high degree of statistical discrimination was achieved based on CpG methylation profiles between groups, with 1008 statistically different CpG sites (p < 0.0025, after false discovery rate correction). A methylation load calculation was used to assess higher order impacts of methylation shifts on genes and pathways and most notably identified FGF3, FGF8, HTT, KMTA5, MIR8073, and YWHAG as differentially methylated genes with high relevance to Parkinson's Disease and neurodegeneration (based on PubMed literature citations). Of these, KMTA5 is a histone methyl-transferase gene and HTT is Huntington Disease Protein or Huntingtin, for which there are well established neurodegenerative impacts. The future need for precision diagnostics now requires more tools for exploring epigenetic processes that may be linked to cellular dysfunction and subsequent disease progression.

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“…Zebrafish maintenance and embryonal exposures were as reported previously [7]. In short, zebrafish were housed in a ZebTec flow-through system with 7.5 L tanks, a light-dark cycle of 14–10 h, and automatically controlled water conditions, including a temperature of 26 ± 1 °C, pH at 7.5 ± 0.5, conductivity at 500 ± 100 µs/cm, and CaO levels at 10–250 mg/L.…”

Section: Methodsmentioning

confidence: 99%

“…Recent reviews summarize experimental approaches and models that could be employed to investigate mechanisms of DOHaD and associated predictive biomarkers [5,6]. One such suggested model is Danio rerio (zebrafish), and therefore we previously explored effects on DNA methylation after embryonal exposure in this model with environmental contaminants [7]. Here, we expand that study with investigations into delayed effects of embryonal exposure to one such contaminant, cadmium.…”

Section: Introductionmentioning

confidence: 99%

“…Here, we expand that study with investigations into delayed effects of embryonal exposure to one such contaminant, cadmium. Cadmium has known epigenetic effects [7,8] and is a ubiquitous environmental pollutant leading to chronic low-dose exposure in humans through vegetables, cereals and tobacco [9]. In the experimental part of this study, zebrafish embryos were exposed (0–72 h post fertilization, hpf) to cadmium (CdCl 2 ).…”

Section: Introductionmentioning

confidence: 99%

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Programmed Effects in Neurobehavior and Antioxidative Physiology in Zebrafish Embryonically Exposed to Cadmium: Observations and Hypothesized Adverse Outcome Pathway Framework

Ruiter

Sippel

Bouwmeester

et al. 2016

IJMS

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Non-communicable diseases (NCDs) are a major cause of premature mortality. Recent studies show that predispositions for NCDs may arise from early-life exposure to low concentrations of environmental contaminants. This developmental origins of health and disease (DOHaD) paradigm suggests that programming of an embryo can be disrupted, changing the homeostatic set point of biological functions. Epigenetic alterations are a possible underlying mechanism. Here, we investigated the DOHaD paradigm by exposing zebrafish to subtoxic concentrations of the ubiquitous contaminant cadmium during embryogenesis, followed by growth under normal conditions. Prolonged behavioral responses to physical stress and altered antioxidative physiology were observed approximately ten weeks after termination of embryonal exposure, at concentrations that were 50–3200-fold below the direct embryotoxic concentration, and interpreted as altered developmental programming. Literature was explored for possible mechanistic pathways that link embryonic subtoxic cadmium to the observed apical phenotypes, more specifically, the probability of molecular mechanisms induced by cadmium exposure leading to altered DNA methylation and subsequently to the observed apical phenotypes. This was done using the adverse outcome pathway model framework, and assessing key event relationship plausibility by tailored Bradford-Hill analysis. Thus, cadmium interaction with thiols appeared to be the major contributor to late-life effects. Cadmium-thiol interactions may lead to depletion of the methyl donor S-adenosyl-methionine, resulting in methylome alterations, and may, additionally, result in oxidative stress, which may lead to DNA oxidation, and subsequently altered DNA methyltransferase activity. In this way, DNA methylation may be affected at a critical developmental stage, causing the observed apical phenotypes.

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Zebrafish embryos as a screen for DNA methylation modifications after compound exposure

Cited by 59 publications

References 63 publications

Differential DNA methylation at conserved non-genic elements and evidence for transgenerational inheritance following developmental exposure to mono(2-ethylhexyl) phthalate and 5-azacytidine in zebrafish

Differential DNA methylation at conserved non-genic elements and evidence for transgenerational inheritance following developmental exposure to mono(2-ethylhexyl) phthalate and 5-azacytidine in zebrafish

Epigenetic DNA Methylation Profiling with MSRE: A Quantitative NGS Approach Using a Parkinson's Disease Test Case

Programmed Effects in Neurobehavior and Antioxidative Physiology in Zebrafish Embryonically Exposed to Cadmium: Observations and Hypothesized Adverse Outcome Pathway Framework

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