The inland silverside, Menidia beryllina, is a euryhaline fish and a model organism in ecotoxicology. We previously showed that exposure to picomolar (ng/L) levels of endocrine disrupting chemicals (EDCs) can cause a variety of effects in M. beryllina, from changes in gene expression to phenotypic alterations. Here we explore the potential for early life exposure to EDCs to modify the epigenome in silversides, with a focus on multi- and transgenerational effects. EDCs included contaminants of emerging concern (the pyrethroid insecticide bifenthrin and the synthetic progestin levonorgestrel), as well as a commonly detected synthetic estrogen (ethinylestradiol), and a synthetic androgen (trenbolone) at exposure levels ranging from 3 to 10 ng/L. In a multigenerational experiment, we exposed parental silversides to EDCs from fertilization until 21 days post hatch (dph). Then we assessed DNA methylation patterns for three generations (F0, F1, and F2) in whole body larval fish using reduced representation bisulfite sequencing (RRBS). We found significant (α = 0.05) differences in promoter and/or gene body methylation in treatment fish relative to controls for all EDCs and all generations indicating that both multigenerational (F1) and transgenerational (F2) effects that were caused by strict inheritance of DNA methylation alterations and the dysregulation of epigenetic control mechanisms. Using gene ontology and pathway analyses, we found enrichment in biological processes and pathways representative of growth and development, immune function, reproduction, pigmentation, epigenetic regulation, stress response and repair (including pathways important in carcinogenesis). Further, we found that a subset of potentially EDC responsive genes (EDCRGs) were differentially methylated across all treatments and generations and included hormone receptors, genes involved in steroidogenesis, prostaglandin synthesis, sexual development, DNA methylation, protein metabolism and synthesis, cell signaling, and neurodevelopment. The analysis of EDCRGs provided additional evidence that differential methylation is inherited by the offspring of EDC-treated animals, sometimes in the F2 generation that was never exposed. These findings show that low, environmentally relevant levels of EDCs can cause altered methylation in genes that are functionally relevant to impaired phenotypes documented in EDC-exposed animals and that EDC exposure has the potential to affect epigenetic regulation in future generations of fish that have never been exposed.

中文翻译：

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生命早期暴露于环境相关水平的内分泌干扰物会推动鱼类模型的多代和跨代表观遗传变化

内陆银鱼，Menidia beryllina，是一种广盐性鱼类，也是生态毒理学的模式生物。我们之前表明，暴露于皮摩尔 (ng/L) 水平的内分泌干扰化学物质 (EDC) 会对 M. beryllina 产生多种影响，从基因表达的变化到表型的改变。在这里，我们探索了生命早期暴露于 EDC 以修饰银侧表观基因组的潜力，重点是多代和跨代效应。EDCs 包括新出现的污染物（拟除虫菊酯杀虫剂联苯菊酯和合成孕激素左炔诺孕酮），以及一种常见的合成雌激素（炔雌醇）和一种合成雄激素（群勃龙），暴露水平为 3 至 10 ng/L。在多代实验中，我们从受精到孵化后 21 天 (dph) 将亲本银边暴露于 EDC。然后，我们使用减少代表性亚硫酸氢盐测序 (RRBS) 评估了全身幼鱼中三代（F0、F1 和 F2）的 DNA 甲基化模式。我们发现处理鱼的启动子和/或基因体甲基化相对于所有 EDC 和所有世代的对照存在显着 (α = 0.05) 差异，表明由 DNA 严格遗传引起的多代 (F1) 和跨代 (F2) 效应甲基化改变和表观遗传控制机制的失调。使用基因本体和通路分析，我们发现了代表生长和发育、免疫功能、繁殖、色素沉着、表观遗传调控、应激反应和修复（包括在致癌作用中重要的途径）。此外，我们发现潜在 EDC 反应基因 (EDCRG) 的一个子集在所有治疗和世代中都发生了差异甲基化，包括激素受体、参与类固醇生成的基因、前列腺素合成、性发育、DNA 甲基化、蛋白质代谢和合成、细胞信号传导、和神经发育。EDCRGs 的分析提供了额外的证据，表明差异甲基化是由 EDC 处理的动物的后代遗传的，有时在从未接触过的 F2 代中遗传。这些发现表明，低、