ABSTRACT

Organisms exposed to endocrine disruptors in early life can show altered phenotype later in adulthood. Although the mechanisms underlying these long-term effects remain poorly understood, an increasing body of evidence points towards the potential role of epigenetic processes. In the present study, we exposed hatchlings of an isogenic lineage of the self-fertilizing fish mangrove rivulus for 28 days to 4 and 120 ng/L of 17-α-ethinylestradiol. After a recovery period of 140 days, reduced representation bisulphite sequencing (RRBS) was performed on the liver in order to assess the hepatic genome-wide methylation landscape. Across all treatment comparisons, a total of 146 differentially methylated fragments (DMFs) were reported, mostly for the group exposed to 4 ng/L, suggesting a non-monotonic effect of EE2 exposure. Gene ontology analysis revealed networks involved in lipid metabolism, cellular processes, connective tissue function, molecular transport and inflammation. The highest effect was reported for nipped-B-like protein B (NIPBL) promoter region after exposure to 4 ng/L EE2 (+ 21.9%), suggesting that NIPBL could be an important regulator for long-term effects of EE2. Our results also suggest a significant role of DNA methylation in intergenic regions and potentially in transposable elements. These results support the ability of early exposure to endocrine disruptors of inducing epigenetic alterations during adulthood, providing plausible mechanistic explanations for long-term phenotypic alteration. Additionally, this work demonstrates the usefulness of isogenic lineages of the self-fertilizing mangrove rivulus to better understand the biological significance of long-term alterations of DNA methylation by diminishing the confounding factor of genetic variability.

摘要

在生命早期暴露于内分泌干扰物的生物体可能在成年后期表现出改变的表型。尽管这些长期影响背后的机制仍然知之甚少，但越来越多的证据表明表观遗传过程的潜在作用。在本研究中，我们将自受精鱼红树林河的同基因谱系的幼体暴露于 4 和 120 ng/L 的 17-α-炔雌醇 28 天。在 140 天的恢复期后，对肝脏进行了降低代表性的亚硫酸氢盐测序 (RRBS)，以评估肝脏全基因组甲基化景观。在所有治疗比较中，共报告了 146 个差异甲基化片段 (DMF)，主要用于暴露于 4 ng/L 的组，表明 EE2 暴露的非单调效应。基因本体分析揭示了涉及脂质代谢、细胞过程、结缔组织功能、分子转运和炎症的网络。据报道，效果最高的是暴露于 4 ng/L EE2 后，nipped -B 样蛋白 B (NIPBL) 启动子区域 (+ 21.9%)，表明 NIPBL 可能是 EE2 长期影响的重要调节剂。我们的结果还表明 DNA 甲基化在基因间区域和可能在转座因子中的重要作用。这些结果支持在成年期早期暴露于内分泌干扰物诱导表观遗传改变的能力，为长期表型改变提供合理的机制解释。此外，这项工作证明了自花红树林河的同基因谱系通过减少遗传变异的混杂因素来更好地理解 DNA 甲基化长期改变的生物学意义。