Mechanisms of bisphenol A induced epigenomic alterations in the mammalian germline

项目总结/摘要 BPA广泛存在于许多消费品中。人类每天都暴露在BPA中，超过90% 的被调查者的尿液中有可检测到的BPA含量。肥胖、代谢紊乱，以及 神经系统疾病与人类和动物接触BPA有关。在啮齿类动物中， BPA的F0代妊娠雌性动物的肥胖表型在F2代中持续存在， F3及以后的世代。这些观察结果表明，暴露于BPA诱导表观遗传 在F1种系的改变，可以遗传间和转代。然而，机制 这些表观遗传改变通过这些机制得以建立、维持并传递给后代 大部分尚未开发。为了解决这个问题，我们将怀孕的雌性小鼠暴露在BPA中， 胚胎期7-13天，此时胎儿的生殖细胞正在被重新编程。我们发现接触BPA 导致一种肥胖表型，该表型通过母系和父系遗传至少4代。精子 从BPA暴露小鼠的肥胖后代中含有新的染色质可及位点， 未暴露的对照对应于CTCF和几种核激素受体的新结合位点。 重要的是，这些新位点存在于F2-F5小鼠的精子中，表明它们直接或间接地与精子中的蛋白质结合。 导致肥胖症的表型。总之，这些结果表明，表观遗传改变诱导的 双酚A可以通过雄性生殖系进行跨代传播。然而，我们不知道这些 改变最初是在生殖系中在重编程过程中获得的，或者它们在缺失的情况下是如何维持的。 BPA的原始刺激。在这里，我们提出实验来研究BPA如何诱导表观遗传变化 在早期生殖细胞中建立和维持。目标1中提出的工作将确定新的 在发育过程中不同时间点BPA暴露诱导的转录因子（TF）结合位点 原始生殖细胞（PGCs），精原细胞和精原干细胞（SSCs），并检查 TF结合的变化是否与这些细胞转录组的变化相关。在目标2中，我 将研究PGCs中TF结合的变化与DNA甲基化、组蛋白 修饰，和组蛋白变异体在新的TF网站在整个分化的男性生殖系。目标3 将分析在不存在以下情况下F2代生殖系中表观遗传改变的维持： BPA暴露，以及TF结合与DNA甲基化变化之间的时间关系， 染色质结构从拟议的研究中获得的知识将首次揭示BPA- 诱导的表观遗传改变在胚胎生殖系中的表观遗传重编程期间建立， 维持在后代的雄性生殖细胞中。这些研究将具有重要意义， 了解内分泌诱导表型的跨代遗传机制 干扰性化学品（EDCs）是设计干预措施以扭转这些影响的第一步。

PROJECT SUMMARY/ABSTRACT BPA is ubiquitously found in many consumer products. Humans are exposed to BPA daily and more than 90% of surveyed individuals have detectable levels of BPA in their urine. Obesity, metabolic disorders, and neurological disorders have been associated with BPA exposure in humans and animals. In rodents, exposure to BPA of F0 pregnant females results in obesity phenotypes in the F2 generation that persist in the unexposed F3 and subsequent generations. These observations suggest that exposure to BPA induces epigenetic alterations in the F1 germline that can be inherited inter- and transgenerationally. However, the mechanisms by which these epigenetic alterations are established, maintained, and transmitted to subsequent generations remain largely unexplored. To address this question, we exposed pregnant mouse females to BPA during embryonic days 7-13, when the germline of the fetus is being reprogrammed. We find that exposure to BPA leads to an obese phenotype that is transmitted maternally and paternally for at least 4 generations. Sperm from the obese progeny of BPA-exposed mice contain new chromatin accessible sites not present in unexposed controls corresponding to new binding sites for CTCF and several nuclear hormone receptors. Importantly, these new sites are present in sperm of F2-F5 mice, suggesting that they are directly or indirectly responsible for the obesity phenotype. Together, these results suggest that epigenetic alterations induced by BPA can be transmitted transgenerationally by the male germline. However, we do not know how these alterations are initially acquired in the germline during reprograming or how they are maintained in the absence of the original stimulus by BPA. Here we propose experiments to study how BPA-induced epigenetic changes are established and maintained in early germ line cells. The work proposed in Aim 1 will identify new transcription factor (TF) binding sites induced by BPA exposure at different time points during the development of the Primordial Germ Cells (PGCs), prospermatogonia, and Spermatogonial Stem Cell (SSCs), and examine whether the changes in TF binding are associated with changes in the transcriptome of these cells. In Aim 2, I will study the relationship between changes in TF binding in PGCs and alterations of DNA methylation, histone modifications, and histone variants at new TF sites throughout the differentiation of the male germline. Aim 3 will analyze the maintenance of epigenetic alterations in the germline of the F2 generation in the absence of BPA exposure, and the temporal relationship between TF binding and changes in DNA methylation and chromatin structure. The knowledge gained from the proposed studies will reveal for the first time how BPA- induced epigenetic alterations are established during epigenetic reprogramming in the embryonic germline and maintained in the male germ cells of subsequent generations. These studies will be highly significant in understanding the mechanisms of transgenerational inheritance of epiphenotypes induced by endocrine disrupting chemicals (EDCs), a first step towards designing interventions to reverse these effects.