Intergenerational Arsenic Exposure on Mouse Epigenetics

项目概述：无机砷（iAs）是影响人类的最大毒性暴露之一 全球有超过1.4亿人通过受污染的饮用水暴露于iAs。iAs暴露 在怀孕期间破坏了发育中的后代的正常DNA甲基化模式，并导致 成人疾病，如II型糖尿病（T2 D）、心血管疾病和癌症。更具体地说， 妊娠期间的破坏减弱了F1发育后代表观基因组的适当重塑 并可能通过破坏胎儿原始生殖细胞（PGCs）而使其F2代子代死亡。但有一个 疾病表型和甲基化谱之间的相关性之间的理解有限， F1的后代此外，我们不知道iAs暴露是否会改变F2大细胞的表观遗传重塑。 如果F2代即使没有直接iAs也有发生iAs相关疾病的风险， exposure.从机制上讲，表观遗传重编程可以在原始生殖细胞内被破坏， 导致F2代后代的DNA甲基化差异和有害的代谢健康。该机制是作为 iAs通过甲基化酶促代谢，使用S-腺苷甲硫氨酸（SAM）作为甲基化酶。 团体捐赠者SAM通常也用于DNA甲基化，以通过酶促甲基化维持表观基因组。 将甲基添加到未修饰的胞嘧啶上。因此，SAM对甲基化的竞争 途径导致甲基缺乏，导致iAs转化减少（毒性增加）和DNA减少 甲基化（基因表达的错误调节）。核心假设是， 子宫内iAs暴露将影响F1和F2成人内的表观遗传特征和疾病表型发作 尽管在这些世代中没有直接接触砷。目标1将建立 iAs对胚胎发生过程中F1表观遗传重塑的影响，重点关注与以下相关的基因： 成年期肥胖和T2 D的发展。为了证实iAs暴露对母体的影响， 表观遗传重构，经验证的AgglomaAvy小鼠模型表观遗传生物传感器将用于指示损失 或通过改变毛色获得DNA甲基化。目标2将描述F2（成年祖代）效应 砷暴露对F1原始生殖细胞表观遗传重构的影响。为了实现这两个目标，全球DNA 将通过液相色谱法定量体细胞和生殖组织中的甲基化和羟甲基化 质谱和通过焦磷酸测序的位点特异性甲基化。葡萄糖耐量和身体组成 将在成年F1和F2后代中测量，作为成年期存在疾病的指征。总的来说，这些 实验将提供一个更好的理解iAs重金属暴露对代际的影响， 表观遗传学，成年疾病在后代中的发病，以及多代表观遗传学的目标 重新编程本研究响应NIEHS战略计划目标，目标1（基础生物学研究）， 目标2（个体易感性）、目标4（暴露体）和目标6（预测毒理学）。

PROJECT SUMMARY: Inorganic Arsenic (iAs) is one of the largest toxic exposures to impact humanity worldwide, with over 140 million people exposed to iAs via contaminated drinking water. Exposure to iAs during pregnancy disrupts normal DNA methylation patterns in developing offspring and leads to the onset of adult diseases such as type II diabetes (T2D), cardiovascular disease, and cancers. More specifically, disruption during pregnancy attenuates the proper remodeling of the epigenome of the F1 developing offspring and potentially its F2 grand-offspring via disruption of fetal primordial germ cells (PGCs). However, there is a limited understanding between the correlation between the disease phenotype and methylation profile within the F1 offspring. Furthermore, we do not know whether iAs exposure alters epigenetic remodeling in F2 grand- offspring and if the F2 generation is also at risk for developing iAs associated diseases even without direct iAs exposure. Mechanistically, epigenetic reprogramming could be disrupted within the primordial germ cells, resulting in differential DNA methylation and detrimental metabolic health in F2 offspring. The mechanism is as follows: iAs is metabolized enzymatically by methylation, using S-adenosylmethionine (SAM) as a methyl group donor. SAM is also normally used in DNA methylation to maintain the epigenome via the enzymatic addition of a methyl group to unmodified cytosines. Thus, the competition of SAM for both methylation pathways results in methyl group deficiency leading to less iAs conversion (increased toxicity) and less DNA methylation (mis-regulation of gene expression). The central hypothesis is that intergenerational effects of in utero iAs exposure will impact the epigenetic profile and onset of disease phenotypes within F1 and F2 adult offspring, despite the life-long absence of direct arsenic exposure within these generations. Aim 1 will establish the effects of iAs on F1 epigenetic remodeling during embryogenesis, focusing on genes associated with development of obesity and T2D during adulthood. To confirm the maternal effect of iAs exposure on epigenetic remodeling, the validated Agouti Avy mouse model epigenetic biosensor will be used to indicate loss or gain of DNA methylation by shifting coat color. Aim 2 will characterize the F2 (adult grand-offspring) effects of arsenic exposure on epigenetic remodeling in F1 primordial germ cells. For both aims, global DNA methylation and hydroxymethylation will be quantified in somatic and germ tissues by liquid chromatography mass-spectrometry and site specific methylation by pyrosequencing. Glucose tolerance and body composition will be measured in adult F1 and F2 offspring as indications of disease present in adulthood. Collectively, these experiments will provide a better understanding of the effects of iAs heavy metal exposure on intergenerational epigenetics, the onset of adult disease in subsequent generations, and targets of multigenerational epigenetic reprogramming. This study responds to the NIEHS Strategic Plan goals, Goal 1 (Basic Biological Research), Goal 2 (Individual Susceptibility) Goal 4 (Exposome) and Goal 6 (Predictive Toxicology).