Mechanisms of EDC Effects via Small-RNA Cargo in Sperm Epididymosomes

精子附睾中小 RNA 货物的 EDC 作用机制

• 批准号: 10592593
• 负责人: ANDREA C GORE
• 金额: $ 19.81万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10592593
• 关键词: Adult; Agriculture; Alcohol consumption; Bioinformatics; Biology; Cell Differentiation process; Chemical Exposure; Chemical Industry; Chemical-Induced Change; Chronic stress; Competence; CpG Islands; DNA; DNA Methylation; Data; Development; Diet; Disease; Dose; Embryo; Endocrine Disruptors; Environment; Epididymis; Epigenetic Process; Epithelium; Exposure to; Fertilization; Fetal Development; Fetus; Functional disorder; Future Generations; Gene Expression; Germ Cells; Health; Heritability; Hormonal; Human; Industrialization; Inherited; Intervention; Lactation; Life; Link; Mammals; Mediating; Methylation; Modeling; Molecular; Oocytes; Outcome; Phenotype; Plastics; Play; Polychlorinated Biphenyls; Predisposition; Pregnancy; Probability; Process; Property; Rattus; Research; Sampling; Signal Transduction; Small RNA; Sperm Maturation; Testing; Transfer RNA; Untranslated RNA; Work; androgenic; bisulfite sequencing; deep sequencing; diphenyl; dosage; egg; epigenome; estrogenic; exposed human population; extracellular vesicles; feeding; fungicide; intergenerational; male; next generation sequencing; novel; offspring; pregnant; programs; reproductive tract; sperm cell; stem cells; success; transcriptome; vinclozolin

ABSTRACT Exposures to environmental endocrine-disrupting chemicals (EDCs), especially during early life, are strongly linked to adverse health outcomes. Furthermore, EDC exposures to the fetus, which also expose the germ cells (precursors to sperm and egg) within the fetus, can cause heritable epigenetic changes that are passed to future generations. Previous work has implicated epimutations of DNA CpG methylation as one molecular mechanism by which EDCs program the germline. Proposed work will explore this mechanism, which is under-studied, and in addition, seek to understand how external factors from the male reproductive tract involved in sperm maturation may themselves contribute to epimutations. More specifically, the proximal portion (caput) of the epididymis has a substantial influence on sperm during their final steps of maturation. Extracellular vesicles (EVs) from the epididymis – epididymosomes - released from the epithelium fuse and transfer their diverse molecular contents to sperm, including small noncoding RNAs (sncRNA), thereby contributing to the functional competency of sperm and to the developmental trajectory of a fertilized embryo. These sncRNAs have the capacity to regulate gene expression and direct DNA methylation in sperm. In the current proposal, we will establish whether and how two different EDC classes that signal via differing hormonal mechanisms cause direct CpG methylation changes to DNA, and/or change the transfer of sncRNA content of epididymosomes to sperm to indirectly cause epimutations. The two EDC classes studied here are the polychlorinated biphenyl mix Aroclor 1221 (A1221), an industrial mixture that has weakly estrogenic properties; and vinclozolin (VIN), an anti- androgenic fungicide. Both EDCs are still environmentally relevant. Rats are exposed to human relevant dosages of A1221 or VIN, or vehicle, through feeding the dams during gestation and lactation. In adult male rats, sncRNA cargo of EVs from caput epididymis and mature sperm from the cauda epididymis will be extracted and used for deep sequencing and bioinformatics. DNA from sperm in the same samples will be subjected to reduced representation bisulfite sequencing to identify epimutations in CpG islands. These data will establish a definitive epigenetic profile that will allow us to pinpoint the origin of EDC induced epimutations. This proposal is biomedically relevant, as all humans are exposed to EDCs, with mechanisms of sperm maturation highly conserved across mammals.

抽象的 暴露于环境内分泌干扰化学物质 (EDC)，尤其是在生命早期，会严重影响 与不良健康结果有关。此外，EDC 暴露给胎儿，这也暴露了生殖细胞 （精子和卵子的前体）在胎儿体内，可能会导致可遗传的表观遗传变化，并传递给未来 几代人。先前的工作表明 DNA CpG 甲基化的表观突变是一种分子机制 EDC 通过它对种系进行编程。拟议的工作将探索这一尚未得到充分研究的机制，并且 此外，寻求了解男性生殖道的外部因素如何影响精子 成熟本身可能会导致表观突变。更具体地说，近端部分（头） 附睾对精子成熟的最后阶段有重大影响。细胞外囊泡 （EV）来自附睾 - 附睾体 - 从上皮融合释放并转移其多样化 精子的分子内容，包括小非编码 RNA (sncRNA)，从而有助于功能 精子的能力和受精胚胎的发育轨迹。这些 sncRNA 具有 调节基因表达和指导精子 DNA 甲基化的能力。在当前的提案中，我们将 确定通过不同激素机制发出信号的两种不同 EDC 类别是否以及如何导致直接 CpG 甲基化改变 DNA，和/或改变附睾体的 sncRNA 内容向精子的转移 间接引起表突变。这里研究的两个 EDC 类别是多氯联苯混合物 Aroclor 1221 (A1221)，一种具有弱雌激素特性的工业混合物；和乙烯菌核利 (VIN)，一种抗 雄激素杀菌剂。这两种 EDC 仍然与环境相关。大鼠暴露于人类相关剂量 A1221 或 VIN 或媒介物，通过在妊娠和哺乳期间喂养母鼠。在成年雄性大鼠中，sncRNA 来自附睾头的 EV 货物和来自附睾尾的成熟精子将被提取并用于 深度测序和生物信息学。同一样本中精子的 DNA 将被减少 代表性亚硫酸氢盐测序来鉴定 CpG 岛的表观突变。这些数据将建立一个明确的 表观遗传图谱将使我们能够查明 EDC 诱导的表观突变的起源。这个提议是 生物医学相关，因为所有人都接触 EDC，精子成熟机制高度相关 在哺乳动物中保守。