Molecular mechanisms by which estrogens affect male germ cell programming during development

雌激素影响发育过程中雄性生殖细胞编程的分子机制

• 批准号: RGPIN-2014-04607
• 负责人: Delbes, Geraldine
• 金额: $ 2.11万
• 依托单位: Institut national de la recherche scientifique
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2015
• 资助国家: 加拿大
• 起止时间: 2015-01-01 至 2016-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=587910
• 关键词: Molecular; mechanisms; estrogens; affect; male

Germ cell development in mammals represents a unique and complex process. The fetal and early postnatal windows of development are critical. Indeed, during this time window, germ cells (gonocytes) undergo epigenetic reprogramming, proliferation, quiescence and differentiation. Recent evidence have indicated that adult reproductive dysfunction can originate from misregulation of germ cell development during this specific period of development. We have demonstrated that this period of development is particularly sensitive to endocrine disruption by estrogen-like compounds. Yet, the signaling mechanisms and molecular basis involved in perinatal gonocyte programming and the targets of xenoestrogens remain poorly understood. My research program aims to elucidate the underlying molecular basis for fetal and neonatal male germ cell development in mammals and determining how the misregulation of these processes might alter reproductive function in adulthood. The central hypothesis is that the epigenetic reprogramming that occurs during gonocyte development, through de novo DNA methylation and histones post-translational modifications, regulates the expression of key regulators genes that are necessary for germ cell programming, and that such mechanism can be targeted by the estrogen signalling pathway. Taking advantages of a transgenic rat model in which only germ cells express the green fluorescent protein, we will elucidate how the estrogen signaling can affect gene expression and the epigenetic machinery in gonocytes by using both in vivo and in vitro models. In Objective 1, we will characterize the short-term impact of in vivo gestational exposure to ethinyl estradiol and genistein when gonocytes proliferate and de novo DNA methylation occurs. Comparison between treatments will discriminate between the two estrogen receptors pathways. The impact on gene expression will be evaluated using genome wide transcriptome analysis. Further analysis of the effect of treatment on epigenetic reprogramming and cell differentiation will be done. Finally, in depth comprehension of epigenetic regulation of genes which expression is affected by estrogenic treatment will be obtained. In Objective 2, we will use two in vitro models and pharmaceutical inhibitors to disrupt the enzymes involved in DNA methylation and histone modifications, we will reveal the crosstalk between the estrogen signaling and the epigenetic machinery in regulating gene expression in gonocytes. We will use an immortalized gonocyte cell line and organ culture to determine how the misregulation of epigenetic mechanisms regulates gonocytes proliferation, differentiation or cell death, and how estrogens alter such regulation. Altogether, these studies will provide a better understanding of how estrogens affect male germ cell development by targeting the epigenetic reprogramming. Fetal development represents a key and unique period in cell programming when cells acquire information for a lifetime. Understanding the molecular mechanisms underlying male germ cell programming and how xenobiotics can disrupt these mechanisms is essential to establish the dependency of reproductive outcomes on fetal programming.

哺乳动物的生殖细胞发育是一个独特而复杂的过程。胎儿和出生后早期的发育窗口是至关重要的。事实上，在此时间窗期间，生殖细胞（生殖细胞）经历表观遗传重编程、增殖、静止和分化。最近的证据表明，成人生殖功能障碍可能源于生殖细胞发育在这一特定时期的发展失调。我们已经证明，这一时期的发展是特别敏感的雌激素样化合物的内分泌干扰。然而，参与围产期生殖细胞编程的信号传导机制和分子基础以及异种雌激素的靶点仍然知之甚少。我的研究项目旨在阐明哺乳动物胎儿和新生儿雄性生殖细胞发育的潜在分子基础，并确定这些过程的失调如何改变成年后的生殖功能。核心假设是，在生殖细胞发育过程中发生的表观遗传重编程，通过从头DNA甲基化和组蛋白翻译后修饰，调节生殖细胞编程所必需的关键调控基因的表达，并且这种机制可以通过雌激素信号通路靶向。利用转基因大鼠模型，其中只有生殖细胞表达的绿色荧光蛋白，我们将阐明雌激素信号如何影响基因表达和表观遗传机制在性腺细胞通过使用在体内和体外模型。在目标1中，我们将描述当生殖细胞增殖和从头DNA甲基化发生时，体内妊娠期暴露于炔雌醇和染料木黄酮的短期影响。治疗之间的比较将区分两种雌激素受体途径。将使用全基因组转录组分析评价对基因表达的影响。将进一步分析治疗对表观遗传重编程和细胞分化的影响。最后，深入理解的表观遗传调控的基因表达的影响，雌激素治疗将获得。目的二是利用两种体外模型和药物抑制剂，通过阻断DNA甲基化和组蛋白修饰相关酶，揭示雌激素信号通路与表观遗传机制在调控生殖细胞基因表达中的相互作用。我们将使用永生化的生殖细胞系和器官培养来确定表观遗传机制的失调如何调节生殖细胞的增殖、分化或细胞死亡，以及雌激素如何改变这种调节。总之，这些研究将提供一个更好地了解雌激素如何影响男性生殖细胞发育的目标表观遗传重编程。胎儿发育是细胞编程中的一个关键和独特的时期，细胞在此期间获得一生的信息。了解男性生殖细胞编程的分子机制以及外源性物质如何破坏这些机制对于建立生殖结果对胎儿编程的依赖性至关重要。