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

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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.