Project 2: Role of Glycosylation in FSH Signaling in FSH Target Cells

项目摘要/摘要--项目2(戴维斯) 卵巢衰老大约从30岁开始，在接下来的10到15年里，明显表现为不孕不育和 卵巢储备的减少反映在抗苗勒氏激素水平的下降， 生物活性低糖基化卵泡刺激素(FSH)与血清低水平 活性完全糖基化的卵泡刺激素。卵巢老化的过程可能会导致转录组内的修饰 卵巢细胞可以降低老年妇女生育亲生子女的几率。调查人员 证实了低糖基化的卵泡刺激素具有更强的生物活性 体外和体内的生理相关模型。已确定FSH可激活多种细胞信号 但FSH糖类如何调节颗粒细胞转录组还不清楚。遗漏自 以前的研究是对CREB和YAP1等关键转录因子在 卵泡刺激素糖体对颗粒细胞增殖、凋亡、分化的影响 衰老。我们的中心假设是FSH18/21和FSH24不同地调节关键转录 CREB和YAP1因子调节颗粒细胞功能。我们假设衰老促进一种与生俱来的 调节卵泡发育的关键转录事件的失调。这个项目利用了老化 小鼠模型和利用从青年和老年生殖年龄回收的颗粒细胞的体外模型 女人和老鼠。该项目将确定FSH糖型对转录程序的影响 幼龄和高龄生殖年龄小鼠和妇女的颗粒细胞。我们将利用基因损失和获得 研究CREB和YAP1的功能，并进行RNA-SEQ，深入分析它们在基因表达中的作用 对FSH糖体的反应。我们将使用最先进的方法(削减和运行)来确定差异化 青年和高龄育龄妇女颗粒细胞中CREB和YAP1的结合模式。 我们还将对这些相同的样本进行atac-seq，以分析年轻和 年纪较大的个体。将进行rna-seq以捕获整个转录组，并进行生物信息学组装。 个体组学方法允许全面了解衰老对反应的影响 转化为FSH糖体。我们将进行综合生物信息学分析，以构建全面的 了解FSH糖型调控的基因组格局和靶基因及细胞通路 作为年龄的函数。成功完成这些目标将揭示独特的卵巢转录信号 使用体外和体内方法相结合的方法，诱导FSH糖型作为年龄的函数。 利用小鼠和人类的衰老模型，结合转录和生物信息学分析，将 揭示新知识，为改善卵巢功能的方法提供信息，并转化为 不仅改善生殖健康，而且改善整体健康和长寿。

PROJECT SUMMARY/ABSTRACT – Project 2(Davis) Ovarian senescence begins about age 30 and over the next 10 to 15 years is manifest by infertility and a diminished ovarian reserve reflected in decreased Anti-Mullerian hormone levels, decreased expression of the biologically active hypo-glycosylated follicle stimulating hormone (FSH) and elevated serum levels of the less active fully glycosylated FSH. The process of ovarian aging may lead to modifications within the transcriptome of ovarian cells that reduce the chances of an older woman having her own biological children. The investigators of this P01 have established that hypo-glycosylated FSH has greater bioactivity when examined in physiologically relevant models in vitro and in vivo. It is established that FSH activates a variety of cell signaling pathways, but how FSH glycoforms regulate the granulosa cell transcriptome is not understood. Missing from previous studies is an understanding of the role that crucial transcription factors like CREB and YAP1 play in relaying the actions of FSH glycoforms to control granulosa cell proliferation, apoptosis, differentiation, and senescence. Our central hypothesis is that FSH18/21 and FSH24 differentially regulate the key transcription factors CREB and YAP1 to modulate granulosa cell function. We hypothesize that aging promotes an inherent dysregulation of key transcriptional events that regulate follicular development. This project makes use of aging mouse models and in vitro models using granulosa cells recovered from young and older reproductive age women and mice. This project will determine the effect of FSH glycoforms on transcriptional programs of granulosa cells of young and advanced reproductive age mice and women. We will employ genetic loss and gain of function of CREB and YAP1 and conduct RNA-seq for an in-depth gene expression analysis on their role in the response to FSH glycoforms. We will use a state-of-the-art approach (CUT&RUN) to identify differential binding patterns of CREB and YAP1 in granulosa cells from young and advanced reproductive aged women. We will also perform ATAC-seq on these same samples to profile chromatin accessibility between young and older individuals. RNA-seq will be performed to capture the entire transcriptome and bioinformatics will assemble the individual omics approaches allowing a comprehensive understanding of the impact of aging on the response to FSH glycoforms. We will conduct an integrative bioinformatics analysis to build a comprehensive understanding of the genomic landscape and target genes and cellular pathways regulated by FSH glycoforms as a function of age. Successful completion of these aims will reveal unique ovarian transcriptomic signatures induced by FSH glycoforms as a function of age using a combination of in vitro and in vivo approaches. Employment of mouse and human aging models coupled with transcriptomics and bioinformatics analysis will reveal new knowledge that informs approaches to improve ovarian function and translate to approaches that improve not only reproductive health, but overall health and longevity.