Programming the oocyte for the next generation

这个研究项目的重点是人体最不寻常的细胞：卵母细胞。事实上，这种细胞具有独特的能力，因为在排卵前几周，它积累指令，以便控制/允许受精后早期胚胎发育。这一异常现象是本次研究计划的重点。我们知道，在某种程度上，卵巢控制着卵母细胞的质量，这体现在它成为胚胎的能力上。然而，起源于卵巢的信号转导过程以及卵母细胞对这些信号的分子反应尚未被描述。根据来源卵泡的大小和分化程度，卵母细胞可能具有不同的质量水平。我们认为，卵泡分化引发了一系列事件，向卵母细胞预示着接下来的轨迹：排卵或闭锁。例如，在奶牛身上，药物被用来刺激几个卵子的排卵，以加速良好遗传学的传播，但尽管我们已经开发出产生良好胚胎的药物干预措施，但控制质量获得的机制仍然难以捉摸。通过收集相当独特的卵泡样本，我们已经生成了大量数据，从而可以创建一个旨在捕捉卵泡分化复杂性的虚拟卵泡。利用独特的激素诱导的卵泡环境与良好的胚胎产量相关，我们已经识别出颗粒细胞分泌的分子可能在排卵前的最后几天影响卵丘细胞和卵母细胞的分化。现在有机会利用组学、生物信息学和体外模型来拼凑卵母细胞能力之谜，我们相信我们处于正确的位置来实现这一目标。 利用最先进的分子工具，如转录组和生物信息学，现在可以探索卵泡中的基因调控和表达，从颗粒细胞到卵丘细胞，最后到连续的卵母细胞。功能基因组学的实际挑战是将数据组织到生理和相关的背景中。这个项目对卵泡向卵母细胞发出的信号和嵌入在卵子中的特殊母体指令感兴趣，这些指令指导着良好胚胎和健康小牛的发育。我们已经有独家数据，这些数据产生了几个有待验证的假设。我们的研究表明，可能启动卵泡向卵母细胞发出信号的事件之一是卵泡排卵能力的发展，这伴随着颗粒细胞上LH受体的出现，并伴随着这些细胞从黄体中的上皮型向间叶型的转换。 我们实验室提供了不同的模型来研究信号级联反应的相关变化：1)颗粒细胞的体外原代培养以评估单个基因对下游靶点的影响；2)卵丘封闭的未成熟卵母细胞培养过程中的信号诱导或模仿；3)卵母细胞显微注射吗啡或带有调控3‘UTR或化学激动剂的特定报告RNA以验证所确定的途径。 有了我们积累的所有特殊数据，我们通过合作建立或获得的工具，以及我们提出的模型，我们希望揭开让奶牛产下好鸡蛋的特定分子序列。这种对功能的理解将允许直接和间接干预奶牛的繁殖，并有助于保持我们行业多年来获得的领先地位。

This research program is focused on the most extraordinary cell of the body: the oocyte. Indeed this cell has unique capacities as, in the weeks preceding ovulation, it accumulates instructions in order to control/allow early embryonic development following fertilization. This exceptional phenomenon is the focus of this research program. We know that somehow, the ovary controls the quality of the oocyte, which is demonstrated by its ability to become an embryo. However, the signal transduction processes originating in the ovary and the oocyte molecular response to these signals are not characterized. Oocytes may have different levels of quality depending on the size and differentiation level of the follicles of origin. We believe that there is a cascade of events triggered by follicular differentiation that indicates to the oocyte the trajectory to follow: ovulation or atresia. In cows for example, drugs are used to stimulate the ovulation of several eggs to accelerate the dissemination of good genetics but although we have developed pharmaceutical interventions that generate good embryos, the mechanisms controlling quality acquisition remain elusive. With quite a unique collection of follicular samples, we have generated a tremendous amount of data allowing for the creation of a virtual folliculome that aims to capture the complexity of the follicle differentiation. Using unique specific hormone-induced follicular contexts associated with good yields of embryos, we have identified molecules secreted by granulosa cells that may impact the differentiation of cumulus cells and the oocyte in the last days prior to ovulation. There is now an opportunity to assemble the puzzle of oocyte competence using omics, bioinformatics and in vitro models and we believe that we are rightly positioned to achieve that. Using state-of-the-art molecular tools such as transcriptomics and bioinformatics, it is now possible to explore the gene regulation and expression in follicles starting with granulosa cells moving on to cumulus cells and finally to oocytes in a continuum. The actual challenge of functional genomics is the organisation of the data into a physiological and relevant context. This program is interested in the follicular signaling to the oocyte and the special maternal instructions that are imbedded in the egg and that direct the development of a good embryo and a healthy calf. We already have exclusive data which generated several hypotheses to be validated. Our studies indicate that one of the events that could initiate the follicular signaling to the oocyte is the development of the capacity of the follicle to ovulate, which occurs concomitantly with the appearance of LH receptors on granulosa cells, and continues with the switch of these cells from an epithelial type to the mesenchymal type found in the corpus luteum. Different models are available in our lab to study the relevant changes in signaling cascade; 1) in vitro primary cultures of granulosa cells to assess individual gene effects on downstream targets, 2) signal induction or mimicking during culture of cumulus-enclosed immature oocytes and 3) oocyte microinjection with morpholinos or specific reporter RNAs with regulatory 3’UTR or chemical agonists to validate the pathway identified. With all the exceptional data that we have accumulated, the tools we have built or acquired through collaboration and the models that we propose, we hope to unravel the specific molecular sequence of events that makes a good egg in dairy cows. This functional understanding will allow direct and indirect interventions in dairy cow’s reproduction and help to maintain the leadership position that our industry has gained over the years.