Function of RUNX transcription factors in COCs

描述(申请人提供)：本方案的总体目标是确定卵丘卵母细胞复合体(COCs)成熟过程中涉及的分子/细胞机制(S)。COC的适当成熟是体内和体外繁殖成功的关键。我们对这一成熟过程缺乏完整的了解，限制了我们在体内管理生育能力的能力，并阻碍了体外受精和体外成熟等辅助生殖技术的实施。我们实验室和其他实验室最近的研究已经开始揭示一个小的核转录因子家族，RUNX1和RUNX2，作为参与COC扩张的关键转录调节因子。我们的初步数据表明，RUNX1和Runx2在排卵周围卵泡的卵丘细胞中被高度诱导表达。我们还发现RUNX1和RUNX2调节壁层颗粒细胞和/或卵丘细胞中Ptgs2(前列腺素生物合成的限速酶)和Hapln1(卵丘基质的稳定剂)的表达。已知这两种基因产物在COC扩展中起着关键作用。最重要的是，我们的初步研究表明，卵丘细胞中显性负RUNX的过度表达阻止了COC的扩张。基于这些新的发现，我们假设RUNX1/2是促性腺激素激增诱导的COC扩张所必需的必要转录调节因子。这一假设将通过首先证明RUNX1/2在卵丘细胞中表达的功能意义来检验(目标1)。其次，我们将确定RUNX1和RUNX2对其靶基因(目标2)的调控作用(S)。接下来，我们将确定卵丘细胞中RUNX1和RunX2表达的调控机制(S)(目标3)。这三个目标将用大鼠COCs进行研究。为了确保从大鼠研究中获得的数据对人类是可翻译的，RUNX1/2及其目标基因表达谱也将使用在整个排卵期收集的人类卵丘细胞进行验证。我们还将使用从接受体外受精的妇女获得的COCs来检验RUNX1/2及其靶基因的堆积表达与卵母细胞质量、受精和胚胎发育的相关性(目标#4)。这些人体研究将为未来的翻译/临床应用奠定基础。从拟议的研究中获得的信息不仅将促进我们对COC成熟所涉及的机制的理解，而且也将有助于未来的翻译/临床应用(S)，从而改善体内和体外生育的管理。

DESCRIPTION (provided by applicant): The overall goal of the present proposal is to determine the molecular/cellular mechanism(s) involved in the maturation process of cumulus oocyte complexes (COCs). Proper maturation of the COC is critical to reproductive success in vivo and in vitro. Our lack of a complete understanding of this maturation process restricts our ability to manage fertility in vivo and hinders implementation of ART such as in vitro fertilization and in vitro maturation. Recent studies by our laboratory and others have begun to shed light on a small family of nuclear transcription factors, RUNX1 and RUNX2, as key transcriptional regulators involved in COC expansion. Our preliminary data demonstrated that Runx1 and Runx2 expression is highly induced in cumulus cells of periovulatory follicles. We also found that RUNX1 and RUNX2 regulate the expression of Ptgs2 (a rate-limiting enzyme in the biosynthesis of prostaglandins) and Hapln1 (a stabilizer of cumulus matrix) in mural granulosa cells and/or cumulus cells. Both gene products are known to play crucial roles in COC expansion. Most importantly, our pilot study demonstrated that over-expression of dominant negative RUNX in cumulus cells blocked COC expansion. Based on these novel findings, we hypothesized that RUNX1/2 are essential transcriptional regulators necessary for the gonadotropin surge-induced COC expansion. This hypothesis will be tested by first demonstrating the functional significance of Runx1/2 expression in cumulus cells (Aim #1). Secondly, we will determine the regulation action(s) of RUNX1 and RUNX2 on their target genes (Aim #2). Next, we will determine the regulatory mechanism(s) of Runx1 and Runx2 expression in cumulus cells (Aim #3). These 3 Aims will be studied using rat COCs. To insure that the data obtained from rat studies is translatable to humans, RUNX1/2 and their target gene expression profile will be also verified using human cumulus cells collected throughout the periovulatory period. We will also examine the correlation of the cumulus expression of RUNX1/2 and their target genes to oocyte quality, fertilization, and embryo development using COCs obtained from women undergoing IVF (Aim #4). These human studies will serve as a foundation for future translational/clinical application. The information obtained from the proposed studies will not only advance our understanding of the mechanism involved in COC maturation, but also be instrumental for future translational/clinical application(s), thus leading to improved management of fertility in vivo and in vitro.