The role of Core Binding Factors (CBFs) in the periovulatory process

核心结合因子 (CBF) 在围排卵过程中的作用

• 批准号: 8127891
• 负责人: MISUNG JO
• 金额: $ 7.13万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-08-12 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8127891
• 关键词: Animal Model; Binding; Cells; Complex; Core-Binding Factor; DNA Binding; Data; Development; Diagnostic; Disease; Evaluation; Family; Female; Fertility; Gene Expression; Gene Expression Profile; Gene Targeting; Genes; Genetically Modified Animals; Genotype; Goals; Gonadotropins; Human; In Vitro; Infertility; Knockout Mice; Knowledge; Laboratories; Lead; Light; Luteinization; Mediator of activation protein; Molecular; Mus; Mutant Strains Mice; Nuclear Family; Oocytes; Ovarian; Ovary; Ovulation; Pathology; Phenotype; Physiological; Physiology; Pilot Projects; Play; Process; Proteins; RUNX1 gene; Regulatory Pathway; Rodent; Role; Staging; Technology; Testing; Time; Transgenic Mice; Up-Regulation; base; corpus luteum; egg; in vitro Model; in vivo; insight; mouse model; novel; null mutation; public health relevance; transcription factor

DESCRIPTION (provided by applicant): The preovulatory gonadotropin surge initiates the final process of follicular maturation that culminates in ovulation of an expanded cumulus-oocyte complex (COC) and formation of the corpus luteum (CL). One essential step elicited by the gonadotropin surge required for successful follicular maturation is the expression of specific transcription factors. However, our knowledge of the identity and regulatory actions of LH-induced key transcription factors remains very limited. Recent studies from our laboratory and others shed light on a small family of nuclear transcription factor, Core binding factor (CBF), as a key transcriptional regulator involved in periovulatory processes. CBF is composed of two subunits; DNA binding alpha-subunit encoded by one of Runx1, Runx2, and Runx3 genes and non-DNA binding beta-subunit, CBF2. To be functional, RUNX proteins need to be dimerized with CBF2. Our preliminary data showed the rapid induction of CBF components (Runx1, Runx2 and CBF2) by the LH surge in periovulatory follicles. Using an in vitro model, we further demonstrated that CBFs (RUNX1/CBF2 and RUNX2/CBF2) regulate the expression of periovulatory genes that are known to be critical for COC expansion, ovulation, and luteinization. Moreover, our pilot study revealed that inhibition of RUNX activity blocked COC expansion in vitro. Based on these novel findings, we hypothesized that CBFs are key transcriptional regulators necessary for successful COC expansion, ovulation, and luteinization. Homozygous null mutation of genes for CBF components in mice results in early lethality and, consequently, fails to define the function of CBFs in the ovary. The additional challenge in demonstrating the physiological importance of CBFs in vivo is the overlapping expression of Runx1 and Runx2 and their functional redundancy in periovulatory follicles. To circumvent this problem, we propose to establish a novel transgenic mouse model in which CBFs are inactivated specifically in ovarian cells. This will be accomplished by deleting CBF2 in ovarian cells using Cre-lox technology. Since CBF2 is a binding partner for both RUNX1 and RUNX2, targeted deletion of CBF2 abrogates the activity of all CBFs (RUNX1/CBF2 and RUNX2/CBF2). Using this transgenic mouse model, we will test the hypothesis that targeted inactivation of CBFs results in defective COC expansion, ovulation, and luteinization by examining the ovarian phenotype of this mutant mouse (Specific Aim #1). Using this mutant mouse ovary, we will identify the genes downstream of CBFs in periovulatory follicular cells and begin to delineate the transcriptional regulatory pathways necessary for the final stage of periovulatory follicle development (Specific Aim #2). These studies will establish a genetically modified animal model not only to define the in vivo ovarian function of CBFs, but also to identify the transcriptional regulatory machinery that controls the periovulatory process. Information derived from this proposal will provide new insight into the mechanism(s) involved in COC expansion, ovulation, and CL formation. Gaining a thorough understanding of cellular/molecular mechanisms of the periovulatory process will lead to better diagnostic evaluation of ovarian pathology and facilitate the physiological manipulation of these processes to either promote or inhibit fertility. PUBLIC HEALTH RELEVANCE: The overall goal of the proposed study is to investigate how the LH surge induces the release of a mature egg from the ovary and corpus luteum formation, which are essential steps for female fertility. The current proposal focuses on determining the function of a small family of transcription factors, Core binding factors (CBFs) as key LH-induced mediators in these processes. Such knowledge can be applied for promoting and inhibiting these critical facets of ovarian physiology, thereby allowing us to better manage fertility, infertility, and ovarian-based disorders.

描述（由申请人提供）：排卵前促性腺激素激增启动卵泡成熟的最终过程，最终导致卵丘卵母细胞复合体（COC）的排卵和黄体（CL）的形成。卵泡成功成熟所需的促性腺激素激增所引发的一个重要步骤是特定转录因子的表达。然而，我们对 LH 诱导的关键转录因子的身份和调控作用的了解仍然非常有限。我们实验室和其他人的最新研究揭示了核转录因子核心结合因子（CBF）的一个小家族，它是参与排卵过程的关键转录调节因子。 CBF由两个亚基组成；由 Runx1、Runx2 和 Runx3 基因之一编码的 DNA 结合 α 亚基和非 DNA 结合 β 亚基 CBF2。为了发挥功能，RUNX 蛋白需要与 CBF2 形成二聚体。我们的初步数据显示，排卵期卵泡中 LH 激增可快速诱导 CBF 成分（Runx1、Runx2 和 CBF2）。使用体外模型，我们进一步证明 CBF（RUNX1/CBF2 和 RUNX2/CBF2）调节排卵期基因的表达，这些基因对于 COC 扩张、排卵和黄体化至关重要。此外，我们的初步研究表明，抑制 RUNX 活性可阻止 COC 体外扩增。基于这些新发现，我们假设 CBF 是 COC 成功扩增、排卵和黄体化所必需的关键转录调节因子。 小鼠 CBF 成分基因的纯合无效突变会导致早期致死，因此无法确定 CBF 在卵巢中的功能。证明 CBF 在体内的生理重要性的另一个挑战是 Runx1 和 Runx2 的重叠表达及其在排卵期卵泡中的功能冗余。为了解决这个问题，我们建议建立一种新型转基因小鼠模型，其中CBF在卵巢细胞中特异性失活。这将通过使用 Cre-lox 技术删除卵巢细胞中的 CBF2 来实现。由于 CBF2 是 RUNX1 和 RUNX2 的结合伴侣，因此 CBF2 的靶向删除会消除所有 CBF（RUNX1/CBF2 和 RUNX2/CBF2）的活性。使用这种转基因小鼠模型，我们将通过检查这种突变小鼠的卵巢表型来测试 CBF 的靶向失活导致 COC 扩增、排卵和黄体化缺陷的假设（具体目标＃1）。利用这种突变小鼠卵巢，我们将鉴定排卵期卵泡细胞中CBF下游的基因，并开始描绘排卵期卵泡发育最后阶段所需的转录调控途径（具体目标#2）。 这些研究将建立转基因动物模型，不仅可以定义 CBF 的体内卵巢功能，还可以识别控制排卵过程的转录调节机制。从该提案中获得的信息将为 COC 扩张、排卵和 CL 形成所涉及的机制提供新的见解。全面了解排卵期过程的细胞/分子机制将有助于更好地诊断评估卵巢病理学，并促进这些过程的生理操作，以促进或抑制生育能力。 公共健康相关性：拟议研究的总体目标是调查 LH 激增如何诱导卵巢释放成熟卵子和黄体形成，这是女性生育能力的重要步骤。目前的提案重点是确定一个转录因子小家族的功能，核心结合因子 (CBF) 作为这些过程中 LH 诱导的关键介质。这些知识可用于促进和抑制卵巢生理学的这些关键方面，从而使我们能够更好地管理生育、不孕和卵巢疾病。