LIM Homeodomain Regulated Genetic Pathways in Oogenesis and Ovarian Failure

我们感兴趣的是了解早期卵子发生的转录调控，特别是有丝分裂后阶段，以及从产前到产后卵巢的过渡，当卵母细胞群开始形成原始卵泡时。卵巢卵泡形成的早期阶段，从生殖细胞囊肿的破裂、原始卵泡的形成和向初级和次级卵泡的过渡开始，是决定生殖寿命和生育能力的关键。大量卵母细胞的丢失发生在卵母细胞发育的早期阶段，未知的促存活因子稳定了这种丢失，如卵母细胞在减数分裂前期停止，原始卵泡形成。在卵泡形成的早期阶段，许多生殖细胞特异性基因的转录是卵泡发育和早期胚胎发生所必需的。我们发现LIM同源结构域基因LhxB在卵母细胞中优先表达，并在卵母细胞分化中起关键作用。此外，在LhxB缺陷卵母细胞中，Kit等多个优先表达于卵母细胞的基因下调。在Specific Aim 1中，我们将验证LhxB直接调控包括卵母细胞特异性转录调控因子在内的许多卵母细胞特异性基因的假设。我们假设LHX8直接调节Kit的表达，Kit是原始卵泡形成的重要调节因子。我们还将使用抗LHX8抗体进行染色质免疫沉淀，以确定LHX8直接结合的基因组区域。此外，我们将研究LhxB缺乏对胚胎性腺转录组的影响，并确定分子病理学的发病。在Specific Aim 2中，我们将验证LHX8蛋白中的两个LIM结构域与其他卵母细胞特异性转录调节因子以及LIM结构域结合蛋白相互作用的假设。LhxB是首个卵母细胞特异性LIM同源结构域基因，在卵发生早期具有关键功能。我们提出的研究将为LhxB的作用机制提供深入的见解，并进一步阐明控制卵子发生的遗传途径。LhxB通路或其协同调节因子可能在人类卵巢早衰中发挥作用。

We are interested in understanding transcriptional regulation during early oogenesis, specifically the post-mitotic stages, and the transition from prenatal to postnatal ovary, when clusters of oocytes begin to form into primordial follicles. Early stages of ovarian follicle formation, beginning with the breakdown of germ cell cysts, formation of primordial follicles and transition to primary and secondary follicles, are critical in qetermining the reproductive life span and fertility. Large oocyte loses occur during these early stages of oocyte development, and unknown pro-survival factors stabilize such loss as oocytes arrest in meiotic prophase I, and primordial follicles form. Transcription of numerous germ cell specific genes, necessary and essential for follicular development and early embryogenesis, is initiated during these early stages of follicle formation. We discovered that LIM homeodomain gene, LhxB, is preferentially expressed in oocytes and critical in oocyte differentiation. In addition, multiple genes preferentially expressed in the oocyte, such as Kit are down-regulated in LhxB deficient oocytes. In Specific Aim 1, we will test the hypothesis that LhxB directly regulates numerous oocyte-specific genes including oocyte-specific transcriptional regulators. We hypothesize that LHX8 directly regulates expression of Kit, an essential regulator of primordial follicle formation. We will also perform chromatin immunoprecipitation with anti-LHX8 antibodies to determine genomic regions that LHX8 directly binds. In addition, we will study the effects of LhxB deficiency on the embryonic gonadal transcriptome and determine the onset of molecular pathology. In Specific Aim 2 we will test the hypothesis that the two LIM domains in the LHX8 protein interact with other oocytespecific transcriptional regulators as well as LIM domain binding proteins. LhxB represents the first oocyte-specific LIM homeodomain gene with critical functions during early oogenesis. Our proposed studies will provide insight into the mechanisms of LhxB action and further elucidate genetic pathways that govern oogenesis. It is likely that LhxB pathway or its co-regulators playa role in human premature ovarian failure.