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过程中的转录调控，特别是有丝分裂后阶段，以及从产前到出生后卵巢的过渡，此时卵母细胞簇开始形成原始卵泡。卵泡形成的早期阶段，首先是生殖细胞囊破裂、原始卵泡的形成和向初级和次级卵泡的过渡，是决定生殖寿命和生育力的关键。在这些卵母细胞发育的早期阶段，会发生大量的卵母细胞丢失，未知的促生存因素可以稳定这种丢失，如减数分裂前期I期卵母细胞停滞和原始卵泡的形成。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育所必需和必不可少的。我们发现LIM同源结构域基因LhxB在卵母细胞中优先表达，在卵母细胞分化中起关键作用。此外，在LhxB缺乏的卵母细胞中，Kit等多种基因在卵母细胞中优先表达下调。在特定的目标1中，我们将检验LhxB直接调控许多卵母细胞特异性基因的假设，包括卵母细胞特异性转录调节因子。我们推测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，我们还将研究LhxB缺乏对胚胎性腺转录组的影响，并确定分子病理学的发生。在特定的目标2中，我们将检验LHX8蛋白中的两个LIM结构域与其他卵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是第一个卵母细胞特异的LIM同源结构域基因，在早期卵子发生过程中具有关键功能。我们提出的研究将提供对LhxB作用机制的洞察，并进一步阐明控制卵子发生的遗传途径。LhxB途径或其辅助调节因子可能在人类卵巢早衰中起作用。

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.