Combinatorial Signals & Gene Expression in Trophoblasts

描述（由申请人提供）：绒毛膜促性腺激素（CG）是一种关键的 灵长类动物发育中的孕体分泌的促黄体激素。内分泌 促进CG亚基最大合成的机制是重要的 成功怀孕的决定因素。初步证据显示， CG α亚基基因以胎盘特异性方式协同诱导 通过由几个顺式元件（两个 克雷斯和JRE），这些是整合多种细胞内信号所必需的。 CRE结合复合物由几种激活剂（CREB，AP-1，CBP）和 辅阻遏物分子（HDAC 2）。JRE是同源框结合位点， 与无远端3（D1 x3）相互作用。该提案详细介绍了一系列复杂的 实验检查的假设，转录激活因子和 阻遏物有助于组合的细胞特异性调节的 α亚基基因此外，滋养层分化取决于基因 由Dlx 3引入的程序。总体目标是全面了解 CG合成机制及调控因素分析 滋养层分化目标1：继续分析 控制CRE依赖性α亚单位基因调控的机制 滋养层CRE依赖性α亚基基因调控及 在原代人滋养层中检测EGF和cAMP。生化 和诱变研究探讨了CRE中辅助调节因子的作用， 结合复合物，并定义了α-羟色胺内染色质重塑的程度。 亚基启动子目的2：探讨Dlx 3基因在细胞凋亡调控中的作用。 α亚基基因研究将分析灵长类动物中Dlx 3的表达模式 胎盘和体外滋养层分化期间 免疫细胞化学生物化学和诱变研究将检查 α亚单位内CRE和JRE结合复合物之间的相互作用 启动子Dlx 3的相互作用伴侣，有助于α亚基基因 调节和滋养层分化将被确定。目标3： 研究Dlx 3在胎盘发育和分化中的作用。Dlx3 敲除小鼠将用于鉴定Dlx 3在鼠中诱导的基因程序 用最先进的基因图谱分析来检测滋养层细胞。以补充 这些研究，调查调解所需的因素， Dlx 3基因启动子的转录激活，这将导致 滋养层分化“上游”效应物的鉴定将 进行。

DESCRIPTION (provided by applicant): Chorionic gonadotropin (CG) is a key luteotropic hormone secreted by the developing conceptus in primates. Endocrine mechanisms contributing to maximal CG subunit synthesis are important determinants of a successful pregnancy. Preliminary evidence suggests that the CG alpha subunit gene is synergistically induced in a placental-specific manner via a complex transcriptional "unit" comprised of several cis elements (two CREs and the JRE) necessary for integration of multiple intracellular signals. The CRE binding complex consists of several activator (CREB, AP-1, CBP) and corepressor molecules (HDAC2). The JRE is a homeobox-binding site that interacts with Distal-less 3 (D1x3). This proposal details a complex series of experiments examining the hypothesis that transcriptional activators and repressors contribute combinatorially to the cell-specific regulation of the alpha subunit gene. Further, trophoblast differentiation depends upon gene programs induced by Dlx3. The overall goal is a comprehensive understanding of mechanisms governing CG synthesis and analysis of factors regulating trophoblast differentiation. The aims are: Aim 1: Continued analysis of the mechanisms that govern CRE-dependent alpha subunit gene regulation in trophoblasts. CRE-dependent alpha subunit gene regulation and the effects of the EGF and cAMP will be examined in primary human trophoblasts. Biochemical and mutagenesis studies investigate the role of coregulators within the CRE binding complex and defines the extent of chromatin remodeling within the alpha subunit promoter. Aim 2: Explore the role of Dlx3 in the regulation of the alpha subunit gene. Studies will analyze the Dlx3 expression pattern in primate placenta and during trophoblast differentiation in vitro by immunocytochemistry. Biochemical and mutagenesis studies will examine interactions between CRE and JRE binding complexes within the alpha subunit promoter. Interacting partners of Dlx3 that contribute to alpha subunit gene regulation and to trophoblast differentiation will be identified. Aim 3: Examine the role of Dlx3 in placental development and differentiation. The Dlx3 knockout mouse will be used to identify gene programs induced by Dlx3 in murine trophoblasts using state-of-the-art gene profiling analysis. To complement these studies, an investigation of the factors required to mediate transcriptional activation of the Dlx3 gene promoter that will lead to identification of "upstream" effectors of trophoblast differentiation will be conducted.