GnRH-Regulated Transcriptome in the Gonadotrope

描述(申请人提供)：调节促性腺激素中的基因转录是决定促性腺激素水平的关键调节步骤，因此，调节性腺功能。两性的配子发生和类固醇的发生都受这两种激素的控制，即来自垂体的卵泡刺激素(FSH)和黄体生成素(LH)，而这两种激素又受来自下丘脑的促性腺激素释放激素(GnRH)的调节。GnRH通过诱导三个即刻早期基因来调节促性腺激素的合成：ATF3诱导？糖蛋白亚基Egr-1诱导黄体生成素？而AP-1诱导FSH？亚单位。这项应用的总体目标是确定促性腺激素细胞中由这些即刻早期基因组成的GnRH调节的转录本。我们提出了两个互补的目标，从更全面的角度剖析转录调控。在第一个目标中，我们将通过芯片来确定这些即刻早期基因中除促性腺激素亚基之外的直接靶基因的补充性，这将使我们能够描绘转录调控网络。已经通过表达阵列分析确定的GnRH诱导的基因与通过芯片确定的这三个最高诱导转录因子的直接基因靶点的交集，将集中在最具功能的结合元件上。此外，这些数据集的重叠将识别其转录可能受到类似于促性腺激素的调控的基因。在第二个目标中，使用来自促性腺激素来源的细胞的双杂交筛选，我们将识别与ATF3、Egr-1或AP-1相互作用的分子，特别强调辅助激活因子和辅助抑制因子。这些辅因子可能在促性腺激素三个亚基的差异表达中起作用。此外，我们将确定从第一个目的中选择的目标基因是否在其启动子中具有相似的结合位点和/或需要相同的辅助因子，这可能有助于调节它们的表达。这种方法将有助于我们从更广泛的角度三角测量促性腺激素亚单位的差异调控，以及它是否通过在各自的启动子上形成不同的复合体来完成，此外还将分析促性腺激素基因转录的更全面的调控。这一提议的结果也将为研究染色质重塑和基础转录机制的激活开辟新的途径。与公共卫生相关：哺乳动物的生育力受不同激素信号的整合调节，这种信号发生在脑下垂体促性腺激素细胞群的水平上。这种整合最终导致了促性腺激素细胞中基因表达的精确调控，从而导致了控制生殖的激素的微妙平衡。因此，了解促性腺激素基因表达的分子机制将有助于深入了解生殖系统的生理学和病理生理学。

DESCRIPTION (provided by applicant): Regulation of gene transcription in the gonadotrope is a key regulatory step that determines the levels of gonadotropin hormones and therefore, regulates gonadal function. Gametogenesis and steroidogenesis in both sexes are controlled by these two hormones, follicle-stimulating hormone (FSH) and luteinizing hormone (LH) from the pituitary, which are, in turn, regulated by gonadotropin-releasing hormone (GnRH) from the hypothalamus. GnRH regulates gonadotropin synthesis through induction of the three immediate early genes: ATF3 induces the ? glycoprotein subunit, Egr-1 induces the LH? subunit, and AP-1 induces the FSH? subunit. The overall goal of this application is to determine the GnRH-regulated transcriptomes comprised of these immediate early genes in the gonadotrope cell. We propose two complimentary aims to dissect transcriptional regulation from a more comprehensive standpoint. In the first aim, we will determine the complement of direct target genes, in addition to gonadotropin subunits, of these immediate early genes by ChIP-chip, which will allow us to delineate the transcriptional regulatory network. The intersection of genes that are induced by GnRH, already identified by expression array analysis, and the direct gene targets of these three most highly induced transcription factors determined herein by ChIP-chip, will focus our analysis on the most functional binding elements. Furthermore, overlap of these data sets will identify the genes whose transcription may be regulated similarly to gonadotropins. In the second aim, using a two-hybrid screen of a cDNA library from gonadotrope-derived cells, we will identify molecules that interact with ATF3, Egr-1 or AP-1, with a special emphasis on coactivators and corepressors. These cofactors may contribute to differential expression of the three gonadotropin subunits. Additionally, we will determine whether selected target genes from the first aim have similar sets of binding sites in their promoters and/or require the same cofactors, which may contribute to regulation of their expression. This approach will help us triangulate, from a broad perspective, the differential regulation of the gonadotropin subunits and whether it is accomplished by formation of diverse complexes on their respective promoters, in addition to analyzing the more global regulation of gene transcription in the gonadotrope. The results from this proposal will also open new avenues to study chromatin remodeling and activation of basal transcriptional machinery. PUBLIC HEALTH RELEVANCE: Fertility in mammals is regulated by integration of different hormonal signals, which occurs at the level of the gonadotrope cell population in the pituitary gland. The integration ultimately results in the precisely orchestrated modulation of gene expression in the gonadotrope cell that leads to delicate balance of hormones that control reproduction. Thus, understanding the molecular mechanisms governing gonadotrope gene expression will lead to insight into the physiology and pathophysiology of the reproductive system.