REGULATION OF GNRH RECEPTOR GENE EXPRESSION

GNRH受体基因表达的调控

• 批准号: 6553567
• 负责人: Colin M Clay
• 金额: $ 10.2万
• 依托单位: COLORADO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-01-01 至 2004-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6553567
• 关键词: biological signal transduction; cell line; gene expression; genetic promoter element; genetically modified animals; gonadotropin releasing factor; hormone receptor; hormone regulation /control mechanism; laboratory mouse; protein structure function; tissue /cell culture

The binding of gonadotropin-releasing hormone (GnRH) to specific, high-affinity receptors located on gonadotrope cells of the anterior pituitary gland is central to reproduction. In the absence of GnRH input, synthesis and secretion of luteinzing hormone and, consequently, normal gonadal function ceases. Thus, the GnRH receptor (GnRHR) is the site that receives and mediates the primary stimulatory input to gonadotropes. We have found that expression of the murine GnRHR in gonadotrope-derived alphaT3-1 cells is mediated by a complex enhancer whose components include a binding site for steroidogenic factor-1 (SF- 1), an AP-1 element, and an element we have termed the GnRH receptor activating sequence (GRAS). This complex enhancer also integrates multiple endocrine inputs. First, we have recently found that GRAS co-localizes with activin regulation of the GnRHR promoter. Unresolved, however, is the identity of the protein(s) that integrate functional activity at GRAS. In Specific Aim 1, we propose to identify the protein(s) that regulate the functional activity, and activin responsiveness of GRAS. Second, AP-1 appears to be the operative element that mediates GnRH regulation; however, important questions remain as to the signal transduction cascades and downstream targets that ultimately lead to GnRH activation at the GnRHR AP-1 site. In Specific Aim 2, our goal is to define the molecular mechanisms underlying GnRH regulation of GnRHR gene expression. We have also found that 1900 bp of proximal promoter is sufficient for tissue-specific expression and GnRH responsiveness in transgenic mice. In Specific Aim 3, we propose to expand these studies to further explore the requirements for tissue/cell-specific expression and hormonal regulation of the GnRHR gene. Finally, we have generated cell lines that express intrinsically fluorescent forms of the GnRHR. These molecules provide a unique opportunity to study the GnRHR as both an occupied and unoccupied receptor in living cells. In Specific Aim 4, we will use fluorescence resonance energy transfer to test the hypothesis that an early event in GnRH signaling is agonist induced receptor self- association. In terms of fertility regulation, the relevance of investigating GnRH and its cognate receptor is clear. However, the use of potent agonists and antagonists of GnRH in the treatment of fibroid tumors, endometriosis, and carcinomas of the breast, prostate, testes, and pituitary underscores the need for a full understanding of GnRH and the GnRHR in both health and disease.

促性腺激素释放激素（GNRH）与位于垂体前腺体促性腺细胞上的特异性高亲和力受体的结合是繁殖的核心。 在没有GNRH输入的情况下，曲曲霉激素的合成和分泌，因此，正常的性腺功能停止。 因此，GNRH受体（GNRHR）是接收并介导促性腺激素的主要刺激输入的部位。 我们发现，鼠gnRHR在性腺衍生的Alphat3-1细胞中的表达是由复杂增强剂介导的，其组分包括类固醇生成因子-1（SF-1），AP-1元素，AP-1元素和我们称为GNRH受体激活序列（GRAS）的结合位点。 该复合物增强子还集成了多个内分泌输入。 首先，我们最近发现，GRA与GNRHR启动子的激活素调节共定位。 然而，尚未解决的是在GRAS上整合功能活性的蛋白质的身份。 在特定目标1中，我们建议鉴定调节功能活性的蛋白质和GRA的激活素反应性。 其次，AP-1似乎是介导GNRH调节的手术元素。但是，关于最终导致GNRHR AP-1位点的GNRH激活的信号转导级联和下游目标仍然存在重要问题。 在特定目标2中，我们的目标是定义GnRHR基因表达的GNRH调节的分子机制。 我们还发现，1900 bp的近端启动子足以使转基因小鼠的组织特异性表达和GNRH反应能力。 在特定目标3中，我们建议扩展这些研究，以进一步探索GNRHR基因组织/细胞特异性表达和激素调节的要求。 最后，我们产生了细胞系，该细胞系表达了GNRHR的内在荧光形式。 这些分子为研究GNRHR作为活细胞中的被占用和未占用的受体提供了独特的机会。 在特定目标4中，我们将使用荧光共振能量转移来检验以下假设：GnRH信号传导中的早期事件是激动剂诱导的受体自我缔合。 在生育率调节方面，研究GNRH及其同源受体的相关性很明显。 然而，使用有效的激动剂和GNRH的拮抗剂在治疗乳腺癌，子宫内膜异位症和乳腺癌，前列腺，睾丸和垂体的癌中强调了对健康和疾病中GNRH和GNRHR的全面了解。