DAX1 in Testis Determination and Function

DAX1 在睾丸测定和功能中的作用

• 批准号: 6675743
• 负责人: James Larry JAMESON
• 金额: $ 31.36万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-11 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6675743
• 关键词: clinical research; developmental genetics; functional /structural genomics; gene expression; gene interaction; gene targeting; immunoprecipitation; in situ hybridization; laboratory mouse; microarray technology; nuclear receptors; point mutation; protein protein interaction; protein structure function; reproductive development; sex determination; sex differentiation; testis; transfection; yeast two hybrid system

DESCRIPTION (provided by applicant): Over the last 8 years, we identified and characterized patients with naturally occurring mutations in the orphan nuclear receptors, DAX1 and SF1, and demonstrated that DAX1 acts in part by inhibiting SF1- mediated transcription. Using targeted mutagenesis, we developed a murine Dax1 knockout (KO) model and identified roles for Dax1 in gonadal determination, testis development, and adult testis function. In this grant, we propose to extend these studies, which have provided unexpected insight into mechanisms of gonadal development. We propose 3 inter-related aims that focus on Dax1 structure and function as a transcriptional repressor, identify genetic pathways regulated by Dax1, and develop animal models to unravel the interplay of Dax1 with other genes involved in testis development and function. Specifically, in Aim 1 we will identify molecular partners that mediate DAX1 transcriptional repression. Naturally occurring DAX1 mutations will be identified and characterized to elucidate key structural domains required for DAX1 function in vivo. Candidate proteins identified in a yeast two-hybrid screen of an embryonic gonadal library will be further characterized. The goal of Aim 2 is to identify the genetic pathways regulated by Dax1 using microarray analyses of genes expressed in wild type versus Dax1-deficient embryonic gonads at 12 dpc, a timepoint when Dax1 expression diverges in males and females. Aim 3 is designed to explore the interaction of Dax1 with other genetic pathways in vivo. Using mice that lack Dax1, we will explore the gonadal phenotypes of mice with selective rescue of Dax1 in various cell types. In addition, Dax1-deficient mice will be crossed to other strains with alterations in genetic pathways proposed to intersect with Dax1 (e.g., Sf1, Sry, Ptc). Success in these aims should provide a critical link between DAX1 and transcriptional repression pathways that link a variety of other transcription factors involved in gonadal development.

描述（由申请人提供）：在过去的 8 年里，我们鉴定并表征了孤儿核受体 DAX1 和 SF1 天然发生突变的患者，并证明 DAX1 部分通过抑制 SF1 介导的转录发挥作用。通过定向诱变，我们开发了小鼠 Dax1 敲除 (KO) 模型，并确定了 Dax1 在性腺决定、睾丸发育和成人睾丸功能中的作用。在这笔赠款中，我们建议扩展这些研究，这些研究为性腺发育机制提供了意想不到的见解。我们提出了 3 个相互关联的目标，重点关注 Dax1 作为转录抑制因子的结构和功能，识别 Dax1 调节的遗传途径，并开发动物模型来揭示 Dax1 与参与睾丸发育和功能的其他基因的相互作用。具体来说，在目标 1 中，我们将鉴定介导 DAX1 转录抑制的分子伙伴。将鉴定和表征自然发生的 DAX1 突变，以阐明 DAX1 体内功能所需的关键结构域。将进一步表征在胚胎性腺文库的酵母二杂交筛选中鉴定的候选蛋白。目标 2 的目标是通过对 12 dpc（Dax1 表达在雄性和雌性中出现差异的时间点）野生型与 Dax1 缺陷胚胎性腺中表达的基因进行微阵列分析来确定受 Dax1 调节的遗传途径。目标 3 旨在探索 Dax1 与体内其他遗传途径的相互作用。使用缺乏 Dax1 的小鼠，我们将通过在各种细胞类型中选择性拯救 Dax1 来探索小鼠的性腺表型。此外，Dax1缺陷小鼠将与其他品系杂交，这些品系的遗传途径发生改变，与Dax1相交（例如Sf1、Sry、Ptc）。这些目标的成功应该在 DAX1 和转录抑制途径之间提供关键联系，这些途径将性腺发育中涉及的各种其他转录因子联系起来。