Mechanisms of androgen-dependent Wolffian duct differentiation

雄激素依赖性沃尔夫管分化机制

• 批准号: 10633606
• 负责人: Fei Zhao
• 金额: $ 45.41万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-01 至 2028-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10633606
• 关键词: ATAC-seq; Address; Affect; Androgen Receptor; Androgens; Apoptosis; Cells; Chromatin; Connective Tissue; Development; Diagnosis; Disease; Ductal Epithelium; Embryo; Embryonic Structures; Ensure; Epididymis; Epithelium; Event; Female; Fluorescence-Activated Cell Sorting; Future Generations; Gene Expression; Gene Expression Profiling; Genetic; Growth Factor; Human; Impairment; Infertility; Knock-out; Knockout Mice; Knowledge; Lead; Maintenance; Male Infertility; Mediating; Mesenchymal; Mesenchyme; Metabolism; Molecular; Morphogenesis; Mutation; Organ; Pathology; Pattern; Physiology; Prevention strategy; Process; Proliferating; Reproduction; Role; Seminal Vesicles; Sex Differentiation; Sexual Development; Signal Transduction; Sorting; Sperm Transport; Structure of mesonephric duct; System; Testing; Testis; Tissues; Variant; WNT Signaling Pathway; androgen sensitive; knockout gene; mRNA sequencing; male; mouse model; paracrine; progenitor; programs; receptor expression; reproductive development; reproductive tract; sperm cell; transcriptome; transcriptome sequencing; transmission process

Project Summary A major cause of male infertility is defective and atypical development of the Wolffian duct, the embryonic structure that give rise to male internal reproductive tract organs. It is known that Wolffian duct differentiation is predominantly driven by the action of the testis-derived androgens. However, how the androgen signaling coordinates the process of the Wolffian duct differentiation remains unclear in the field of reproduction. The androgen action in Wolffian duct differentiation is mediated by the androgen receptor (AR), which is expressed in both Wolffian duct epithelium and mesenchyme. Using a new mesenchyme-specific Ar knockout mouse model, we provided the first genetic evidence that the AR in the mesenchyme is essential for Wolffian duct differentiation. By comparing chromatin accessibilities and transcriptomes of Wolffian duct mesenchymes from female and male embryos, we discovered a set of androgen- induced chromatin accessible regions and a new androgen-induced mesenchymal factor R- Spondin 3 (Rspo3). RSPO3 is a WNT signaling activator secreted from the mesenchyme to activate epithelial Wnt signaling that is essential for Wolffian duct morphogenesis. While the mesenchyme governs epithelial differentiation, the epithelium has the reciprocal inductive effects on the mesenchyme by synthesizing a paracrine growth factor WNT9B. We found that the loss of Wnt9b caused Wolffian duct degeneration at the sexual differentiation window when the androgen signaling was supposed to promote Wolffian duct survival. These observations lead to and support our central hypothesis: the androgen-dependent Wolffian duct differentiation requires the stimulation of the epithelium-derived WNT9B, and the androgen signaling in Wolffian duct differentiation is mediated by the mesenchymal AR and executed by the androgen-induced mesenchymal factor RSPO3 via epithelial-mesenchymal interactions. We will determine the mechanisms of WNT9B, AR, RSPO3 actions in promoting Wolffian duct differentiation by utilizing a combination of tissue-specific gene knockouts, gene expression assays, fluorescence-activated cell sorting, RNA-seq, ATAC-seq, and single cell mRNA-seq. AR and WNT9B variants in humans have been associated with defective androgen-dependent male reproductive tract differentiation. Therefore, the completion of our proposal will not only yield fundamental knowledge of basic mechanisms underlying androgen-dependent Wolffian duct differentiation but also provide knowledge directly relevant to our understanding of disorders of male sexual development in humans.

项目摘要 男性不育症的主要原因是沃尔夫管道的有缺陷和非典型发育， 产生男性内部生殖道器官的胚胎结构。众所周知 沃尔夫管道分化主要由睾丸衍生的作用驱动 雄激素。但是，雄激素信号如何协调沃尔夫管的过程 在繁殖领域，分化尚不清楚。沃尔夫管中的雄激素作用 分化是由雄激素受体（AR）介导的，该雄激素受体（AR）在两个狼队中都表达 管道上皮和间质。使用新的间充质特异性AR敲除鼠标 模型，我们提供了第一个遗传证据，表明间充质中的AR对于 沃尔夫管道分化。通过比较染色质的可访问性和转录组 来自雌性和雄性胚胎的沃尔夫（Wolffian）导管间充质，我们发现了一组雄激素 - 诱导染色质的区域和新的雄激素诱导的间充质因子R- Spondin 3（RSPO3）。 RSPO3是从间充质分泌的Wnt信号传导激活剂 激活上皮Wnt信号传导，这对于沃尔夫管形态发生至关重要。而 间充质控制上皮分化，上皮具有相互感应性 通过合成旁分泌生长因子Wnt9b对间充质的影响。我们发现 Wnt9b的损失在性别分化窗口中导致沃尔夫人管道变性 雄激素信号传导被认为促进了沃尔夫管的存活。这些观察 导致和支持我们的中心假设：依赖雄激素的狼管道分化 需要刺激上皮衍生的Wnt9b，并在 沃尔夫管道分化是由间充质AR介导的，并由 雄激素诱导的间充质因子RSPO3通过上皮间质相互作用。我们将 确定WNT9B，AR，RSPO3在促进Wolffian管道中的机制 通过利用组织特异性基因敲除，基因表达的结合来分化 测定，荧光激活的细胞分选，RNA-SEQ，ATAC-SEQ和单细胞mRNA-Seq。 ar 人类中的Wnt9b变体与雄激素依赖性雄性有缺陷有关 生殖道分化。因此，我们的提案的完成不仅将产生 依赖雄激素依赖雄激素的基本机制的基本知识 差异化，但也提供与我们对疾病的理解直接相关的知识 男性性发展在人类中。