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.

项目概要 男性不育的一个主要原因是沃尔夫管发育缺陷和不典型。 产生男性内生殖道器官的胚胎结构。据了解 沃尔夫管分化主要由睾丸来源的作用驱动 雄激素。然而，雄激素信号如何协调沃尔夫管的过程 生殖领域的分化仍不清楚。沃尔夫管中雄激素的作用 分化是由雄激素受体 (AR) 介导的，该受体在 Wolffian 和 Wolffian 细胞中均表达 导管上皮和间质。使用新型间充质特异性 Ar 敲除小鼠 模型中，我们提供了第一个遗传证据，表明间充质中的 AR 对于 沃尔夫管分化。通过比较染色质可及性和转录组 来自雌性和雄性胚胎的沃尔夫管间充质，我们发现了一组雄激素- 诱导的染色质可及区域和新的雄激素诱导的间充质因子 R- Spondin 3 (Rspo3)。 RSPO3 是一种从间充质分泌的 WNT 信号激活剂 激活对沃尔夫管形态发生至关重要的上皮 Wnt 信号传导。虽然 间充质控制上皮分化，上皮具有相互诱导性 通过合成旁分泌生长因子 WNT9B 对间充质的影响。我们发现 Wnt9b 的缺失导致性分化窗口处的沃尔夫管变性 雄激素信号传导被认为可以促进沃尔夫管的存活。这些观察 导致并支持我们的中心假设：雄激素依赖性沃尔夫管分化 需要刺激上皮来源的 WNT9B，以及雄激素信号传导 沃尔夫管分化由间充质 AR 介导并由 雄激素通过上皮-间质相互作用诱导间充质因子 RSPO3。我们将 确定 WNT9B、AR、RSPO3 促进沃尔夫管的作用机制 通过利用组织特异性基因敲除、基因表达的组合进行分化 分析、荧光激活细胞分选、RNA-seq、ATAC-seq 和单细胞 mRNA-seq。增强现实 人类中的 WNT9B 变异与有缺陷的雄激素依赖性男性有关 生殖道分化。因此，完成我们的提案不仅会产生 雄激素依赖性沃尔夫管基本机制的基础知识 区分，而且还提供与我们对疾病的理解直接相关的知识 人类男性性发育。