Transcriptional networks coordinating luminal environment in the human epididymis: the role of the androgen receptor.

雄激素是类固醇激素，在决定男性特征方面具有关键作用， 与雄激素受体（AR）结合以指导独特的转录程序。虽然最好的研究是在 在人类前列腺和前列腺癌中，AR转录网络还控制其他细胞的正常功能， 男性生殖道组织，包括附睾和输精管。一个关键的功能， 附睾是男性生殖管的近端部分，将睾丸产物输送出体外， 精子成熟离开睾丸的未成熟精子在发育过程中获得使卵子受精的能力。 穿过附睾的头部和身体的通道。许多因素有助于建立和维持 正常精子成熟所需的特殊附睾腔环境。这些包括 激素和其他睾丸产物进入导管，尽管附睾内衬的上皮细胞具有 关键角色近端腔液的pH值和碳酸氢盐浓度较低，以保持精子活力 以及相对高的钾浓度以促进精子静止。多离子通道，离子 附睾顶膜或基底膜中的交换体、转运体和水通道 上皮细胞参与对腔液的微调。基于a）关于AR的大量初步数据 人附睾上皮（HEE）细胞中的转录网络，这表明该TF的新辅因子 参与附睾，连同B）创新的HEE类器官模型，我们的目标是测试 雄激素受体是建立和维持正常功能的关键的总体假设 人类附睾上皮细胞。此外，通过招募特定的辅助因子，AR 在这个组织中协调了一个独特的转录程序。在第一个具体目标中，我们将确定 雄激素影响人类附睾分化和功能的机制 上皮我们将使用一种新的附睾类器官模型与单细胞RNA一起进行功能研究， 测序（scRNA-seq）。这些实验将提供新的和重要的见解的作用 人类附睾上皮生物学中的雄激素和协调的转录网络 其功能。第二个目标的研究将确定AR与其他共刺激因子相互作用的机制。 包括RUNX 1和CEBPβ在内的因子以及这些因子如何整合到转录网络中， 人附睾上皮我们还将研究AR、RUNX 1和 使用全基因组和基因座特异性测定的CEBPβ。一个特别感兴趣的基因是囊性纤维化 跨膜传导调节因子（CFTR），其建立并维持氯化物和碳酸氢盐 附睾腔内的梯度。CFTR在HEE细胞中招募了一组独特的顺式调节元件，其中一些是 它也结合AR。这些数据可能有助于确定男性避孕药的新靶点， 也对恢复男性附睾疾病生育能力的治疗方法产生深远影响。

SUMMARY: Androgens are steroid hormones with a pivotal role in determining male characteristics, which bind to the androgen receptor (AR) to direct a unique transcriptional program. Although best studied in the human prostate and in prostate cancer, the AR transcriptional network also controls normal functions of other male reproductive tract tissues, including the epididymis and the vas deferens. A critical function of the epididymis, the proximal part of the male genital ducts that transport testicular products out of the body, is in sperm maturation. Immature sperm leaving the testis acquire the capacity to fertilize an egg during their passage through the head and body of the epididymis. Many factors contribute to establishing and maintaining the specific epididymis luminal environment that is required for normal sperm maturation. These include hormones and other testicular products that enter the duct, though the epithelium lining the epididymis has a pivotal role. The proximal luminal fluid has a low pH and bicarbonate concentration to preserve sperm motility and relatively high potassium concentrations to facilitate sperm quiescence. Multiple ion channels, ion exchangers, transporters and water channels in the apical or basolateral membranes of the epididymis epithelium participate in fine-tuning the luminal fluid. Building upon a) substantial preliminary data on the AR transcriptional network in human epididymis epithelial (HEE) cells, which suggest novel co-factors for this TF are involved in the epididymis, together with b) an innovative HEE organoid model, our goal is to test the overarching hypothesis that the Androgen Receptor is pivotal to establishing and maintaining normal functions of the human epididymis epithelium. Moreover, that through the recruitment of specific co-factors, AR orchestrates a unique transcriptional program in this tissue. In the first specific aim we will determine the mechanisms through which androgens impact the differentiation and function of the human epididymis epithelium. We will use a novel epididymis organoid model for functional studies together with single cell RNA- sequencing (scRNA-seq). These experiments will provide novel and important insights into the role of androgens in the biology of the human epididymis epithelium and the transcriptional network that coordinates its functions. Studies in the second aim will determine the mechanisms of interaction of AR with other co- factors including RUNX1 and CEBPβ and how these factors integrate into the transcriptional network in the human epididymis epithelium. We will also investigate the hierarchy of dependency of AR, RUNX1 and CEBPβ using genome wide and locus-specific assays. A gene of particular interest is the cystic fibrosis transmembrane conductance regulator (CFTR), which establishes and maintains chloride and bicarbonate gradients in the epididymis lumen. CFTR recruits a unique set of cis-regulatory elements in HEE cells, some of which also bind AR. These data may facilitate the identification of novel targets for male contraceptives and also have a profound impact on therapeutic approaches to restore male fertility in epididymis disease.