Diversification of cell types during male and female external genital development

生殖泌尿发育分子解剖学项目（GUDMAP）的这项提案将使用小鼠模型来研究男性和女性外生殖器之间的差异如何在单细胞水平上产生。先天性外生殖器畸形（CAEG）是最常见的人类出生缺陷之一，影响约1：150的活产男婴。尿道下裂是一种CAEG，其特征在于阴茎腹侧的尿道异位开口。在严重的尿道下裂中，尿道板可以沿着阴茎的整个下侧打开，使其看起来像阴蒂。这种情况被称为模棱两可的生殖器。尿道下裂的遗传原因一直难以捉摸。全球流行导致对环境因素的审查增加，特别是可在动物模型中诱导生殖器异常的内分泌干扰物（EDCs）。胚胎生殖结节（GT）分化为阴茎或阴蒂的性别分化受性腺雄激素和雌激素的调节。性激素在GT发展中的中心作用产生了对EDC的敏感性;然而，介导EDC对GT影响的机制尚不清楚，特别是在细胞水平上。引起尿道下裂的内分泌干扰物作用于GT中的所有细胞，还是针对特定的细胞类型？如果尿道反应细胞的亚群控制尿道形成，那么缓冲这些细胞对抗EDC可能是预防尿道下裂的方法（类似于叶酸预防神经管缺陷的能力）。对于遗传性尿道下裂，识别细胞特异性基因功能可以区分尿道形成的直接调节因子和具有间接作用的基因（例如对内分泌干扰物的敏感性）。了解外生殖器发育中细胞类型的多样性对于确定CAEG的原因和制定预防策略至关重要。CAEG预防和治疗进展的一个主要障碍是缺乏对外生殖器组织中特定细胞类型的了解。该项目旨在通过使用单细胞RNA测序结合新的成像方式来填补这些关键的知识空白，以绘制正常雄性和雌性小鼠发育中的外生殖器的细胞类型多样性。

This proposal for the GenitoUrinary Development Molecular Anatomy Project (GUDMAP) will use the mouse model to investigate how differences between male and female external genitalia arise at the single cell level. Congenital malformations of the external genitalia (CAEG) are among the most prevalent human birth defects, affecting ~1:150 live male births. Hypospadias is a CAEG that is characterized by ectopic opening(s) of the urethra on the ventral side of the penis. In severe hypospadias, the urethral plate can open along the entire underside of the penis, giving it a clitoris-like appearance. This condition is termed ambiguous genitalia. Genetic causes of hypospadias have been elusive. The global prevalence has led to increased scrutiny of environmental factors, particularly endocrine disrupting chemicals (EDCs) that can induce genital anomalies in animal models. Sexual differentiation of the embryonic genital tubercle (GT) into a penis or a clitoris is regulated by gonadal androgens and estrogen. The central role of sex hormones in GT development creates sensitivity to EDCs; however, the mechanisms that mediate EDC effects on the GT are unknown, particularly at a cellular level. Do EDCs that cause hypospadias act on all cells in the GT, or do they target specific cell types? If subpopulation(s) of hormonally-responsive cells control urethragenesis, then buffering those cells against EDCs could be a method for prevention of hypospadias (analogous to folic acid’s ability to prevent neural tube defects). For genetic hypospadias, identifying cell-specific gene functions can distinguish direct regulators of urethragenesis from genes with indirect effects (e.g. sensitivity to EDCs). Understanding cell type diversity in developing external genitalia is essential for identifying the causes of CAEG and for developing preventative strategies. A major obstacle to progress in prevention and treatment of CAEG is the lack of knowledge of the specific cell types in external genital tissues. This project aims to fill these critical knowledge gaps by using single cell RNA-sequencing combined with novel imaging modalities to map cell type diversity in the developing external genitalia of normal male and female mice.