In the mammalian auditory system, frequency discrimination depends on numerous morphological and physiological properties of the organ of Corti, which gradually change along the apex-to-base (tonotopic) axis of the organ. For example, the basilar membrane stiffness changes tonotopically, thus affecting the tuning properties of individual hair cells. At the molecular level, those frequency-specific characteristics are mirrored by gene expression gradients; however, the molecular mechanisms controlling tonotopic gene expression in the mouse cochlea remain elusive. Through analyzing single-cell RNA sequencing (scRNA-seq) data from E12.5 and E14.5 time points, we predicted that morphogens, rather than a cell division-associated mechanism, confer spatial identity in the extending cochlea. Subsequently, we reconstructed the developing cochlea in 3D space from scRNA-seq data to investigate the molecular pathways mediating positional information. The retinoic acid (RA) and hedgehog pathways were found to form opposing apex-to-base gradients, and functional interrogation using mouse cochlear explants suggested that both pathways jointly specify the longitudinal axis.

在哺乳动物听觉系统中，频率辨别取决于柯蒂氏器的众多形态和生理特性，这些特性沿着器官的顶端到基部（音位）轴逐渐变化。例如，基底膜硬度发生音调变化，从而影响单个毛细胞的调谐特性。在分子水平上，这些频率特异性特征由基因表达梯度反映；然而，控制小鼠耳蜗音调基因表达的分子机制仍然难以捉摸。通过分析 E12.5 和 E14.5 时间点的单细胞 RNA 测序 (scRNA-seq) 数据，我们预测形态发生素（而不是细胞分裂相关机制）在延伸的耳蜗中赋予空间同一性。随后，我们根据 scRNA-seq 数据在 3D 空间中重建了发育中的耳蜗，以研究介导位置信息的分子途径。视黄酸（RA）和刺猬通路被发现形成相反的顶端到基底梯度，并且使用小鼠耳蜗外植体进行的功能询问表明这两条通路共同指定纵轴。