Hair Cell Development in the Mammalian Cochlea

听觉和前庭功能依赖于功能性内耳的形成。虽然这两个系统都有多个组成部分，但本实验室专注于感觉上皮的发育，其中包含机械感觉毛细胞和相关细胞，称为支持细胞，以及来自VIIIth(听前庭)脑神经的神经元对这些毛细胞的神经支配。这三种细胞类型都来自耳囊，耳囊是一种胎盘结构，在发育早期形成于后脑附近。确定指定每一种细胞类型的因素，然后将它们组装成功能单位是发育神经科学科的一个关键目标。在过去的一年里，实验室的不同成员检查了这些发育过程的几个不同方面。 从2020年3月开始的强制关闭严重影响了实验室的整体活动。然而，我们能够公布一个长期项目的结果，该项目试图使用单细胞RNA测序来表征发育中的耳蜗内多个时间点的所有细胞类型。在这项研究中，我们从四个不同的发育时间点--E14、E16、P0和P7--收集了30,000多个细胞。随后对这些细胞的分析揭示了新的细胞类型，并使我们能够生成已知细胞(如毛细胞)和新细胞类型(如位于Kollikers器官中的细胞)的发育轨迹，Kollikers器官是一种瞬时发育结构，与耳蜗几个独特方面的生成有关。我们还发现了一条新的信号通路，由转化生长因子β调控，对Corti器官内两种独特的细胞类型--外毛细胞和Deiters细胞的发育至关重要。该项目正在进行的工作已经开始产生更全面地描述转化生长因子β作用所需的条件小鼠突变。 根据NIH基因组数据共享政策，我们将我们所有的单细胞数据存放在两个存储库中：GEO和Gear门户，这是一个旨在共享内耳单细胞数据的在线网站。Gear跟踪数据的使用情况，我们非常高兴地看到该数据集的广泛使用。 我们还发表了第二项单细胞RNA-Seq研究，其中我们比较了出生后早期和成年后的支持细胞。结果表明，随着支持细胞的发育，基因表达发生了变化，但也表明，在新生儿和成年耳蜗中都存在两种独特的支持细胞类型。这些数据也同时存放在GEO和GEAR中。 作为正在进行的合作的一部分，我们为研究NAD+补充对Cockayne综合征小鼠模型毛细胞存活的影响的项目做出了贡献，Cockayne综合征是一种早衰的早衰模型，并研究了YAP在哺乳动物椭圆体毛细胞再生中的作用。

Auditory and vestibular function are dependent of the formation of a functional inner ear. While there are multiple components for both of these systems, this laboratory focuses on the development of the sensory epithelia, which contain mechanosensory hair cells and associated cells called supporting cells and on the innervation of those hair cells by neurons from the VIIIth (acousticovestibular) cranial nerve. All three of these cell types are derived from the otocyst, a placodal structure that forms adjacent to the hindbrain early in development. Identifying the factors that specify each of these cell types and then direct their assembly into functional units is a key goal of the Section on Developmental Neuroscience. During the previous year, different members of the laboratory have examined several different aspects of these developmental processes. Overall activity in the laboratory was significantly affected by the mandatory shutdown beginning in March, 2020. However, we were able to publish the results of a long-term project which sought to characterize all of the cell types within the developing cochlea at multiple time points using single cell RNA-sequencing. In this study, we collected just over 30,000 cells from four different developmental time points, E14, E16, P0 and P7. Subsequent analysis of these cells revealed novel cell types and allowed us to generate developmental trajectories for both known cells, such as hair cells, and novel cell types, such as those located in Kollikers organ, a transient developmental structure that has been implicated in the generation of several unique aspects of the cochlea. We also identified a novel signaling pathway, regulated by Transforming Growth Factor Beta, as important for the development of two unique cell types within the organ of Corti, outer hair cells and Deiters cells. Ongoing work on this project has begun to generate the conditional mouse mutants required to more completely characterize the role of TGF Beta. In compliance with the NIH Genome Data Sharing Policy, we deposited all of our single cell data in two repository, GEO and the gEAR Portal, an online site designed and maintained for the purpose of sharing inner ear single cell data. gEAR tracks data usage and we have been very happy to see extensive use of this data set. We also published a second single cell RNA-Seq study in which we compared early post-natal and adult supporting cells. Results indicated changes in gene expression as supporting cells develop but also showed that two unique supporting cell types are present in both newborn and adult cochleae. These data were also deposited in both GEO and gEAR. As part of ongoing collaborations, we contributed to projects examining the effects of NAD+ supplementation on hair cell survival in mouse models of Cockayne syndrome, a progeria-like model of premature aging, and examined the role of YAP in hair cell regeneration in the mammalian utricle.