Single Cell Transcriptional Characterization of the Developing Organ of Corti Identifies Etv4/5/1 as Regulators of Lateral Cell Identity

The mammalian auditory sensory epithelium, the organ of Corti, is comprised of a complex and highly ordered cellular mosaic that includes multiple unique cell types. The small number of each of these cell types and their close association with one another has limited efforts to transcriptionally characterize the development of these cells. To provide a better understanding of the development events that regulate cell type specification, we dissociated developing cochlear ducts at time points between embryonic day 11 and 16 and collected duct epithelial cells for single cell RNA-sequencing. From the resulting data we were able to build a developmental trajectory from undifferentiated precursor cells to each of the major cell types in the organ of Corti. Bioinformatic analyses including SCENIC, TradeSeq and CellOracle were then used to identify transcription factors and other genes that may regulate different phases of cell type development. To validate the analysis, the role of one set of candidate transcription factors, Etv4/5/1 was examined by generating triple knock out mice. As predicted from the bioinformatic analysis, results demonstrated key roles for Etvs in the development of inner and outer pillar cells. and accurately identified down-stream Etv targets. This data set will provide significant insights regarding the specification of unique cochlear cell types which will have implications for understanding congenital deficits and potential applications in the development of regenerative strategies.