Unraveling Cochlear Otic Mesenchyme Cells: The Role of POU3F4 During Cochlear Development

Abstract

The cochlea consists of diverse cellular populations working in harmony to convert mechanical stimuli into electrical signals for the perception of sound. One such cell type are otic mesenchyme cells (OMCs), which are a specialized type of neural crest and cranial paraxial mesoderm that express multiple unique transcription factors (e.g., POU3F4), all of which are known deafness genes, highlighting the importance of OMCs in auditory function. OMCs are known to terminally differentiate into spatially and functionally distinct cell types, including fibrocytes of the lateral wall and spiral limbus, modiolar osteoblasts, and specialized tympanic border cells of the basilar membrane. Interestingly, consequences of Pou3f4 mutations are diverse and include a complete loss of endocochlear potential, shortening of the cochlear duct, and defective pathfinding and survival of spiral ganglion neurons (SGNs), indicating diverse roles of POU3F4 in each OMC-derived cell type. Here, we aim to illuminate the molecular distinctness and functionality of OMCs and show how loss of Pou3f4 impacts cochlear development. By utilizing scRNA sequencing, we elucidated that OMCs divide into four transcriptionally distinct subpopulations well before the onset of hearing, each of which corresponding to one of the OMC-derived cochlear structures. Furthermore, we show OMC subpopulations display distinct functional roles corresponding to their spatial localization. We also unravel the cochlear cellular communication pathways showcasing OMCs are the main contributors of outgoing signaling during cochlear development, including both global and subpopulation specifying signaling pathways. Finally, we indicate how Pou3f4 expression regulates gene expression in each OMC subpopulation and which signaling pathways are lost in Pou3f4 mutants which may be the cause of the defects in surrounding cell types. Our data suggest that OMC diversification occurs not long after the formation of the otocyst with further refinement until the onset of hearing, well before terminal differentiation. Our data also suggests OMCs are the main contributors of paracrine signaling during cochlear development, showcasing their importance in influencing surrounding cochlear cell types. Finally, we show how loss of Pou3f4 affects each OMC subpopulation differently, leading to diverse phenotypes in Pou3f4 mutants. Without cochlear OMCs and their later terminally differentiated cell types, normal auditory function would not be feasible highlighting the importance of tissue specific mesenchymal cells in cochlear development.