Differential expression of mechanotransduction complex genes in auditory/vestibular hair cells in zebrafish.

Ciliated sensory cells such as photo- and olfactory receptors employ multiple types of opsins or hundreds of unique olfactory G-protein coupled receptors to respond to various wavelengths of light or odorants. With respect to hearing and balance, the mechanotransduction machinery involves fewer variants; however, emerging evidence suggests that specialization occurs at the molecular level. To address how the mechanotransduction complex varies in the inner ear, we characterized the expression of paralogous genes that encode components required for mechanotransduction in zebrafish hair cells using RNA-FISH and bioinformatic analysis. Our data indicate striking zonal differences in the expression of two components of the mechanotransduction complex which are known to physically interact, the transmembrane channel-like 1 and 2 (tmc1/2) family members and the calcium and integrin binding 2 and 3 (cib2/3) paralogues. tmc1, tmc2b, and cib3 are largely expressed in peripheral or extrastriolar hair cells, whereas tmc2a and cib2 are enriched in central or striolar hair cells. In addition, a gene implicated in deaf-blindness, ush1c, is highly enriched in a subset of extrastriolar hair cells. These results indicate that specific combinations of these components may optimize responses to mechanical stimuli in subtypes of sensory receptors within the inner ear.

Epimorphic regeneration in the mammalian tympanic membrane.

Adult mammals are generally believed to have limited ability to regenerate complex tissues and instead, repair wounds by forming scars. In humans and across mammalian species, the tympanic membrane (TM) rapidly repairs perforations without intervention. Using mouse models, we demonstrate that the TM repairs itself through a process that bears many hallmarks of epimorphic regeneration rather than typical wound healing. Following injury, the TM forms a wound epidermis characterized by EGFR ligand expression and signaling. After the expansion of the wound epidermis that emerges from known stem cell regions of the TM, a multi-lineage blastema-like cellular mass is recruited. After two weeks, the tissue architecture of the TM is largely restored, but with disorganized collagen. In the months that follow, the organized and patterned collagen framework of the TM is restored resulting in scar-free repair. Finally, we demonstrate that deletion of Egfr in the epidermis results in failure to expand the wound epidermis, recruit the blastema-like cells, and regenerate normal TM structure. This work establishes the TM as a model of mammalian complex tissue regeneration.

A Hierarchy of Proliferative and Migratory Keratinocytes Maintains the Tympanic Membrane.

The tympanic membrane (TM) is critical for hearing and requires continuous clearing of cellular debris, but little is known about homeostatic mechanisms in the TM epidermis. Using single-cell RNA sequencing, lineage tracing, whole-organ explant, and live-cell imaging, we show that homeostatic TM epidermis is distinct from other epidermal sites and has discrete proliferative zones with a three-dimensional hierarchy of multiple keratinocyte populations. TM stem cells reside in a discrete location of the superior TM and generate long-lived clones and committed progenitors (CPs). CP clones exhibit lateral migration, and their proliferative capacity is supported by Pdgfra+ fibroblasts, generating migratory but non-proliferative progeny. Single-cell sequencing of the human TM revealed similar cell types and transcriptional programming. Thus, during homeostasis, TM keratinocytes transit through a proliferative CP state and exhibit directional lateral migration. This work forms a foundation for understanding TM disorders and modeling keratinocyte biology.