EDNRB2 regulates fate determination, migration and maturation of hair-cell precursors in regenerating avian auditory epithelia
The mammalian cochlea loses the ability for hair cell (HC) regeneration after birth, while in the avian auditory epithelium, namely the basilar papilla (BP), supporting cells (SCs) retain the capability for HC regeneration throughout life. Our previous study using single-cell RNA sequencing indicated stepwise fate conversion of SCs to HCs via the precursor state, in which EDNRB2 exhibited specifically high expression, in the process of HC regeneration in chick BP. The present study aimed to reveal a distinct role of EDNRB2 in HC regeneration in chick BP. During HC regeneration in chick BP explant cultures, EDNRB2 expression emerged in a part of ATOH1-expressing SCs immediately after HC loss and decreased according to the progress of HC regeneration process. In embryonic chick BP, EDNRB2 expression was specifically identified in the precursor cell state, which confirmed that mature SCs were reprogrammed to the precursor state in response to HC loss. Pharmacological inhibition of endothelin receptor type B (EDNRB) signaling decreased regenerated HCs in chick BP explant cultures. RNA sequencing revealed that inhibition of EDNRB signaling downregulated genes for HC differentiation and maturation together with genes for cell migration. Histological assessments clarified the deterioration of migration of HC precursors and the delay of HC regeneration processes by inhibition of EDNRB signaling. These results indicate that the expression of EDNRB2 contributes to the fate determination of HCs and that EDNRB signaling regulates the migration and maturation of HC precursors during chick HC regeneration. Overall design: Bulk RNA-seq of chick basilar papaillae during hair cell regeneration using an explant culture