ATG5-FOXA3 Axis Contributes to Lysosomal Biogenesis and Auditory Function in Kölliker's Organ

Kölliker's organ (KO) support cells undergo orderly, time-dependent degeneration that is essential for auditory development and is accompanied by precisely regulated autophagic activity; however, the molecular hierarchy linking autophagy to this remodeling remains obscure. This study aimed to elucidate the regulatory mechanisms connecting autophagic flux to lysosomal biogenesis and auditory function during cochlear development.

We established an Atg5^flox/flox; Sox2Cre+ mouse model with deletion of the autophagy gene Atg5 in cochlear-supporting cells. Auditory function was assessed via Auditory Brainstem Response (ABR) testing. Transcriptomic profiling of the neonatal basilar membrane was performed to screen for downstream targets. Mechanistic validation included spatiotemporal immunofluorescence mapping (E18-P30) and in vitro functional assays using siRNA-mediated knockdown and lysosomal tracking.

At 2 months of age, Atg5^flox/flox; Sox2Cre+ mice exhibited moderate-to-severe sensorineural hearing loss accompanied by significant outer hair cell loss. Bulk RNA-seq of the basilar membrane identified fork-head box A3 (Foxa3) as a significantly downregulated transcription factor within the lysosomal-autophagy network. Spatiotemporal immunolabelling from embryonic day 18 to postnatal day 30 revealed that FOXA3 expression becomes progressively restricted to KO cells during postnatal development, with ATG5 loss reducing FOXA3 protein levels by 62.4%. In vitro, deficiency of either Atg5 or Foxa3 in primary KO cells resulted in comparable reductions in LAMP1-positive puncta.

These findings support a model wherein the ATG5-FOXA3 axis contributes to lysosomal biogenesis in developing KO cells, with implications for understanding mechanisms of congenital sensorineural hearing loss.