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The inner ear is a complex structure responsible for hearing and balance, and organ pathology is associated with deafness and balance disorders. To evaluate the role of epigenomic dynamics, we performed whole genome bisulfite sequencing at key time points during the development and maturation of the mouse inner ear sensory epithelium (SE). Our single-nucleotide resolution maps revealed variations in both general characteristics and dynamics of DNA methylation over time. This allowed us to predict the location of non-coding regulatory regions and to identify several novel candidate regulatory factors, such as Bach2, that connect stage-specific regulatory elements to molecular features that drive the development and maturation of the SE. Constructing in silico regulatory networks around sites of differential methylation enabled us to link key inner ear regulators, such as Atoh1 and Stat3, to pathways responsible for cell lineage determination and maturation, such as the Notch pathway. We also discovered that a putative enhancer, defined as a low methylated region (LMR), can upregulate the GJB6 gene and a neighboring non-coding RNA. The study of inner ear SE methylomes revealed novel regulatory regions in the hearing organ, which may improve diagnostic capabilities, and has the potential to guide the development of therapeutics for hearing loss by providing multiple intervention points for manipulation of the auditory system.

BackgroundThe mammalian inner ear is a complex morphological structure responsible for hearing and balance, and its pathology is associated with deafness and balance disorders. To evaluate the role of epigenomic dynamics in the development and maturation of mouse inner ear sensory epithelium, we performed whole-genome bisulfite sequencing on inner ear tissue, yielding temporal base-pair resolution methylomes at key developmental time points.ResultsWe found a late accumulation of non-CpG methylation, indicating a similarity between the inner ear sensory epithelium and neuronal tissue. Moreover, annotation of both unmethylated and low methylated regions pointed to regulatory elements active in the inner ear in proximity of and distal from transcriptional units. Finally, we identified differentially methylated regions across the transition periods. An analysis of these regions led us to identify several novel candidate regulatory factors, connecting regulatory elements from specific time points in development to molecular features that drive the development and maturation of the inner ear sensory epithelium. The GJB6 locus putative regulatory region was shown to upregulate distal GJB6 gene expression and a non-coding RNA.ConclusionsOur analysis of inner ear sensory epithelium DNA methylation sheds light on novel regulatory regions in the hearing organ, and may help boost diagnostic capabilities and guide the development of therapeutics for hearing loss, by providing multiple intervention points for manipulation of the auditory system.