Direct targets of the transcription factor Six1 identify novel candidate deafness genes

Branchio-otic (BOS) and Branchio-oto-renal (BOR) syndromes are autosomal dominant disorders featuring multiple birth defects including ear, renal and branchial malformations. Mutations in the homeodomain transcription factor SIX1 and its co-factor EYA1 have been identified in about 50% of BOS/BOR patients, while causative mutations are unknown in the other half. We hypothesise that SIX1 target genes represent new BOS/BOR candidates. Using published transcriptomic and epigenomic data from chick ear progenitors, we first identify putative Six1 targets. Next, we provide evidence that Six1 directly regulates these targets: Six1 binds to their enhancers and functional experiments in Xenopus and chick confirm that Six1 controls their expression. Finally, we show that most chick Six1 targets are also expressed in the human developing ear and are associated with known deafness loci. Together, our results not only characterise the molecular mechanisms that mediate Six1 function in the developing ear, but also provide new candidates for human congenital deafness. Molecular characterisation of developing otic vesicle (OV) in humans is currently limited. We present the first transcriptome analysis of human embryonic OV from Carnegie stage (CS) 13 and CS14. OV (right and left), dorsal hind brain (DHB) and ventral hind brain (VHB) fresh tissues was dissected from a single embryo per replicate to perform bulk RNA seq. In total two bilogical replicated for each tissue was collected for both the stages respectively. To identify genes enriched in the OV at each of the developmnetal time points (CS13 &CS14) differential gene expression analysis using DESeq2 was performed comparing the bulk RNA seq data obtained from OV and DHB tissues from respective stages.