Dissecting the Role of the Six1‐Associated Genes in Craniofacial Development and Disease

Abstract

Genetic variants of SIX1, a transcription factor implicated in many cellular processes during craniofacial development, and of its co‐factor EYA1 underlie 50% of Branchio‐oto‐renal syndrome (BOR) cases. BOR is an autosomal dominant disorder characterized by various craniofacial defects, including malformed middle ear ossicles leading to conductive hearing loss. Interestingly, some BOR variants in SIX1 also underlie craniosynostosis (CRS) which indicates that SIX1 has a crucial role in craniofacial bone development. One important aspect of SIX1 function is that it interacts with co‐factors that can induce or repress its transcriptional activity. In this work, we expand our knowledge of the Six1 gene regulatory network with respect to craniofacial development. We first used a Six1 knockout mouse line to assess gene expression profiles of dissected E10.5 mandibular arches which give rise to the middle ear ossicles and lower jaw, via RNA‐sequencing. Our transcriptomic analysis led to the identification of 122 genes, including components of the FGF and WNT signaling pathways, known contributors to craniofacial bone development. In parallel, we used in vitro studies to characterize three novel co‐factors of Six1: Sobp, Mcrs1 and Pa2g4 that were recently identified in Xenopus. Our findings indicate that Sobp and Mcrs1 are bona fide Six1 co‐factors in mouse. We are now studying the genes identified in the RNA‐seq and the novel Six1 co‐factors using in vitro culture of mouse pre‐osteoblastic cells (MC3T3‐E1) to identify their role in bone formation. Altogether, our results identified novel Six1‐associated genes that are potential candidate genes for the unresolved cases of BOR and CRS.