Unexpected role of <i>SIX1</i> variants in craniosynostosis: expanding the phenotype of <i>SIX1</i>-related disorders.

Abstract

Pathogenic heterozygous <i>SIX1</i> variants (predominantly missense) occur in branchio-otic syndrome (BOS), but an association with craniosynostosis has not been reported. We investigated probands with craniosynostosis of unknown cause using whole exome/genome (n=628) or RNA (n=386) sequencing, and performed targeted resequencing of <i>SIX1</i> in 615 additional patients. Expression of SIX1 protein in embryonic cranial sutures was examined in the <i>Six1</i>^{<i>nLacZ/+</i>} reporter mouse. From 1629 unrelated cases with craniosynostosis we identified seven different <i>SIX1</i> variants (three missense, including two de novo mutations, and four nonsense, one of which was also present in an affected twin). Compared with population data, enrichment of <i>SIX1</i> loss-of-function variants was highly significant (p=0.00003). All individuals with craniosynostosis had sagittal suture fusion; additionally four had bilambdoid synostosis. Associated BOS features were often attenuated; some carrier relatives appeared non-penetrant. SIX1 is expressed in a layer basal to the calvaria, likely corresponding to the dura mater, and in the mid-sagittal mesenchyme. Craniosynostosis is associated with heterozygous <i>SIX1</i> variants, with possible enrichment of loss-of-function variants compared with classical BOS. We recommend screening of <i>SIX1</i> in craniosynostosis, particularly when sagittal±lambdoid synostosis and/or any BOS phenotypes are present. These findings highlight the role of <i>SIX1</i> in cranial suture homeostasis.