Unexpected role of SIX1 variants in craniosynostosis: expanding the phenotype of SIX1-related disorders.

The identification of disease-causing mutations has greatly benefited from the introduction of next-generation sequencing technologies. In particular, whole exome and genome sequencing (WES/WGS) have both accelerated the discovery of new genes involved in rare genetic diseases and, because of their unbiased nature, have enabled expansion of clinical phenotypes associated with known disease genes. Moreover retrospective clinical investigation of the patient and family members can reveal additional, previously unrecognised clinical features.1 The initial aim of this work was to investigate a possible genetic cause in a proband affected by speech and language delay, sensorineural hearing loss (HL) and craniosynostosis (CRS), the premature fusion of one or more cranial sutures of the skull. We identified a de novo mutation in SIX1 , which encodes a homeodomain-containing transcription factor of the sine oculis class, originally described in Drosophila .2 Further analysis of additional unsolved CRS cases using WES/WGS, RNA sequencing or targeted resequencing identified heterozygous variants in SIX1 in seven further patients with CRS from six unrelated families. Dominantly inherited SIX1 variants were previously reported in branchio-otic syndrome (BOS; MIM 608389),3 4 non-syndromic HL (MIM 605192)5 and (rarely) in branchio-oto-renal (BOR) syndrome, associated in addition with renal malformation.3 This work uncovers a previously unrecognised role of SIX1 in the maintenance of cranial suture patency. ### Patients Written informed consent was obtained from all participants/legal guardians. The clinical diagnosis of CRS was confirmed by three-dimensional CT scanning of the skull. When clinically indicated, samples were tested for mutation hotspots in FGFR2 , FGFR3 , TWIST1 , TCF12 and ERF , and significant chromosome aneuploidy was investigated using array comparative genomic hybridisation; samples harbouring mutations known to cause CRS were excluded. ### Genetic analyses WES or WGS of unrelated probands with CRS of unknown cause (n=103 and n=525 in Oxford and Yale cohorts, respectively) and subsequent …