Saethre-Chotzen syndrome (SCS), one of the most common forms of syndromic 
craniosynostosis (premature fusion of the cranial sutures), results from 
haploinsufficiency of TWIST1, caused by deletions of the entire gene or 
loss-of-function variants within the coding region. To determine whether 
non-coding variants also contribute to SCS, we screened 14 genetically 
undiagnosed SCS patients using targeted capture sequencing, and identified novel 
single nucleotide variants (SNVs) in the 5' untranslated region (UTR) of TWIST1 
in two unrelated SCS cases. We show experimentally that these variants, which 
create translation start sites in the TWIST1 leader sequence, reduce translation 
from the main open reading frame (mORF). This is the first demonstration that 
non-coding SNVs of TWIST1 can cause SCS, and highlights the importance of 
screening the 5' UTR in clinically diagnosed SCS patients without a coding 
mutation. Similar 5' UTR variants, particularly of haploinsufficient genes, may 
represent an under-ascertained cause of monogenic disease.