Background: Homozygous loss-of-function mutations in the FOXE1 gene have been reported in several patients with partial or complete Bamforth syndrome (congenital hypothyroidism [CH] with thyroid dysgenesis [usually athyreosis], cleft palate, and spiky hair, with or without choanal atresia and bifid epiglottis). Here, our objective was to evaluate potential functional consequences of FOXE1 mutation in a patient with a similar clinical phenotype. Methods: FOXE1 was sequenced in 8 patients with thyroid dysgenesis. Transient transfection was performed in HEK293 cells using the thyroglobulin (TG) and thyroid peroxidase (TPO) promoters in the luciferase reporter plasmid to assess the functional impact of the FOXE1 mutations. Primary human thyrocytes transfected with wild-type and mutant FOXE1 served to assess the impact of the mutation on endogenous TG and TPO expression. Results: We identified and characterized the function of a new homozygous FOXE1 missense mutation (p.R73S) in a boy with a typical phenotype (athyreosis, cleft palate, and partial choanal atresia). This new mutation located within the forkhead domain was inherited from the heterozygous healthy consanguineous parents. In vitro functional studies in HEK293 cells showed that this mutant gene enhanced the activity of the TG and TPO gene promoters (1.5-fold and 1.7-fold, respectively vs. wild-type FOXE1, P<0.05), unlike the five mutations previously reported in Bamforth syndrome. The gain-of-function effect of the FOXE1-p.R73S mutant gene was confirmed by an increase in endogenous TG production by primary human thyrocytes. Conclusion: We identified a new homozygous FOXE1 mutation responsible for thyroid-function gene activation in a boy whose phenotype is similar to that reported previously in patients with loss-of-function FOXE1 mutations. This finding further delineates the role for FOXE1 in both thyroid and palate development and shows that enhanced gene activity should be considered among the mechanisms underlying Bamforth syndrome.