Mutations in the NKX2.5 gene and the PAX8 promoter in a girl with thyroid dysgenesis.

Context: Screening of the known candidate genes involved in thyroid organogenesis has revealed mutations in a small subset of patients with congenital hypothyroidism due to thyroid dysgenesis (TD).

Objective: We studied a girl with TD who had mutations in two transcription factors involved in thyroid development.

Results: Sequencing analysis of candidate genes involved in thyroid gland development revealed a new paternally inherited heterozygous mutation in the NKX2.5 gene (S265R) and a new maternally inherited heterozygous mutation in the PAX8 promoter region (-456C>T). Both parents and a brother, who was also heterozygous for both mutations, were phenotypically normal. Immunofluorescence microscopy showed a correct nuclear localization of both wild-type (WT) and mutant NKX2.5 proteins. EMSA demonstrated that the mutant NKX2.5 binds to the NKE_2, DIO2, TG, and TPO promoter elements equally well as the WT protein. However, the mutant NKX2.5 protein showed a 30-40% reduced transactivation of the thyroglobulin and the thyroid peroxidase promoters and a dominant-negative effect of the mutant NKX2.5. EMSA studies of the WT and mutant PAX8 promoter sequences incubated with nuclear extracts from PCCL3 cells exhibited a loss of protein binding capacity of the mutant promoter. In addition, the mutant PAX8 promoter showed a significantly reduced transcriptional activation of a luciferase reporter gene in vitro. Thus, this promoter mutation is expected to lead to reduced PAX8 expression.

Conclusions: We identified new heterozygous mutations in both NKX2.5 and PAX8 genes of a girl with TD. Both defects might contribute to the phenotype.