Endocrine Abstracts

Mutations leading to haploinsufficiency of the dual zinc finger transcription factor GATA3 result in the hypoparathyroidism-deafness-renal dysplasia (HDR) syndrome which is an autosomal dominant disorder. We investigated 21 HDR probands and 14 patients with isolated hypoparathyroidism for GATA3 abnormalities. Venous blood was obtained after informed consent, as approved by the local ethical committee, and leukocyte DNA extracted. GATA3 specific primers were used for PCR amplification and the DNA sequences of both strands determined. Thirteen different heterozygous germline mutations were identified in patients with HDR. These consisted of 6 frameshifting deletions, 2 frameshifting insertions, 3 nonsense mutations, one missense (Leu348Arg) mutation, and one acceptor splice site mutation. The splice site mutation was demonstrated to cause a pre-mRNA processing abnormality leading to the use of an alternative acceptor site 8-bp downstream of the normal site, resulting in a frameshift and prematurely terminated protein. Electrophoretic mobility shift assays (EMSAs) revealed three classes of GATA3 mutations: those that lead to a loss of DNA binding which represent over 90% of all mutations, and involved a loss of the carboxy-terminal zinc finger; those that resulted in a reduced DNA binding affinity; and those (e.g., Leu348Arg) that did not alter DNA binding or the affinity but likely altered the conformational change that occurs during binding in the DNA major groove as predicted by three-dimensional modelling. These results elucidate further the molecular mechanisms underlying the altered functions of mutants of this zinc finger transcription factor and their role in causing this developmental anomaly. No mutations were identified in patients with isolated hypoparathyroidism. Thus, our analysis indicates that in the clinical setting a search for GATA3 abnormalities is likely to be productive in patients with either 3 or 2 of the phenotypic features of the HDR syndrome, but not in those with only one, such as isolated hypoparathyroidism.