Endocrine Abstracts (2009) 19 P202

Characterisation of a novel GATA3 mutation in hypoparathyroidism, deafness, and renal dysplasia (HDR) syndrome

K Gaynor1, I Grigorieva1, M Nesbit1, T Cranston2, T Gomes3, L Gortner3 & R Thakker1


1Oxford Centre for Diabetes, Endocrinology and Metabolism, Churchill Hospital, University of Oxford, Oxford, UK; 2Genetics Laboratories, Churchill Hospital, University of Oxford, Oxford, UK; 3Abteilung für Allgemeine Pädiatrie und Neonatologie, Universitätskliniken des Saarlandes, Homburg/Saar, Germany.


GATA3 mutations cause the congenital autosomal dominant Hypoparathyroidism, Deafness and Renal dysplasia (HDR) syndrome. GATA3 belongs to a family of dual zinc-finger transcription factors that recognise the consensus (A/T)GATA(A/G) motif and are involved in vertebrate embryonic development. We investigated a HDR proband 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 sequence of both strands determined. This revealed a novel missense mutation, Thr272Ile, in zinc finger 1 (ZnF1) of GATA3. Electrophoretic mobility shift assays (EMSAs) revealed that the Thr272Ile mutant caused a reduction in DNA binding affinity, but had no effect on the stability of binding. Luciferase reporter assays established that Ile272 GATA3 activation of gene transcription was significantly reduced (P<0.01) compared to the wild-type protein. In addition, yeast two-hybrid assays demonstrated the mutation resulted in the loss of interaction with zinc finger 1 and 6 of FOG2, a known cofactor of GATA3. The effects of the Thr272Ile are intermediate to previously characterised ZnF1 mutations, which cause either a loss of interaction with FOG2 (Trp275Arg) or reduced stability of DNA binding (Arg276Pro). Three-dimensional modelling revealed that Thr272 lies between these two residues on the surface of the protein, accounting for the unique characteristic of affecting both DNA binding ability and interaction with the cofactor FOG2. The mutation results in a polar neutral, Thr, being substituted for a non-polar neutral, Ile. This subtle change in charge affects the protein–protein interactions between GATA3 and FOG2, with the change in electrostatic interactions affecting the specificity and strength of binding. These results elucidate further the molecular mechanisms of the altered function of this zinc finger transcription factor and its role in causing this developmental anomaly.

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