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Title: The molecular basis of hereditary vitamin D resistant rickets
Author: Rut, Andrew
Awarding Body: University of London
Current Institution: University College London (University of London)
Date of Award: 1995
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Hereditary vitamin D resistant rickets is an autosomal recessive disorder characterised by severe rickets, hypocalcaemia, secondary hyperparathyroidism and occasionally, the absence of body hair. It has been shown that the pathological process involves resistance of target tissues to the actions of calcitriol [1,25(OH)2D3], the hormonal form of vitamin D. Calcitriol mediates its actions through a nuclear receptor (VDR) that has been cloned and shown to be a member of the superfamily of steroid/thyroid/retinoic acid receptors that act as gene trans-activating factors. These receptors are organised into two discrete functional domains, one for ligand binding at the COOH-terminus and one for DNA binding at the NH2-terminus. All nuclear hormone receptors have marked sequence conservation, particularly in the DNA binding domain. Crystallisation of this domain in the glucocorticoid receptor (GR) and has provided a structural model, enabling a stereochemical explanation of mutations in VDR that cause vitamin D resistant rickets. Five patients with target organ resistance to the actions of 1,25(OH)2D3 were studied. VDR from their skin fibroblasts were characterised. RNA was isolated, reverse transcribed, amplified, cloned and sequenced. Independent missense mutations were identified in four of the individuals, two localised to the hormone binding domain (Q149X & R271L) and two to the DNA binding domain (K42E & F44I). However, one patient with the classical phenotype had a normal cDNA sequence. The mutant receptors were expressed in VDR-deficient CV-1 cells and were found to demonstrate impaired trans-activation ability of an osteocalcin reporter gene linked to chloramphenicol transferase Finally, knowledge of the crystal structure of the DNA binding domain of GR, and its extensive homology with VDR, enabled computer-assisted modelling of the K42E & F44I mutations as well as five previously reported mutations. This provided an explanation for the defective phenotypes at the level of protein/DNA interactions.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available