THE ROLE OF RECEPTORS IN THE ACTION OF 1,25-DIHYDROXYVITAMIN D3

摘要

The aim of this thesis has been to contribute to the further understanding of the mechanism of action of 1,25-dihydroxyvitamin D as a steroid hormone at both physiological and biochemical levels. The background section discusses the general mechanism of action, biochemical properties, and tissue distribution of 1,25-dihydroxyvitamin D receptors. This is followed by a description of experimental results which examined the areas of receptor regulation, receptor structure, and patients with vitamin D resistance due to receptor mutations.;The data on receptor regulation demonstrate that the number or abundance of 1,25-dihydroxyvitamin D receptors can be regulated by a variety of physiologically relevant manipulations. These studies were performed in a cultured kidney cell line as well as in vivo in mice and rats. Modification of tissue culture growth conditions and the effects of glucocorticoids, age, and estrogen were examined. The data indicate that receptor regulation may be an additional factor whereby functional response to hormone is modulated.;Receptor structure was examined by analysis of receptors after proteolysis by an endogenous serine protease found in intestine. Proteolysis resulted in the selective cleavage of the receptor DNA binding site leaving a receptor form with unique biochemical characteristics. Additional studies examined the "activation" process whereby receptors acquire the ability to bind to DNA. Unactivated or non-DNA binding receptor forms were isolated and activated in vitro by exposure to hypertonic salt conditions. The various receptor forms were further characterized by sucrose gradients and DNA-celulose binding. The data provide evidence for activation of the 1,25-dihydroxyvitamin D receptor and the presence of separate binding domains on the receptor for steroid and DNA.;Skin fibroblasts were cultured from biopsies of several individuals representing four kindreds with target organ resistance to vitamin D. Cells from affected children, unaffected siblings, and parents were cultured. Two categoreis of 1,25-dihydroxyvitamin D receptor defects were described; (i) defects in the steroid binding domain and (ii) defects in the DNA binding domain. Functional reponse to vitamin D metabolites revealed that cells with defective receptors did not respond to 1,25-dihydroxyvitamin D. Collectively, these studies aid in the elucidation of the receptor pathway, generate evidence for the postulated mechanism of action for 1,25-dihydroxyvitamin D, and provide a biochemical basis for the clinical findings in the syndrome of vitamin D resistance.